2023年度中國肉牛產業發展大會-夷陵牛產業發展模式白皮書（97頁）.pdf

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1、白白皮皮書書發發布布單單位位：國國家家肉肉牛牛牦牦牛牛產產業業技技術術體體系系宜宜昌昌市市人人民民政政府府枝枝江江市市人人民民政政府府20232023年度中國肉牛產業發展大會年度中國肉牛產業發展大會夷陵牛產業發展模式夷陵牛產業發展模式目目錄錄1.夷陵牛產業發展模式之一政策模式012.夷陵牛產業發展模式之二技術模式143.夷陵牛產業發展模式之三商業模式234.附件：夷陵牛部分研究成果4.1.夷陵黃牛品種資源調查報告(一)夷陵黃牛形成過程的考證354.2.夷陵黃牛品種資源調查報告(二)夷陵黃牛的產地與分布情況394.3.夷陵黃牛品種資源調查報告(三)夷陵黃牛肉用選育及產業化開發思路414.4.夷陵

2、黃牛的保種及雜交改良利用444.5.基于全長轉錄組測序的牛脂肪組織可變剪接事件分析_夏晗454.6.夷陵黃牛 mtDNA+D-loop 區遺傳多樣性研究534.7.Genetic background analysis and breed evaluation of Yiling yellowCattle574.8.Effects of Age and Rice Straw Inclusion Levels in the Diet ofYiling Cull Cows on Growth Performance,Meat Quality,andAntioxidant Status of Tis

3、sues684.9.Serum Biochemical Parameters,Rumen Fermentation,and RumenBacterial Communities Are Partly Driven by the Breed and Sex ofCattle When Fed High-Grain Diet82夷陵牛產業發展模式之“政策模式”余峰枝江市人民政府近年來，在省委省政府和宜昌市委市政府的堅強領導下，在國家肉牛牦牛產業技術體系的大力支持下，枝江市深入學習貫徹習近平總書記關于“三農”工作的重要論述，始終把肉牛產業作為農業主導產業來抓，以用促保強種業、綠色托底優養殖、聯農帶農

4、保增收，把政策項目落實到產業鏈條上，把科技研究書寫在生產發展上，把豐收碩果結在廣大農戶中，奮力譜寫枝江夷陵牛產業發展新篇章。一一、手執手執“牛耳朵牛耳朵”，以用促保踐行種業安全以用促保踐行種業安全，推進種推進種源保護開發源保護開發夷陵牛是一頭血統純正的中國夷陵牛是一頭血統純正的中國“老老”黃牛黃牛。農業種質資源是農業的“芯片”。習近平總書記強調，農業現代化，種子是基礎，必須把民族種業搞上去，實現種業科技自立自強、種源自主可控。夷陵牛種質資源保護開發，是深入貫徹落實習近平總書記重要講話精神、扎實做好“土特產”文章的生動實踐，是“執牛耳”的大事。一是從古到今一是從古到今，挖掘夷陵牛的前世今生挖掘夷

5、陵牛的前世今生。宜昌古稱夷陵，所以，本地的黃牛就有了“夷陵?！边@個古韻悠長的名字。相傳大禹治水，夷陵牛就是開山牛。據夷陵州志記載，宜昌農戶養殖夷陵牛已有千百年的歷史。2018 年 1 月，夷陵-1-牛被農業農村部列入國家畜禽遺傳資源目錄，正式成為我國黃牛的一個新品種。圖圖 1 1專家團隊在峽江沿岸探尋夷陵牛歷史專家團隊在峽江沿岸探尋夷陵牛歷史二是從點到面二是從點到面，做好夷陵牛種源保護做好夷陵牛種源保護。為了保護好夷陵牛的種質資源，我們籌措農業發展項目資金 3000 多萬元，建立夷陵牛核心保種場及良種繁育基地，核心群種牛規模達到 400 頭以上。同時，推廣“公司+村集體+農戶”的夷陵牛代養模式

6、，建立“農戶代養母牛、公司集中育肥”的良性循環機制，實現基礎母牛擴繁增量目標。目前，全市發展代養示范村 15 個、代養農戶 183 戶，代養夷陵母牛 550 頭。-2-圖圖 2 2宜昌市第二屆夷陵牛評鑒會宜昌市第二屆夷陵牛評鑒會三是從有到優三是從有到優，推動夷陵牛開發利用推動夷陵牛開發利用。我們堅持“以保為先、以保促用、保用結合”，在國家肉牛牦牛產業技術體系專家的悉心指導下，利用夷陵牛成功培育出 A3 級以上雪花牛肉，牛肉價格從 100 元/公斤提高到 1200 元/公斤，每頭牛價值從 1.5 萬元提高到 8 萬元，將資源優勢轉化為產業優勢、經濟優勢。圖圖 3 3培育夷陵牛雪花牛肉培育夷陵牛雪

7、花牛肉-3-二二、抓住抓住“牛鼻子牛鼻子”，積極引進行業頂尖力量積極引進行業頂尖力量，科技賦科技賦能產業發展能產業發展夷陵牛是一頭技術頂尖的高端科技牛夷陵牛是一頭技術頂尖的高端科技牛。我們堅持把科技興農作為推動農業高質量發展的重要路徑，緊緊扭住科技創新這個“牛鼻子”，加快推進農業科技成果轉化，大力推廣新品種、新技術、新模式。一是集聚產業頂尖人才一是集聚產業頂尖人才?！叭櫭]”求賢引才，2015年以來多次赴漢進京，拜訪國家肉牛牦牛產業技術體系首席科學家曹兵海和李俊雅、孫寶忠、郭愛珍等崗位科學家，為夷陵牛產業發展把脈問診。目前，曹兵海首席科學家工作站落戶枝江，專注于夷陵牛資源保護與雪花牛肉開發。

8、圖圖 4 4專家指導夷陵牛雪花牛肉培育專家指導夷陵牛雪花牛肉培育-4-二是搭建科技創新平臺二是搭建科技創新平臺。與中國農業大學、華中農業大學、湖北省農科院等科研機構展開務實合作，共同開展農業技術攻關和成果轉化，建立產學研示范基地，解決了飼料調配與飼養管理、養殖糞污資源化利用、動物疫病綜合控制與凈化、種養循環一體化等技術難點。三是開展技術推廣應用三是開展技術推廣應用。結合養殖戶實際需求，積極開展動物疫病防控、床場一體化養殖等主推技術培訓，每年技術培訓達 2400 多人次，全面提升肉牛養殖技術水平，進一步降低養殖風險、提高養殖效益，疫病死亡率由 10下降到1.5，日增重由 1.25 公斤提高到 1

9、.4 公斤，出欄重由 650公斤提高到 700 公斤，規模養殖場床場一體化養殖覆蓋率達到 100%。圖圖 5 5開展養殖技術培訓開展養殖技術培訓-5-三三、壯實壯實“大牛腿大牛腿”，深入推進投入機制創新深入推進投入機制創新，夯實產夯實產業基礎支撐業基礎支撐夷陵牛是一頭四肢強健的開拓創新牛夷陵牛是一頭四肢強健的開拓創新牛。我們堅持“政府主導、部門協同、社會參與、市場驅動”的原則，創新肉牛產業發展多元化投入模式，形成財政、金融、土地、品牌“四大支撐”，推動夷陵牛產業高質量發展。一是強化財政撬動一是強化財政撬動。2015 年以來，我們深入落實各級關于推進牛羊產業發展的意見精神，制定肉牛產業發展實施方

10、案，發揮財政資金“四兩撥千斤”的作用，每年安排財政資金 200 萬元以上，對牛舍建設、夷陵牛保種、品牌創建、貸款貼息、社會化服務、疫病防控等方面進行獎補，引導帶動廣大養殖戶科學發展肉牛產業。二是強化金融創新二是強化金融創新。搶抓全省首個農村合作金融創新試點機遇，成功探索“農合聯+農擔+銀行+農業獎補+農業保險+政務平臺”的政擔銀企多方合作、風險共擔機制，以金融“活水”激發“三農”活力。2022 年，全省金融服務鄉村振興共建創新示范區現場推進會在枝江召開，“枝江模式”全省推廣。成立 500 萬元肉牛產業發展擔?；?，按 1:8 的比例放大，為肉牛養殖戶提供無抵押貼息貸款，2022 年發放肉牛貼息

11、貸款 5000 余萬元。推出肉牛政策性保險，保險費率3.5%，保費補貼 60%，進一步降低了養殖風險，解決了后顧之憂。-6-圖圖 6 6“犇牛貸犇牛貸”發放儀式發放儀式三是強化用地保障三是強化用地保障。編制地方國土空間規劃時，統籌解決肉牛養殖用地需求。實行肉牛產業發展用地限時辦結，對符合畜禽養殖“三區”規劃的肉牛養殖用地，統一辦理設施農業用地，占用林地的，依法依規辦理使用林地手續。嚴格落實環境保護“三同時”要求，提升肉牛養殖項目環評審批質效。四是強化品牌賦能四是強化品牌賦能。成功打造湖北省首個縣域農產品公共品牌“枝滋有味”，支持“牛味央”、“仙女牛郎山”肉牛品牌融入“枝滋有味”品牌建設，形成“

12、公共品牌+行業品牌+自主品牌”的農業品牌體系。夷陵牛品牌在央視種子種子和科技苑等欄目宣傳推介，品牌使用龍頭企業達到 7 家，品牌產品銷售額達到 2 億元；夷陵牛雪花牛肉獲評首屆中國牛優質牛肉品鑒大會“最具特色獎”和“最具效益獎”，逐步實現從“賣資源”向“賣品牌”轉變。-7-圖圖 7 7 夷陵牛雪花牛肉獲評首屆中國牛夷陵牛雪花牛肉獲評首屆中國牛 優質牛肉品鑒大會優質牛肉品鑒大會“最具特色獎最具特色獎”和和“最具效益獎最具效益獎”四四、勇鉆勇鉆“牛角尖牛角尖”，延鏈強鏈補齊發展短板延鏈強鏈補齊發展短板，促進產促進產業深度融合業深度融合夷陵牛是一頭延鏈強鏈的產業拓荒牛。夷陵牛是一頭延鏈強鏈的產業拓荒

13、牛。我們牢固樹立“鏈式發展”思維，以“鉆牛角尖”的執著精神，深入推進一二三產業融合發展，加快打造肉牛產業發展全產業鏈，推動夷陵牛產業“?！眲葆绕?。一是聚焦一是聚焦“一產一產”，夯實產業基礎夯實產業基礎。大力推廣肉牛養殖“3321”模式（即一個養殖戶新建標準化牛舍 300 平方米、每年飼養牛 30 頭、利用秸稈 200 噸、年收入 10 萬元），已發展肉牛養殖示范村 12 個、示范戶 129 個、標準化牛舍 269棟，戶均增收 13.5 萬元。2022 年，全市肉牛規模養殖場達到 43 個，肉牛存欄 2.86 萬頭，出欄 1.45 萬頭，每年正以-8-20%以上的速度增長，肉牛產業已成為枝江市鄉

14、村振興、農民增收的重要產業。圖圖 8 8“33213321”肉牛養殖模式示范戶肉牛養殖模式示范戶二是二是對接對接“二產二產”，發展精深加工。，發展精深加工。按照“集中屠宰、冷鏈配送、生鮮上市”的要求，支持龍頭企業建立肉牛自動化屠宰分割生產線。高標準建成牛肉精深加工車間，生產牛肉干、牛肉包、牛肉醬等各類產品，滿足市場多樣化、品質化需求。同時，搶抓預制菜產業發展風口，推出古法牛肉、金湯牛仔骨等熱銷產品。2022年，全市精深加工牛肉產品產量達30余噸。圖圖 9 9 牛肉精深加工系列產品牛肉精深加工系列產品-9-三是三是緊連緊連“三產三產”，推進推進牧牧旅融合旅融合。搶抓國家級美麗鄉村示范村建設機遇，

15、瞄準創建國家4A級旅游景區，加快推進仙女向巷村牛郎山小鎮片區建設，優化完善賽牛場、觀光牧場、林間步道和吉吉村桔子集市、桔子工坊、桔子餐廳、桔子樂園等基礎設施，每年接待游客5萬人次以上，逐步實現美麗鄉村向美麗經濟轉化。圖圖 1010舉行舉行“牛的盛宴牛的盛宴“主題活動主題活動五五、騎上騎上“大牛背大牛背”，聯農帶農助力鄉村振興聯農帶農助力鄉村振興，推動實推動實現共同富?，F共同富裕夷陵牛是一頭扎根鄉土的發展致富牛夷陵牛是一頭扎根鄉土的發展致富牛。我們始終堅持生活、生產、生態“三生融合”發展理念，以肉牛產業高質量發展為突破口，優化聯農帶農機制，全面推進鄉村振興，讓更多農戶騎上“大牛背”，共享高標準生

16、產、高顏值生態、高品質生活。一是推動高標準生產一是推動高標準生產。探索建立“龍頭企業+合作社+示-10-范戶+農戶”聯農帶農機制，依托省級龍頭企業豐聯佳沃，成功打造省級肉牛產業化聯合體，推行統一種源管理、統一飼料供應、統一技術服務、統一銷售渠道“四統一”的服務模式，幫助養殖戶解決用地、資金、技術、銷售等難題，讓更多農戶深度融入產業鏈、供應鏈、價值鏈，讓他們有錢掙、得實惠、能致富，推動共同富裕邁出堅實步伐。2022 年，全市農村居民人均可支配收入達到 29204 元、居全省第二，城鄉居民收入之比縮小至 1.482，連續 3 年居全省同類縣市區第一。圖圖 1111 合作社開展送牛服務合作社開展送牛

17、服務二是打造高顏值生態二是打造高顏值生態。以深入實施農村人居環境整治提升五年行動為契機，整市推進肉牛床場一體化技術，肉牛養殖糞污資源化利用率達 98%以上，有效解決了肉牛養殖戶糞污處理難、亂堆亂放造成的環境“臟亂差”問題。同時，加快推進秸稈飼料化利用，既避免了焚燒秸稈帶來的環境污染，-11-又降低了肉牛養殖成本，實現了經濟效益和生態效益“雙提升”。枝江市獲評全國美好環境與幸福生活共同締造試點縣、全國村莊清潔行動先進縣，長江枝江段水質穩定達到類，飲用水水源地水質達標率 100%，2022 年空氣質量優良天數比率達到 89.8%，位居全省一類縣市區第二。三是創造高品質生活三是創造高品質生活。通過大

18、力發展夷陵牛產業，不僅富了廣大養殖戶的“口袋”，還壯大了村級集體經濟。2022年，全市村級集體經濟總收入 4590 萬元，村平經營性收入17.55 萬元，176 個村成功實現債務清零。隨著集體經濟的發展壯大，村級更有能力投入鄉村建設，推動農村基礎設施和公共服務提檔升級，實現村民購物、農產品銷售、看病、金融服務、政務服務“五個不出村”。同時，進一步突出群眾主體地位，廣泛開展美好環境與幸福生活共同締造，著力打造美麗花園、宜居公園、生態田園、富足樂園、精神家園“五園”，構建生活共同體、利益共同體、情感共同體、責任共同體、治理共同體、精神共同體“六個共同體”，共同締造幸福美好生活。圖圖 1212 扶貧

19、牛車間扶貧牛車間-12-下一步，枝江市將深入貫徹落實習近平總書記關于“三農”工作的重要論述，全面落實本次大會精神，聚焦夷陵牛產業發展，著力打造集飼料生產、綠色養殖、精深加工、品牌銷售于一體的肉牛全產業鏈，進一步叫響“夷陵?！眳^域公用品牌，讓“夷陵?！背蔀橹斎什蛔尩摹包S金名片”！-13-夷陵牛產業發展模式之“技術模式”曹兵海李俊雅 孫寶忠劉繼軍 郭愛珍國家肉牛牦牛產業技術體系地方黃牛品種資源是祖先留下的財富還是負擔？是“保著、供著”還是合理開發利用？如何高效地開發利用？這些問題一直是困擾養牛業的重要問題，既是技術問題，也是產業政策問題。夷陵牛產業在較短時間內摸索了一條行之有效的產業發展模式，

20、包括產業政策模式、技術模式和商業模式，三者相輔相成，初步實現了夷陵?！耙杂么！比a業鏈發展。本文整理了夷陵牛產業發展技術模式，包括慧眼視?；蜩b寶定抓手、專家合議高端定位繪藍圖、技術攻關破瓶頸（精準飼喂高檔育肥、雜交改良提檔升級、精深加工增值增效）、苦練內功集成創新鑄造核心競爭力、疾病凈化健康護航等五個方面，期望為我國地方黃牛品種的有效保護和利用提供借鑒和參考。一、慧眼視牛，基因鑒寶，錨定抓手一、慧眼視牛，基因鑒寶，錨定抓手2015年9月27-29日，國家肉牛牦牛產業技術體系首席、中國農業大學曹兵海教授，疾病防治研究室主任、華中農業大學郭愛珍教授一行，和當時宜昌市畜牧獸醫局沈洪學局長、宜昌市

21、畜牧獸醫推廣站和枝江市畜牧獸醫局的相關領導一道，行走于“黃牛山”腳下的峽江兩岸，一邊聽著“神牛助禹開三峽”、“屈鄉靈?！钡墓适?，一邊尋找屬于這塊神奇土地的黃牛的故事。為了應證傳說中的真實，沿途拜訪了黃-14-牛廟，親睹了蜀相諸葛亮所作黃牛廟記的碑刻。專家們或走家串戶，了解養牛戶的飼養情況；或草坡駐足，審視神牛故鄉放牧牛群的神韻。終于，在瑪瑙河畔的河灘上，專家們發現一群長相顯眼的本地黃牛（圖 1），審視再三，喜不自禁。在曹兵海首席建議下，將其命名為：“夷陵黃?！?，以宜昌古稱“夷陵”冠名。圖圖 1A1A瑪瑙河畔的河灘上放牧的本地黃?，旇Ш优系暮訛┥戏拍恋谋镜攸S牛圖圖 1B1B專家們專注觀察瑪瑙河畔

22、河灘上放牧的本地黃牛專家們專注觀察瑪瑙河畔河灘上放牧的本地黃牛-15-2016 年 3 月 30 至 4 月 2 日，曹兵海首席、郭愛珍教授、體系遺傳育種研究室主任、中國農業科學院畜牧獸醫研究所李俊雅研究員，環境設施研究室主任、中國農業大學劉繼軍教授，加工研究室專家、中國農業科學院畜牧獸醫研究所孫寶忠研究員等 5 位體系專家和宜昌市畜牧獸醫局、宜昌市畜牧獸醫推廣站和枝江市畜牧獸醫局等相關領導和企業負責人一道，再訪夷陵黃牛。初步確定了夷陵黃牛的體型外貌特征，主要為：體型中等，外貌基本一致，毛色以棕黃色、板栗色、黑色為主，公母牛均有角，主要特征為“兩黑一筍”，即眼圈黑、鼻鏡黑、筍角；頭清秀且稍長，

23、腰背平直，腹部圓大，尻下斜，尾帚大過飛節；公牛頭頸粗短，肩峰較高；母牛頭頸細長，肩峰較小。曹首席暗許承諾，要為現代神牛續寫“大文章”！2016 年 7 月，李俊雅研究員團隊來湖北采樣，擬為夷陵牛進行基因鑒定。選取了 47 頭夷陵黃牛、19 頭鄖巴黃牛、34 頭恩施黃牛、36 頭棗北黃牛、21 頭秦川牛等品種牛進行DNA 分析。通過 GeneSeek Genomic Bovine LD(GGP-LD)芯片平臺，對樣本進行基因分型。10 月發布研究結果，為夷陵黃牛驗明正身：從遺傳背景角度，夷陵黃牛明顯區別于周邊其他品種，已經形成了自身特有的遺傳特征，是我國牛品種大家庭中不可缺少的成員（圖 2）。后

24、來，體系專家雷朝初教授證實，夷陵黃牛 mtDNA 遺傳多樣性比較豐富，具有普通牛和瘤牛兩大母系起源，屬南方本地黃牛。-16-圖圖 2 2NJNJ 進化樹（進化樹（154154 個個體）。紅色分支表示夷陵黃牛個個體）。紅色分支表示夷陵黃牛二、專家合議，高端定位，揮筆藍圖二、專家合議，高端定位，揮筆藍圖面對地方品種保種難的世紀難題，專家們充分肯定了夷陵黃牛品種的不可替代性，從品種資源調查和特色挖掘、品種選育和保護、精準飼養和高檔育肥、環境控制、疫病防控、屠宰和精深加工、餐飲體驗店等方面開啟了“以用代?！比a業鏈技術模式。宜昌市畜牧獸醫局設專項支持專家的科技研究。圖圖 3 3體系專家們為夷陵黃牛的保

25、種和利用進行整體策劃體系專家們為夷陵黃牛的保種和利用進行整體策劃-17-2017 年 1 月 15 日，農業農村部組織國家畜禽遺傳資源委員會牛馬駝專業委員會專家組對夷陵黃牛進行現場鑒定，專家建議更名為“夷陵?！?。11 月 22 日，夷陵牛通過了國家畜禽遺傳資源委員會的專家鑒定，確定為中國肉牛地方遺傳資源。2018 年 1 月 8 日，農業農村部第 2637 號公告公布，夷陵牛為我國第 55 個地方黃牛品種！進一步依托省級農業產業化龍頭企業-湖北豐聯佳沃農業開發有限公司建設了宜昌市級夷陵牛核心品種資源場，在主產區選購優質種公牛和種母牛，開展品種選育。劉繼軍教授設計了保種場牛舍，規劃和優化了養殖環

26、境條件。李俊雅研究員牽頭制定了夷陵黃牛地方品種標準，指導師完成了資源普查，建立了保種場和保種區，完善了系譜和技術管理檔案，并建立了優質夷陵牛的凍精凍胚基因庫。三、問診把脈，技術攻關，突破瓶頸三、問診把脈，技術攻關，突破瓶頸（一）精準飼喂，高檔育肥（一）精準飼喂，高檔育肥地方黃牛長得慢、產肉少、效益低是保種難的卡脖子問題，因此提高養殖效益是有效保種必需攻克的關鍵技術問題。曹兵海首席高遠立意、高端布局，率團研究夷陵牛的代謝和生長發育特點，通過按階段精準飼喂和營養動態調控技術實現高檔育肥（圖 4）；同時，通過夷陵?；铙w評價、屠宰性能測定、胴體評級、分割肉質測定等研究，指導企業構建了夷陵牛高檔育肥、屠

27、宰、成熟、分割與包裝技術體系。2017年 1 月 16 日，夷陵牛雪花牛肉新品發布會成功召開，夷陵牛生產的高檔雪花牛肉閃亮登場！-18-圖圖 4 4曹兵海首席指導夷陵牛高檔育肥、生產雪花牛肉曹兵海首席指導夷陵牛高檔育肥、生產雪花牛肉（二）（二）雜交改良，提檔升級雜交改良，提檔升級進一步提高養殖效益是推動夷陵?！耙杂么！辈呗缘年P鍵。在本品種擴群選育基礎上，曹兵海首席探討了和牛和夷陵牛、安格斯牛和夷陵牛的二元雜交研究。雜交改良的 24月齡公牛體重由 800 斤增加到 950 斤，母牛由 650 斤增加到800 斤；30 月齡高檔育肥公牛體重由 1000 斤增加到 1250 斤，生產 A3 級雪花

28、肉量由 40 斤增加到 80 斤，大大提高了生產效益（圖 5）。圖圖 5 5曹兵海首席和肖賢方董事長在鑒賞夷陵牛的雪花牛肉曹兵海首席和肖賢方董事長在鑒賞夷陵牛的雪花牛肉-19-（三）（三）精深加工，增值增效精深加工，增值增效如何將夷陵牛的雪花牛肉變成各類高端菜品，將其他牛肉通過菜品開發提檔升級，尤其是將牛皮、牛頭、牛血、牛肚等大量副產物進行開發利用，實現“吃干榨盡”，是加工專家技術孫寶忠研究員和曹兵海教授的技術攻關重點；同時，產品預處理、烹飪、搭配，預制菜的保鮮和口味還原等，每一道工序都是技術和情懷的結合，匠心獨運，方得始終！2018年 9 月，夷陵雪花牛肉榮獲第二屆中國牛肉美食烹飪大賽“中國

29、優質牛肉（西餐紅肉）特優食材獎”。2020 年 12 月，夷陵牛雪花牛肉參與首屆“中國牛優質牛肉品鑒大會”，榮獲本次大會“最具特色獎”、“最具效益獎”和“高品質牛肉生產推介品種”榮譽證書。夷陵牛和夷陵牛雪花肉均獲國家地理標志商標。四、苦練內功，集成創新，鑄造核心競爭力四、苦練內功，集成創新，鑄造核心競爭力龍頭企業是夷陵牛產業發展的載體，而企業的核心競爭力是特色的技術思路和具有自主知識產權的特色產品。比爾蓋茨曾說，如果你給人們一些工具，讓他們發揮自己的天賦和好奇心，他們就會開發出出乎你意料之外的東西。湖北省豐聯佳沃農業開發有限公司在向專家問技的同時苦練內功，在夷陵?！耙员４谩蹦Ｊ街嘘J出了一片天

30、地，名為“牛郞山”。企業從養殖、屠宰分割、加工到餐飲、市場各環節配備了人才隊伍和設備設施，結合自身條件和產業以及市場需求，在預制、菜單、口味、品色、包裝、設計等方面，進行技術集成和自主研發。目前夷陵牛的胴體根據雪花肉的含-20-量和分布，按 25-33 個部位精細分割。除了雪花牛肉產品外，以展現夷陵牛肉質特征與滿足當地市場需求為宗旨，以“楚菜”為特色，開發了牛肉干、牛肉醬、牛肉火鍋共 15 個單品，其中預制火鍋產品 10 款，并且開發了金湯帶皮牛肉火鍋、金湯牛脊排等七款預制菜產品（圖 6，圖 7）。產品線上線下面向全國銷售，同時，將銷售終端延伸到精準客戶身邊，深受市場歡迎。圖圖 6 6夷陵牛肉

31、深加工車間夷陵牛肉深加工車間圖圖 7 7金湯帶皮牛肉火鍋金湯帶皮牛肉火鍋-21-五、疾病凈化，健康護航五、疾病凈化，健康護航安全是美味的基礎，健康是效益的基礎。郭愛珍教授從牛場生物安全體系建設、重要疾病監測和凈化等著手，為保障夷陵牛的健康、高效繁育、牛肉和食品安全等提供了技術支撐，企業牛群在?？谔阋?、牛布魯氏菌病和結核病等重要牛病維持凈化狀態，消費者對夷陵牛產品吃得安心，吃得放心，吃得舒心！-22-夷陵牛產業發展模式之“商業模式”肖賢方湖北豐聯佳沃農業開發有限公司根據原宜昌市畜牧獸醫局肉牛發展的整體布局和關于印發宜昌市“夷陵黃?！碑a業發展總體方案的通知精神，湖北豐聯佳沃農業開發有限公司（以下簡

32、稱“公司”）承擔了夷陵牛保種和開發利用的示范任務。公司位于枝江市仙女鎮腹地，占地 1500 畝，近 10 年來，在國家肉牛牦牛產業技術體系、部省市和枝江市的高度重視、引導及大力支持下，癡心不改探索耕耘夷陵牛保護和養殖、屠宰和加工、銷售和餐飲服務、品牌培育等領域，已經步入了種、養、加、銷、游、網一二三產業融合，循環發展、綠色發展的良性軌道，基本形成了夷陵?！耙杂么俦！比a業鏈發展的商業模式?，F將發展模式整理成文并發布，期望能為我國地方黃牛品種保護和開發利用提供借鑒和參考，以促進我國肉牛業的高效可持續發展。一、在一、在“養養”上下功夫，為上下功夫，為“商商”生產豐富原料生產豐富原料一是種質保護一是

33、種質保護。2016 年，根據原宜昌市畜牧獸醫局關于印發宜昌市“夷陵黃?！碑a業發展總體方案的通知精神，公司增加投資、流轉土地、編制規劃，在體系專家指導下，高標準建設了宜昌市第一個夷陵牛保種場。隨后，公司按照夷陵?！皟珊谝还S”（眼圈黑、黑鼻鏡、筍角）的主要特征，深入枝江、五峰等縣市區的肉牛養殖戶，購買 200 頭夷陵黃牛。精準開展選種、選配、擴群，逐步建立了 6 個家系的原-23-種夷陵牛種群，2017 年夷陵牛通過國家畜禽遺傳資源委員會專家鑒定。2018 年原農業部第 2637 號公告夷陵牛為我國地方畜禽遺傳資源。湖北省和宜昌市、枝江市三級政府每年安排 100 萬左右支持公司開展夷陵牛保護。截止

34、目前公司夷陵牛存欄 416 頭，其中公牛 16 頭、母牛 400 頭。圖圖1 1曹兵海首席帶領專家團隊來宜昌考察驚喜發現瑪瑙河邊本地黃牛曹兵海首席帶領專家團隊來宜昌考察驚喜發現瑪瑙河邊本地黃牛圖圖 2 2產業技術體系崗位科學家李俊雅研究員指導夷陵牛保種工作產業技術體系崗位科學家李俊雅研究員指導夷陵牛保種工作-24-圖圖 3 3夷陵牛核心群夷陵牛核心群圖圖 4 4宜昌市第二屆夷陵牛評鑒會宜昌市第二屆夷陵牛評鑒會二是品種改良二是品種改良。在國家肉牛牦牛產業技術體系曹兵海首席科學家領銜的專家團隊的指導下，公司積極開展夷陵牛精準飼喂和營養動態調控、雜交改良等技術攻關。2017 年 1 月成功培育出夷陵

35、牛雪花牛肉，經國內專家品鑒，已經達到日-25-本雪花牛肉分級標準 A3 級別，市場價格達到 1200 元/公斤，填補了宜昌雪花牛肉的空白。2018 年，試驗將安格斯牛、和牛與夷陵牛雜交改良，已成功繁育出“夷安”雜交牛 1 代、“夷和”雜交牛 1 代，提高了夷陵牛生產性能，增加了雜交牛日增重、體重和雪花牛肉產量。圖圖 5 5夷陵牛雪花牛肉夷陵牛雪花牛肉三是規模養殖三是規模養殖。2014 年公司投資 3000 多萬元建成肉牛標準化養殖場，養殖能力達到 1000 頭。2016 年在產業技術體系崗位科學家劉繼軍教授的技術指導下，新增投資建成夷陵牛保種場，養殖能力達到 500 頭。2017 年在枝江市政

36、府精心組織下，公司牽頭組建了枝江市聯強農牧專業合作社，積極推行“公司+合作社+農戶”、“公司+農戶”、“公司+扶貧車間”等肉牛養殖經營模式。產業技術體系崗位科學家郭愛珍教授牽頭組織專家團隊，深入合作社開展牛病防控技術服務。目前，公司共聯農帶農肉牛飼養量達到 1.2 萬頭，其中公司核心養殖場 1500 頭、帶動合作社農戶飼養肉牛 1.05萬頭。-26-圖圖 6 6產業技術體系崗位科學家郭愛珍教授指導疫病防控工作產業技術體系崗位科學家郭愛珍教授指導疫病防控工作二、在二、在“加加”上作文章，為上作文章，為“商商”開發優質產品開發優質產品一是屠宰分割一是屠宰分割。為實現優良的肉牛賣出優良的價格，公司從

37、“賣?！鞭D變為“賣肉”是第一步，從所有部位牛肉賣一個價轉變為不同部位不同售價是第二步。2016 年 11 月，在枝江市政府的大力支持下，公司建成肉牛標準化屠宰加工車間，年屠宰加工肉牛能力可達到 10000 頭。按照屠宰劈半檢疫排酸分割包裝的生產流程，增加牛肉的嫩度和風味，對牛肉精細分割達到 16 個部位，分別輔以獨立包裝，分部位分級定價，低則 60 多元斤、高則118 元斤，極大提升了生鮮肉品的銷售收益。圖圖 7 7公司公司+合作社合作社+農戶農戶-27-圖圖 8 8分部位牛肉分部位牛肉二是精深加工二是精深加工?；陔x消費者嘴巴越近的商品價值會越高的特點，在產業技術體系崗位科學家孫寶忠研究員的

38、現場指導下，公司對非暢銷的牛體部位進行改頭換面，將牛全身吃干榨凈，提高產品附加值。公司將碎肉制作成包子餡，開發生產風味獨特的牛肉包子，受到市民熱捧，每天供不應求；公司將整張低價出售的生牛皮，精細加工成“不吹牛皮”涼菜，變成了可口獨特的熟食品；公司又將過去用作飼喂寵物的牛肝，加工精細制作成“怪味牛肝”菜品，成為廣受歡迎的佐酒佳肴。公司搶抓預制菜產業發展風口，進軍牛肉預制菜領域。2020 年 12 月，公司建成牛肉食品加工車間，研發生產五香、麻辣風味牛肉干等休閑食品；牛肉包、牛肉餃子、牛肉醬等風味食品；紅燒牛肉、金湯牛肉、牛三鮮、原味牛寶等系列牛肉預制菜，受到消費者的廣泛青睞。2023 年 1 月

39、“牛味央”預制菜入圍湖北省“地道荊楚味”首批 30 家企業推薦品牌。-28-圖圖 9 9曹兵海首席多次來牛郎山指導夷陵牛品種開發利用曹兵海首席多次來牛郎山指導夷陵牛品種開發利用圖圖 1010產業技術體系崗位科學家孫寶忠研究員指導肉牛產品加工產業技術體系崗位科學家孫寶忠研究員指導肉牛產品加工-29-圖圖 1111特色牛肉包、牛肉干特色牛肉包、牛肉干圖圖 1212“不吹牛皮不吹牛皮”、“鴻運當頭鴻運當頭”特色菜肴特色菜肴圖圖 1313牛肉預制菜牛肉預制菜三是品牌培育。三是品牌培育。農產品的最佳出路是標準化和品牌化。宜昌市、枝江市兩級政府真金白銀支持公司鞏固擴大夷陵牛品種保護與利用的成果，培育打造牛

40、肉產品系列品牌，實現了產品提質增效。公司先后注冊了“仙女牛郎山”、“牛味央”、“國?！钡犬a品商標。宜昌市畜牧獸醫中心成功登記-30-注冊夷陵牛和夷陵牛雪花牛肉國家地理標志證明商標，并授權公司使用。2018 年 9 月，夷陵雪花牛肉榮獲第二屆中國牛肉美食烹飪大賽“中國優質牛肉（西餐紅肉）特優食材獎”。2020 年 12 月，夷陵牛雪花牛肉參與首屆“中國牛優質牛肉品鑒大會”，榮獲本次大會“最具特色獎”、“最具效益獎”和“高品質牛肉生產推介品種”榮譽證書。2021 年“牛味央”牛肉產品通過綠色食品認證，榮獲“枝江三寶”稱號。圖圖 1414夷陵牛雪花牛肉榮獲獎項夷陵牛雪花牛肉榮獲獎項三、在三、在“銷銷

41、”上開新局，為上開新局，為“商商”拓展廣闊渠道拓展廣闊渠道一是餐飲服務銷售一是餐飲服務銷售。公司在枝江、宜昌城區建設了 2 個特色牛肉主題餐飲店、2 個品牌社區餐飲店，4 個連鎖牛肉面館、3 個牛肉專營店，從賣牛到賣肉，從賣肉到賣菜，大大提高了肉牛產業附加值。除了直面消費客戶，發展忠實粉絲的同時，瞄準武漢、粵港澳灣區等經濟發達地區，大力拓展行業大客戶。-31-圖圖 1515特色牛肉餐飲店特色牛肉餐飲店二是直達終端銷售。二是直達終端銷售。隨著牛郎山系列產品市場占有率、消費認知度越來越高，公司大力發展牛肉產品營業網點，深耕本地市場，將產品直達零售終端，先后進駐枝江、宜昌、武漢等城市政府機關、企事業

42、單位食堂和職工超市共 50 多家。2021 年 4 月公司產品率先進駐枝滋有味致祥路體驗店，隨后進駐宜昌 710 職工超市、宜昌市委職工超市、宜昌市政府職工超市、宜昌市農業農村局職工超市，今年還進駐湖北省住房和城鄉建設廳、葛洲壩中心醫院、夷陵婦幼醫院、宜昌供銷社等單位食堂。圖圖 1616產品進駐政府機關、企事業單位食堂、超市產品進駐政府機關、企事業單位食堂、超市-32-三是電商平臺銷售三是電商平臺銷售。充分發揮“牧場直供”的優勢，公司積極探索線上線下融合銷售新模式。通過天貓、京東、微店、抖音等公共電商平臺，面向全國市場，銷售牛郎山夷陵牛系列產品?；谖⑿啪€上、線下互相導流，建立私域流量池，深度

43、耕耘全省市場。同時，借助國家扶持農村“土特產”農產品銷售的大勢，拓展進駐中國三峽集團、國家電網、建行善融商務、鐵路 12306 等細分垂直電商領域。2022 年線上牛肉產品銷售收入 0.5 億元。-33-附：夷陵牛部分研究成果附：夷陵牛部分研究成果1 沈洪學,朱德江,田彥孜,熊雄,馬曄.夷陵黃牛品種資源調查報告（一）夷陵黃牛形成過程的考證.中國牛業科學.2016,42(4):76-79.2 沈洪學,朱德江,田彥孜,熊雄,馬曄.夷陵黃牛品種資源調查報告（二）夷陵黃牛的產地與分布情況.中國牛業科學.2016,42(5):76-77.3 沈洪學,朱德江,田彥孜,熊雄,馬曄.夷陵黃牛品種資源調查報告（

44、三）夷陵黃牛肉用選育及產業化開發思路.中國牛業科學.2016,42(6):78-80.4 李發柱.夷陵黃牛的保種及雜交改良利用.農家致富顧問.2018,2:61.5 夏小婷,賈媛,趙明,熊雄,黨瑞華,藍賢勇,黃永震,陳宏,雷初朝.夷陵黃牛 mtDNA D-loop 區遺傳多樣性研究.中國牛業科學.2017,43(1):4-7.6 夏晗,李凡,彭玲偉,杜玉琴,郭愛珍,楊利國,周揚.基于全長轉錄組測測序的牛脂肪組織可變剪接事件分析.華中農業大學學報.2022,41(6):176-183.7 Xu Ling,Zhang Wen-gang,Li Jun-ya,Zhu De-jiang,Xu Xiao-

45、cheng,Tian Yan-zi,Xiong Xiong,Guo Ai-zhen,Cao Bing-hai,Niu Hong,Zhu Bo,Wang Ze-zhao,LiangYong-hu,Shen Hong-xue,Chen Yan.Genetic background analysis and breedevaluation of Yiling yellow cattle.Journal of Integrative Agriculture.2017.16(10):2246-2256.8 Qiu X,Qin X,Chen L,Qiu Q,Wang H,Aziz Ur Rahmanand

46、 M,Cao B,Su H.Effectsof Age and Rice Straw Inclusion Levels in the Diet of Yiling Cull Cows on GrowthPerformance,Meat Quality,and Antioxidant Status of Tissues.Animals(Basel).2021,11(6):1732.9 Qiu X,Qin X,Chen L,Chen Z,Hao R,Zhang S,Yang S,Wang L,Cui Y,Li Y,Ma Y,CaoB,Su H.Serum Biochemical Parameter

47、s,Rumen Fermentation,and RumenBacterial Communities Are Partly Driven by the Breed and Sex of Cattle WhenFed High-Grain Diet.Microorganisms.2022,10(2):323.-34-35-36-37-38-39-40-41-42-43-湖北湖北-44-第41卷 第6期2022年11月華中農業大學學報Journal of Huazhong Agricultural UniversityVol.41No.6Nov.2022，176183基于全長轉錄組測序的牛脂肪組

48、織可變剪接事件分析夏晗，李凡，彭玲偉，杜玉琴，郭愛珍，楊利國，周揚華中農業大學動物科學技術學院、動物醫學院/農業動物遺傳育種與繁殖教育部重點實驗室，武漢 430070摘要為探究可變剪接影響下更為復雜的脂肪沉積調控網絡，明晰可變剪接事件的發生對脂肪沉積調控機制的影響，本研究基于PacBio測序平臺的第三代測序技術，對夷陵黃牛腹腔脂肪、皮下脂肪、肌間脂肪進行全長轉錄組測序分析。共發現15 445個基因檢測到50 520個可變剪接，占牛全部基因的33.4%。對這些基因進行富集分析發現，83個GO條目顯著富集，這些顯著富集的通路可能參與脂肪沉積網絡的調控，其中16個與脂質合成及代謝相關，27條KEGG

49、通路顯著富集，其中15條與脂質合成及代謝相關。使用KOG、KEGG、NR、SwissProt、GO數據庫對測序序列進行注釋，80 756 條序列共注釋到 69 259 個基因，94 458條CDS序列及對應的pep序列。其中GO分析中15 039條序列富集到507個生物學過程條目，7 907條序列富集到214個細胞組分條目，30 672條序列富集到559個分子功能條目。KEGG數據分析富集到34條通路共49 710條序列，其中7 002條序列參與信號轉導途徑。以上結果表明，牛脂肪組織存在大量可變剪接事件，并且這些發生可變剪接事件的基因對脂肪沉積的調控發揮重要作用，這可從基因可變剪接角度為解析脂

50、肪沉積的調控網絡提供必要的資源和理論依據。關鍵詞脂肪沉積；功能注釋；可變剪接；全長轉錄組；夷陵黃牛中圖分類號S814.8文獻標識碼A文章編號1000-2421（2022）06-0176-08肌內脂肪的含量以及沉積分布影響牛肉的嫩度、系水力、剪切力值、風味和多汁性，適當提高牛肉肌內脂肪含量可改變牛肉品質1-2。但是，當皮下脂肪的積累較多時會降低飼料報酬，造成資源損耗3。脂肪沉積受復雜的調控網絡影響，Utrera等4針對包括脂肪沉積能力的牛胴體性狀遺傳力進行了72 項研究，也未能完全了解其遺傳機制?？勺兗艚邮羌艚芋w對前體 mRNA 選擇性的修飾，使同一基因可以表達不同成熟mRNA剪接異構體的現象5

51、?？勺兗艚颖徽J為在基因中廣泛存在，有研究顯示，在高通量測序結果中95%以上的人類基因存在可變剪接，由此產生表達多種不同的剪接異構體6?？勺兗艚邮够虮磉_具有更多可能性，展示了基因功能的多樣性7。研究顯示可變剪接的發生會導致蛋白結構的變化，導致同一基因的不同蛋白亞型在功能上出現差異8；且有研究表明，可變剪接會對牛的脂肪相關基因表達產生重要影響9。例如，NOVA 依賴性的可變剪接會調節白色脂肪組織褐變10、質量減輕導致的皮下脂肪中 TCF7L2 可變剪接調節可導致脂肪組織中的高血糖和胰島素作用受損11等。所以了解可變剪接事件的發生對于明晰脂肪沉積調控網絡有著重要的作用?；赑acBio 平臺的全長

52、轉錄組測序是第三代測序技術的代表，能直接測序得到RNA轉錄本的全長片段，可以規避短片段帶來的缺失信息的影響，提供更完整的轉錄組信息，更好地進行差異表達基因分析及功能注釋12。同時，長讀技術有利于更精確、更準確、更多地發現可變剪接事件，有助于提高對脂肪沉積調控網絡的認識13。目前，第三代測序技術已經被廣泛應用于植物的遺傳信息挖掘，并取得了較好的結果，例如：赪桐的種質資源挖掘14、紫娟茶樹葉片呈色機理及物質合成途徑15、掌葉魚黃草的遺收稿日期：20220418基金項目：國家自然科學基金項目（31902148）夏晗，E-mail：通信作者：周揚，E-mail：夏晗，李凡，彭玲偉，等.基于全長轉錄組測

53、序的牛脂肪組織可變剪接事件分析 J.華中農業大學學報，2022，41（6）：176183.DOI：10.13300/ki.hnlkxb.2022.06.020傳信息挖掘16等。但是，目前基于全長轉錄組的研究較少，尤其是牛類動物繁殖周期長、生長緩慢更是限制了其研究的進展。本研究利用基于PacBio測序平臺的第三代測序技術對夷陵黃牛脂肪組織遺傳信息進行挖掘，以期為正確認識可變剪接對脂肪沉積調控網絡的機制提供理論參考。1材料與方法1.1試驗材料供試牛為湖北省宜昌地區、年齡在3歲、發育正常、體態良好的3頭健康夷陵黃牛。屠宰后，使用消毒手術刀，分別在每頭牛相同部位采集5 cm3大小的皮下脂肪、腹腔脂肪、

54、肌間脂肪樣品。使用75%乙醇徹底清洗組織，PBS清洗殘留在組織上的乙醇，吸干水分后于-80 液氮保存，用于總RNA 提取。1.2總RNA提取使用 TRIZOL 法提取夷陵黃牛脂肪組織總RNA。提取后的 RNA 需使用 1%凝膠電泳檢測樣品純凈度，并經過Nano Drop儀器檢測，確定總RNA的OD260/OD280值在 1.92.1、OD260/OD230值在1.41.8。并使用 Qubit對 RNA 進行定量，Agilent 2100、Bioanalyzer 等進一步進行質量檢測，提取的總 RNA質量滿足要求后，低溫保存，待后續進行文庫構建。1.3文庫構建及測序構建文庫所使用的RNA需要使用

55、 Qiagen 試劑盒進行回收純化預處理，以獲得更高質量的 RNA。隨后使用 SMARTerPCR cDNA Synthesis Kit 試劑盒反轉得到cDNA。依據cDNA質量優化擴增程式，使用KAPA HiFi PCR Kits 試劑盒進行擴增，對獲得的全長 cDNA 片段使用 SMRT bell templateprep kit 1.0試劑盒構建文庫。同時，對cDNA進行末端修復、損傷修復處理，并在全長cDNA的雙端添加stem loop sequencing adapter 進行篩選，文庫構建成功后，利用 PacBio Sequel測序平臺開展全長轉錄組測序。1.4全長轉錄組數據組裝與

56、篩選對 建 庫 測 序 得 到 的 原 始 數 據，首 先 使 用SMRTLINK 5.1 軟件對其進行質控，然后使用SNR（Signal Noise Ratio）篩選剔除低質量數據及接頭序列，得到環形一致序列（circular consensus sequence，CCS）。去除含 5 primer、3 primer、poly A 片段，獲得的全長轉錄組序列經過校正和去冗余后，可用于后續分析。1.5可變剪接檢測試驗使用ICE（iterative isoform-clustering）中的algorithm 模塊進行聚類，使用 DAGCon 得到一致性序列，并通過arrow 進行校正。將二代數

57、據校正后的全長序列與參考基因組比對，去除融合基因及非注釋數據后，將比對到參考基因組同一基因上的不同isoform進行提取，即為可變剪接。根據可變剪接不同的形成方式進行分類。1.6全長轉錄組功能注釋與分類獲得高質量的全長轉錄組序列后，使用KOG 蛋白質真核同源數據庫、GO基因本體數據庫、KEGG代謝通路數據庫、NR非冗余蛋白數據庫、Swiss Prot精準蛋白數據庫對預測的Isoforms進行注釋，用以評估脂肪組織中基因的功能注釋信息及其參與的代謝通路及調控網絡，并判斷基因的物種相似性。2結果與分析2.1測序質量分析對采集的夷陵黃牛的皮下脂肪、腹腔脂肪、肌間脂肪樣品使用PacBio平臺第三代測序

58、技術進行全長轉錄組測序，共獲得19.81 G的原始數據，篩除接頭和長度小于50 bp的原始離線數據后，得到13 014 954條Subreads，平均長度為 1 474 bp（Full Passes1、最小預測準確度 0.8），核驗下機數據中的 CCS（circularconsensus sequence）的質量，共獲得 779 345 個 CCSreads，CCS總堿基數為 1 812 898 181 bp，CCS Read平均長度2 320.43 bp，平均 Full Pass 數量為11。通過驗證5 引物、3 引物和poly-A尾的保留，獲得高質量 的 FLNC（full-length

59、nonchimeric reads）640 205條，占比 82.15%，平均長度為2 091 bp，以上結果說明全長轉錄組測序數據質量較好，可以用于后續試驗。2.2全長轉錄組 Reads 聚類與校正利用來源于相同樣品的二代測序數據對PacBio平臺的第三代測序reads進行校正17。校正數據后，我們獲得了779 345個consensus reads。使用ICE去除缺失序列和冗余序列后，進一步使用Lordec軟件，以二代數據作為參照對356 772 條一致性序列結果進行校正，獲得可用于進一步分析的一致性序列總長為790 043 023 bp。-45-第 6 期夏晗 等：基于全長轉錄組測序的牛

60、脂肪組織可變剪接事件分析傳信息挖掘16等。但是，目前基于全長轉錄組的研究較少，尤其是牛類動物繁殖周期長、生長緩慢更是限制了其研究的進展。本研究利用基于PacBio測序平臺的第三代測序技術對夷陵黃牛脂肪組織遺傳信息進行挖掘，以期為正確認識可變剪接對脂肪沉積調控網絡的機制提供理論參考。1材料與方法1.1試驗材料供試牛為湖北省宜昌地區、年齡在3歲、發育正常、體態良好的3頭健康夷陵黃牛。屠宰后，使用消毒手術刀，分別在每頭牛相同部位采集5 cm3大小的皮下脂肪、腹腔脂肪、肌間脂肪樣品。使用75%乙醇徹底清洗組織，PBS清洗殘留在組織上的乙醇，吸干水分后于-80 液氮保存，用于總RNA 提取。1.2總RN

61、A提取使用 TRIZOL 法提取夷陵黃牛脂肪組織總RNA。提取后的 RNA 需使用 1%凝膠電泳檢測樣品純凈度，并經過Nano Drop儀器檢測，確定總RNA的OD260/OD280值在 1.92.1、OD260/OD230值在1.41.8。并使用 Qubit對 RNA 進行定量，Agilent 2100、Bioanalyzer 等進一步進行質量檢測，提取的總 RNA質量滿足要求后，低溫保存，待后續進行文庫構建。1.3文庫構建及測序構建文庫所使用的RNA需要使用 Qiagen 試劑盒進行回收純化預處理，以獲得更高質量的 RNA。隨后使用 SMARTerPCR cDNA Synthesis Ki

62、t 試劑盒反轉得到cDNA。依據cDNA質量優化擴增程式，使用KAPA HiFi PCR Kits 試劑盒進行擴增，對獲得的全長 cDNA 片段使用 SMRT bell templateprep kit 1.0試劑盒構建文庫。同時，對cDNA進行末端修復、損傷修復處理，并在全長cDNA的雙端添加stem loop sequencing adapter 進行篩選，文庫構建成功后，利用 PacBio Sequel測序平臺開展全長轉錄組測序。1.4全長轉錄組數據組裝與篩選對 建 庫 測 序 得 到 的 原 始 數 據，首 先 使 用SMRTLINK 5.1 軟件對其進行質控，然后使用SNR（Sign

63、al Noise Ratio）篩選剔除低質量數據及接頭序列，得到環形一致序列（circular consensus sequence，CCS）。去除含 5 primer、3 primer、poly A 片段，獲得的全長轉錄組序列經過校正和去冗余后，可用于后續分析。1.5可變剪接檢測試驗使用ICE（iterative isoform-clustering）中的algorithm 模塊進行聚類，使用 DAGCon 得到一致性序列，并通過arrow 進行校正。將二代數據校正后的全長序列與參考基因組比對，去除融合基因及非注釋數據后，將比對到參考基因組同一基因上的不同isoform進行提取，即為可變剪接

64、。根據可變剪接不同的形成方式進行分類。1.6全長轉錄組功能注釋與分類獲得高質量的全長轉錄組序列后，使用KOG 蛋白質真核同源數據庫、GO基因本體數據庫、KEGG代謝通路數據庫、NR非冗余蛋白數據庫、Swiss Prot精準蛋白數據庫對預測的Isoforms進行注釋，用以評估脂肪組織中基因的功能注釋信息及其參與的代謝通路及調控網絡，并判斷基因的物種相似性。2結果與分析2.1測序質量分析對采集的夷陵黃牛的皮下脂肪、腹腔脂肪、肌間脂肪樣品使用PacBio平臺第三代測序技術進行全長轉錄組測序，共獲得19.81 G的原始數據，篩除接頭和長度小于50 bp的原始離線數據后，得到13 014 954條Sub

65、reads，平均長度為 1 474 bp（Full Passes1、最小預測準確度 0.8），核驗下機數據中的 CCS（circularconsensus sequence）的質量，共獲得 779 345 個 CCSreads，CCS總堿基數為 1 812 898 181 bp，CCS Read平均長度2 320.43 bp，平均 Full Pass 數量為11。通過驗證5 引物、3 引物和poly-A尾的保留，獲得高質量 的 FLNC（full-length nonchimeric reads）640 205條，占比 82.15%，平均長度為2 091 bp，以上結果說明全長轉錄組測序數據質

66、量較好，可以用于后續試驗。2.2全長轉錄組 Reads 聚類與校正利用來源于相同樣品的二代測序數據對PacBio平臺的第三代測序reads進行校正17。校正數據后，我們獲得了779 345個consensus reads。使用ICE去除缺失序列和冗余序列后，進一步使用Lordec軟件，以二代數據作為參照對356 772 條一致性序列結果進行校正，獲得可用于進一步分析的一致性序列總長為790 043 023 bp。177-46-第 41 卷華 中 農 業 大 學 學 報2.3可變剪接分析校正后的數據與參考基因組進行比較，得到與2 095個融合基因相關的2 768條reads。將剩余數據進一步過濾

67、去冗余。使用Match Annot軟件將比對后結果與注釋信息進行比較，并將三代注釋結果和原基因組結果進行合并，得到基因注釋的 PB isoform 63 400 條，非基因區序列有36 480 條。將比對到參考基因組 cDNA區域的序列進行檢測，共識別到可變剪接事件50 520個。其中2 406個基因發生外顯子跳躍、1 184個基因發生外顯子替換、1 262個基因發生外顯子接受、4 085個基因發生內含子保留、2 365個基因發生外顯子位置替換、4 143個基因發生其他可變剪接事件（表1）。將檢測到可變剪接事件的基因進行GO富集分析，顯示與83個條目顯著相關（P0.95.4)Call rate

68、,average call rate of individual in each breed.5)Average minor allele frequency.6)Average expected heterozygosity.7)Average observed heterozygosity.-60-2250XU Ling et al.Journal of Integrative Agriculture 2017,16(10):224622563.2.Principal components analysisFig.4 showed the PCA results of PC1 vs.PC2

69、,with PC1 dimension accounting for mainly total variance(6.96%)among breeds.Yiling yellow cattle and Qinchuan cattle grouped together in PC1 dimension,respectively,leading to two obvious independent separation from other breeds.While ZB,BS,and WL individuals blurred into each other in PC1 dimension.

70、Furthermore,it is worth noting that compared to other breeds,individuals from YL clustered together more obviously.3.3.neighbor-joining analysisNeibour-joining tree(NJ)relating the 154 individuals was constructed in Fig.5.Consistent with PCA results,YL and QC could be clearly distinguished,indicatin

71、g that the YL individuals performed a cluster.Nevertheless,BS,WL and ZB were performed to cluster together with disordered genetic variance.Genetic distance among individuals were measured by the length of branches.YL showed a distant genetic relationship with other breeds,and BS displayed the close

72、st genetic relationship with WL.3.4.neis genetic distanceTo provide additional insight into the genetic variation of five breeds,the Neis distance was used to evaluate genetic dis-tance at population level.Table 3 showed pairwise genetic distance among five breeds,in which Neis distance ranged from

73、0.011 to 0.085 with a mean of 0.034.As expect,YL and QC possessed the most distinct genetic distance about 0.085,and WL and ZB had the closest genetic relationship of 0.011.Based on the values of Neis distance,we further con-structed the NeighbourNet graph(Fig.6)to infer the rela-tionship between br

74、eeds.BS,ZB and WL breeds clustered intermediate position between YL and QC.In accordance with PCA result,QC cattle located distant to the other four breeds whereas YL cattle was located nearer to the cluster Fig.3 Frequency histogram of minor allele frequency(MAF)in five cattle breeds(YL,Yiling yell

75、ow cattle;BS,Bashan cattle;WL,Wuling cattle;ZB,Zaobei cattle;QC,Qinchuan cattle).Fig.4 Principle component analysis(PCA)plot of the genetic relationship matrix for 154 individuals of five breeds studied.PC1 explained 6.69%of total variation;PC2 explained 1.40%of total variation.YL,Yiling yellow catt

76、le;BS,Bashan cattle;WL,Wuling cattle;ZB,Zaobei cattle;QC,Qinchuan cattle.0.30.20.10FrequencyIII III IV VIII III IV VIII III IV VIII III IV VIII III IV VYLBSWLZBQCIMAF0.1II 0.1MAF0.2III 0.2MAF0.3IV 0.3MAF0.4V 0.4MAF0.5YLZBWLQCPC1PC2BS204020204060801000101020304050607080YLZBWLQCPC1PC2BS204020204060801

77、00010102030405060708040202040608010004020204060801000-61-2251XU Ling et al.Journal of Integrative Agriculture 2017,16(10):22462256from BS,ZB and WL,and especially,YL had formed an independent branch.3.5.Linkage disequilibriumLinkage disequilibrium(LD)decay of five breeds was es-timated,and Fig.7 ill

78、ustrated the level of LD decreased with the physical distance increasing.When the distance between markers was approximately 2 Mb,in comparison with others,LD of YL was the lowest(r2=0.025).From 0 to 100 kb,the degree of LD of YL decreased from 0.32 to 0.1,LD of QC decreased from 0.46 to 0.12,which

79、indicated that YL had a history of long-term selection,compared with QC.When physical distance extended to 100 kb,the LD 0.03Fig.5 Neighbor-Joining tree relating the 154 individuals in the five cattle breeds.Font color:YL,Yiling yellow cattle,red;BS,Bashan cattle,purple;WL,Wuling cattle,green;ZB,Zao

80、bei cattle,blue;QC,Qinchuan cattle,black.Table 3 Pair-wise Neis distance among five cattle breedsBreed1)WLBSYLQCZBWL00.0167100.0141640.0581330.010939BS0.01671000.0224240.0482940.014474YL0.0141640.02242400.0854360.012377QC0.0581330.0482940.08543600.054520ZB0.0109390.0144740.0123770.05452001)YL,Yiling

81、 yellow cattle;BS,Bashan cattle;WL,Wuling cattle;ZB,Zaobei cattle;QC,Qinchuan cattle.-62-2252XU Ling et al.Journal of Integrative Agriculture 2017,16(10):22462256of each breeds were weak and tended to stability,with YL was 0.1,QC was 0.12,ZB was 0.1,BS was 0.13,and WL was 0.1,respectively.3.6.Select

82、ion signal analysisIn this study,to observe the selection strength of YL in the process of domestication,genome-wide selective sweep analysis was implemented to screen the selected locus in YL(Fig.8).The result revealed that pairwise breed of YL vs.QC had the highest FST average value(0.048),followe

83、d by the YL vs.BS(0.012).YL vs.ZB,YL vs.WL both had an equal FST average value(0.011).After the analysis of selective sweep,the loci of average FST value ranking top 30(1/1 000 loci)were selected for gene annotation.In order to investigate the unique selection signature of YL,we com-pared the signif

84、icant loci of YL vs.three close geographical breeds(BS,WL,and ZB)as showed in the Venn diagram with pairwise comparisons(Fig.9).A total of 90 significant loci distributed in 24 chromosomes,with the most in chro-mosome 1(10)and 9(10)and the least in chromosome 13(1),16(1),17(1),29(1).After selection

85、signal screening,we obtained 86 annotat-ed genes including 76 protein coding genes retrieved from the DAVID database(https:/david.ncifcrf.gov)participating in biological pathways(32 genes,42.5%),cellular compo-nent(22 genes,9.1%),and molecular function(37 genes,48.6%).Meanwhile,27 enrichment items(T

86、able 4)were found by gene-term enrichment analysis,involved in the biological pathway(14 items,51.9%),cellular component(2 items,7.4%),and molecular function(11 items,40.7%).Significantly,the top three pathways were mTOR signalin pathway,Wnt signaling pathway and pancreatic polypeptide receptor acti

87、vity with P-value lower than 0.01.4.DiscussionChina has abundant cattle resources.After long-term domestication,native breeds had formed their own unique genetic characteristics.According to FAO-DAD-IS(http:/dad.fao.org/),there are more than 70 cattle breeds in China,including 53 Chinese indigenous

88、cattle breeds.Currently,Hubei Province has four registered indigenous yellow cattle breeds,including WL,ZB,Huangpi yellow cattle(HP)and BS,in geographically distributed four corners of the province.0.01QCBSZBWLYLFig.6 NeighborNet graph of five cattle breeds of this study.YL,Yiling yellow cattle;BS,B

89、ashan cattle;WL,Wuling cattle;ZB,Zaobei cattle;QC,Qinchuan cattle.Fig.7 Average linkage disequilibrium(LD)decay by r2 values for five population.Upper figure showed LD decays from 0 up to 100 kb for five cattle breeds,and lower figure showed LD decays from 0 to 5 Mb.YL,Yiling yellow cattle;BS,Bashan

90、 cattle;WL,Wuling cattle;ZB,Zaobei cattle;QC,Qinchuan cattle.Fig.8 Scan of FST values distribution for Yiling yellow cattle(YL)vs.the other four breeds by chromosome in selective sweeps.BS,Bashan cattle;WL,Wuling cattle;ZB,Zaobei cattle;QC,Qinchuan cattle.YLBSZBQCWL12345Distance(Mb)Distance(kb)20406

91、08010012000.050.100.150.200.250.300.350.400.45r20.80.60.40.200.2YL vs.ZB YL vs.BS YL vs.WL YL vs.QC SNP position in the genomePlot of FST estimation-63-2253XU Ling et al.Journal of Integrative Agriculture 2017,16(10):22462256YL,which is distributed in the central of Hubei Province,is not yet nominat

92、ed in the list of Animal Genetic Resources in China Bovines.In the past,YL mainly distributed in Zigui County,Yiling District,but now in Zigui County,Changyang Autonomous County,Wufeng Autonomous County and Yiling District.According to the incomplete statistics of relevant counties(districts)by the

93、end of 2015,in Yichang area,the number of yellow cattle population reached more than 43 000,in-cluding about 10 000 YL.Compared with WL and ZB cattle breeds in productive performance,YL has a strong labor performance and unique beef flavor,although the carcass yield of YL is slightly lower than thes

94、e two breeds,with the dressing percentage of 47.1%and meat yield of carcass percentage of 82.6%(Liu and Nu 2007;Li et al.2014).Adult males of YL cattle weigh on 379 kg and females 320 kg with an average withers height of 130 and 120 cm,respectively.In previous investigations,YL had an abundance of h

95、istor-ical background(Shen et al.2016c),and formed a good commercial performance and potential genetic resources under the unique geographical environment and local cultural impaction(Shen et al.2016b).The number scale of YL develops gradually,but conservation system and nu-cleus breeding population

96、 have not been established(Shen et al.2016a).Therefore,the priority is to build the system of monitoring and evaluation for Yiling yellow cattle genetic resource.By comparing with the geographically nearby four indigenous breeds,we implemented a comprehensive genetic background investigation of YL c

97、attle,and provided theoretical basis for YL breed identification and approval.In our study,polymorphisms analysis of MAF,expected heterozygosity(he),and observed heterozygosity(ho)have been applied to estimate genetic diversity among five breeds.According to 27 223 effective makers,we found YL had a

98、 lower polymorphism information content and less MAF(0.2)of more than 50%markers.This result indicated the less genetic variance and higher gene conservation in YL cattle.In addition,PCA and NJ tree analysis were carried out for a better understanding of genetic relationship and distance of YL with

99、Hubei indigenous breeds.According to these two analyses results,YL individuals clustered clearly,which indicated that YL had formed a uniform genetic background.While ZB,BS,and WL individuals blurred into each other,indicated the population structure of the three breeds is disorder.In the assessment

100、 of the population genetic relationships,NeighborNet graph result suggested that YL formed an independent branch with the furthest genetic relationship with QC and slightly close relationship with BS,ZB and WL.Overall,YL contained a unique genetic structure and processed a dependent demonstration fr

101、om other breeds.This phenomenon maybe can be explained by isolated geographical environment and artificial selec-tion.On one hand,Yichang area is in the transition zone of mountain area from western Hubei Province to Jianghan Plain surrounding by the Wushan Mountain,Wuling Moun-tain,and Qiyue Mounta

102、in.Thus traffic inconvenience and complex topography caused a limitation of large-scale gene introgression from other breeds,which provided a possibility to purify the breed.Meanwhile,YL had formed adaptive fitness under the environment of poor soil,high mountains and steep slopes.On the other hand,

103、with local peoples demands of beef flavor,the unique genetic resource of YL cattle was further developed and utilized.unique selection history and population structure for each breeds leads to LD decay difference(Qan-bari et al.2014;Al-Mamun et al.2015).In previous studies,OBrie et al.(2014)calculat

104、ed Fleckvinh LD with Illumina BovineHD assay,resulting r2=0.15 when physical distance was 100 kb.Porto-Neto et al.(2014)revealed LD decreased to 0.46 with a distance of 10 kb in Angus.According to the results of our study,the LD of YL decreased slowly than other breeds up to 100 kb.When the distance

105、 excessed 100 kb,BS had the highest LD,indicating a limit of current effective population size of BS(Flury et al.2010).Since both of these two foreign breeds were currently selected by commercial purposes,the LD of these breeds were higher than Chinese indigenous breeds reasonably.Furthermore,the an

106、notated genes were implemented in YL vs.WLYL vs.ZBYL vs.BS2511224263Fig.9 Venn diagram of the significant loci of Yiling yellow cattle(YL)vs.the other three breeds.Each circle of 30 significant loci represents each of the pairwise cattle(YL vs.WL,YL vs.ZB,YL vs.BS)with numbers within circles or over

107、lapping areas indicating the number of loci.BS,Bashan cattle;WL,Wuling cattle;ZB,Zaobei cattle.-64-2254XU Ling et al.Journal of Integrative Agriculture 2017,16(10):22462256Table 4 Results of gene-term enrichment and pathway annotationCategoryTermGene numbersP-valueBIOCARTAmTOR Signaling Pathway30.00

108、67INTERPROSerine/threonine protein kinase,active site60.009GOTERM_MF_FATPancreatic polypeptide receptor activity20.009INTERPROSerine/threonine protein kinase-related60.011SP_PIR_KEYWORDSSerine/threonine-protein kinase60.013SP_PIR_KEYWORDSPhosphoprotein370.013SMARTS_TK_X30.014GOTERM_BP_FATNegative re

109、gulation of secretion30.016uP_SEQ_FEATuREDomain:AGC-kinase C-terminal30.017GOTERM_CC_FATCytosol110.018GOTERM_MF_FATPeptide YY receptor activity20.018INTERPROAGC-kinase,C-terminal30.018GOTERM_MF_FATProtein serine/threonine kinase activity60.019INTERPROProtein kinase,ATP binding site60.029SMARTIGc240.

110、029GOTERM_BP_FATProtein kinase B signaling cascade20.031uP_SEQ_FEATuREActive site:Proton acceptor70.033INTERPRONeuropeptide Y receptor20.034INTERPROProtein kinase,core60.034GOTERM_BP_FATSteroid metabolic process40.035GOTERM_MF_FATNeuropeptide Y receptor activity20.036GOTERM_BP_FATSteroid biosyntheti

111、c process30.037GOTERM_MF_FATPeptide binding40.037SP_PIR_KEYWORDSKinase70.039GOTERM_CC_FATRibosome40.042GOTERM_BP_FATCholesterol metabolic process30.042INTERPROImmunoglobulin subtype 240.043SP_PIR_KEYWORDSPalmitate40.043PIR_SuPERFAMILYPIRSF002353:S-100 protein20.048GOTERM_BP_FATSterol metabolic proce

112、ss30.05SP_PIR_KEYWORDSAtp-binding100.05GOTERM_MF_FATAdenyl nucleotide binding110.053SMARTS_TKc40.053SP_PIR_KEYWORDSPolymorphism500.055GOTERM_MF_FATPurine nucleoside binding110.057GOTERM_MF_FATNucleoside binding110.06GOTERM_BP_FATPhosphorylation70.062uP_SEQ_FEATuREMutagenesis site130.067GOTERM_MF_FAT

113、Protein kinase activity60.067GOTERM_BP_FATRegulation of GTPase activity30.071INTERPROSerine/threonine protein kinase40.075GOTERM_BP_FATNegative regulation of catalytic activity40.075GOTERM_BP_FATProtein modification by small protein conjugation30.08GOTERM_MF_FATATP binding100.08SP_PIR_KEYWORDSRibonu

114、cleoprotein40.081uP_SEQ_FEATuRESequence variant510.082GOTERM_BP_FATNegative regulation of transport30.083INTERPROBTB/POZ30.083GOTERM_BP_FATBehavior50.084SP_PIR_KEYWORDSLipoprotein60.084GOTERM_MF_FATAdenyl ribonucleotide binding100.086GOTERM_BP_FATProtein amino acid phosphorylation60.086uP_SEQ_FEATuR

115、EDomain:Protein kinase50.093uP_SEQ_FEATuREDomain:BTB30.096INTERPROImmunoglobulin I-set30.096GOTERM_BP_FATRegulation of system process40.096INTERPROS100/CaBP-9k-type,calcium binding,subdomain20.097-65-2255XU Ling et al.Journal of Integrative Agriculture 2017,16(10):22462256gene-term enrichment analys

116、is to explore unique genetic background of YL.The top three significant pathways were found:“Wnt signaling pathway”is related to cell cycle activity,stem cell self-renewal and differentiation regulation(Yin and Song 2011);“mTOR signaling pathway”is involved in the cell growth cycle and related to th

117、e antibacterial immune(Abdel-Nour et al.2014),growth and development(Cong et al.2016);“pancreatic polypeptide receptor activity”par-ticipates in promoting feed intake,regulating intestinal canal activity and affecting internal secretion in chicken(Berntson et al.1993;Denbow et al.1988).These three s

118、ignificant pathways involved in growth,development,energy me-tabolism and other important physiological activities may have positive effects on ecological flexibility,environmental adaptability,crude feed tolerance and constitution specificity of Yiling cattle,and need further research.In general,PC

119、A,NJ tree and genetic distance analysis results revealed that YL individuals clustered together and separated from other breeds clearly.And gene annotation and enrichment results obtained several function pathways related to the development and growth,illustrated that Yiling yellow cattle had formed

120、 a unique genetic resources after long-term natural or artificial selection history.5.ConclusionYiling yellow cattle is a distinct indigenous cattle breed of Yichang region.Our study evaluated genetic background of Yiling yellow cattle and other four geographical nearby breeds.The results showed tha

121、t Yiling yellow cattle has clearly genetic distance between Wuling cattle,Bashan cattle,Zaobei cattle,and Qinchuan cattle,suggesting its distinct genetic resource.This knowledge will give insight into comprehensive understanding the formation of Yiling yellow cattle,and provide genetic evidence for

122、Yiling yellow cattle identification and breed authorization.acknowledgementsThis work was funded in part by the National Natural Science Foundation of China(31402039,31472079,31372294),the Beijing Natural Science Foundation(6154032),the Species and Breed Resources Conservation of the Ministry of Agr

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145、H.2015.Impact of parental Bos taurus and Bos indicus origins on copy number variation in traditional Chinese cattle breeds.Genome Biology and Evolution,7,23522361.(Managing editor ZHANG Juan)-67-3.231ArticleEffects of Age and Rice StrawInclusion Levels in the Diet of YilingCull Cows on Growth Perfor

146、mance,Meat Quality,and AntioxidantStatus of TissuesXinjun Qiu,Xiaoli Qin,Liming Chen,Qinghua Qiu,Haibo Wang,Muhammad Aziz ur Rahmanand,Binghai Cao and Huawei Suhttps:/doi.org/10.3390/ani11061732-68-animalsArticleEffects of Age and Rice Straw Inclusion Levels in the Diet ofYiling Cull Cows on Growth

147、Performance,Meat Quality,andAntioxidant Status of TissuesXinjun Qiu1,Xiaoli Qin1,Liming Chen2,Qinghua Qiu1,Haibo Wang1,Muhammad Aziz ur Rahmanand3,Binghai Cao1,*and Huawei Su1,*?Citation:Qiu,X.;Qin,X.;Chen,L.;Qiu,Q.;Wang,H.;Aziz ur Rahmanand,M.;Cao,B.;Su,H.Effects of Age and Rice StrawInclusion Leve

148、ls in the Diet of YilingCull Cows on Growth Performance,Meat Quality,and Antioxidant Statusof Tissues.Animals 2021,11,1732.https:/doi.org/10.3390/ani11061732Academic Editor:Angela SchwarmReceived:18 March 2021Accepted:8 June 2021Published:10 June 2021Publishers Note:MDPI stays neutralwith regard to

149、jurisdictional claims inpublished maps and institutional affil-iations.Copyright:2021 by the authors.Licensee MDPI,Basel,Switzerland.This article is an open access articledistributedunderthetermsandconditions of the Creative CommonsAttribution(CC BY)license(https:/creativecommons.org/licenses/by/4.0

150、/).1State Key Laboratory of Animal Nutrition,College of Animal Science and Technology,China AgriculturalUniversity,Beijing 100193,China;(X.Q.);(X.Q.);(Q.Q.);(H.W.)2Hubei Fulljoywo Agricultural Development Company Limited,Yichang 443200,China;3Institute of Animal and Dairy Sciences,University of Agri

151、culture,Faisalabad 35200,Pakistan;drazizurrahmanuaf.edu.pk*Correspondence:(B.C.);(H.S.);Tel.:+86-010-6281-4346(B.C.);+86-138-1034-5826(H.S.)Simple Summary:Several negative attributes are associated with carcass and meat from cull cows,including lower carcass weights,inferior tenderness,etc.These und

152、esirable characteristics arenormally more distinct with increasing age.In addition,there are no recommended standards forforage levels in the diet of Chinese local cattle to produce high marbled beef.In the current study,we evaluated the effects of age and forage levels on the finishing performance

153、of Yiling cull cowsunder high-energy ration conditions,and also investigated their antioxidant status related to age anddiet.We found that younger age and adequate forage had better growth performance and carcasstraits.Our findings highlight the importance of selecting according to age and providing

154、 adequateforage feed for Yiling cull cows.Moreover,our study also demonstrated the excellent capability ofproducing high marbled beef for Yiling cattle.Abstract:The objectives of this study were to investigate the effects of age and dietary straw lev-els on growth performance,carcass and meat traits

155、,as well as tissue antioxidant status of Yil-ing cull cows.Twenty-four Yiling cull cows were arranged in a 22 factorial design:two ageclasses consisting of younger cull cows(YCC;appearing with three or four pairs of permanentteeth)and older cull cows(OCC;worn out teeth);two dietary treatments consis

156、ting of lower andhigher rice straw levels(LRS and HRS;providing 0.7 kg/d and 1.2 kg/d rice straw per headbased on air-dry basis,respectively).Cows were fed twice a day.Straw was offered at halfof the predetermined weight each meal;concentrate was separately supplied ad libitum.After300 d of feeding,

157、final body weight(BW),total BW gain,average daily gain and gain:feed intakewere higher(p 0.01)in the YCC group than in the OCC group.Total dry matter intake washigher(p=0.03)in the HRS group than in the LRS group,but neutral detergent fiber apparentdigestibility was negatively affected(p=0.01)by inc

158、reased straw levels.Decreased C15:0,C17:0,C20:5n3c,and saturated fatty acids(SFAs)proportion as well as increased C18:1n9c and monoun-saturated fatty acids(MUFAs)proportion in meat from YCC with HRS diet were observed ascompared to that in meat from YCC with LRS diet(p 0.05)werefound for meat qualit

159、y attributes except for cooking loss,which was higher(p=0.02)in the HRSgroup than in the LRS group.Both YCC group and HRS group had higher(p 0.05)cold carcassweight compared to OCC group and LRS group.Moreover,catalase activity of liver tissue was higher(p=0.045)in YCC than in OCC,while superoxide d

160、ismutase activity of muscle tissue was higher(p=0.01)in LRS than in HRS.Based on results,we concluded that younger age and feeding high-levelstraw can improve the finishing performance of Yiling cull cows.Animals 2021,11,1732.https:/doi.org/10.3390/ani11061732https:/ 2021,11,17322 of 13Keywords:age;

161、antioxidant status;straw levels;meat quality;Yiling cull cows1.IntroductionIn beef meat production systems,carcass and meat traits depend on breed,age,anddiets of slaughtered animals.Previous studies have reported that the tenderness of meathas a negative correlation with the age of cull cows 1,2 du

162、e to the chemical nature ofconnective tissue 3.Furthermore,negative attributes associated with carcass and meatfrom cull cows,including lower carcass weights,smaller Longissimus dorsi muscle(LM)areas,inferior muscling,yellower external fat color,and darker lean color 4,5.Generally,providing high-ene

163、rgy ration to cull cows before slaughter improves carcass characteristicsand meat quality 6,7 and reverses such an influence.The inclusion and the type of forage in the diet of finishing cattle depend upon theavailability,price,and its influence on fattening and meat quality 8.Providing foragein suf

164、ficient amounts is considered as the key factor in preventing ruminal acidosis dueto increased saliva production 9.However,excessive amounts of forage tend to reducedry matter intake(DMI),which may be related to its big bulk,slow rumen passage rateand low apparent digestibility 10,11 and consequentl

165、y influence finishing performance.Therefore,the appropriate amounts of forage are required to maintain rumen health andmaximize finishing performance.In high marbled beef production system,especially with cull cows,the study onthe effect of age and forage levels in the diet could be of great signifi

166、cance.In previousstudies with cull cows,few experiments have been conducted dealing with the effect ofthe breed 1,12,finishing diet 13,14,and age 1,2.They have shown that breed andinteraction with finishing diet can modify the fat deposition in the carcass and meat,themuscle fiber type,the collagen

167、content and composition,and in consequence the meatquality in terms of tenderness,flavor,and juiciness.Furthermore,few experiments havestudied the relationships of different types of finishing diets with beef meat quality ofheifers 15,young bulls 16,and cull beef cows 17,but they did not study facto

168、rs suchas age and their interaction with finishing diet.Yiling cattle originated in Yichang City and has been identified as a new breed ofChinese yellow cattle.Its history as a draft animal can be traced back to 4000 years.Yiling cattle is currently considered as a draught and meat type breed.Previo

169、us studieshave evaluated the genetic background of Yiling cattle 18,but the information about thefinishing performance and meat traits of Yiling cattle is still partially unclear.Antioxidant systems are an important determinant of the oxidative stability of storedmeat,influenced by breed,age,diet,an

170、d environment.The aging process is caused bydeleterious effects of reactive oxygen species(ROS)generated spontaneously from normalcellular metabolism 19.In beef production,cull cows had higher myoglobin or heme-Fecontents 20,which accelerates the oxidation process 21.Previous study elaborated thatin

171、clusion of forage in the diet of beef cattle during the finishing phase altered antioxidantstatus 22.However,the antioxidative status of tissues(i.e.,liver and muscle tissues)incull cows in the finishing phase concerning animal age is still needed to be investigated.For these reasons,our objectives

172、were to assess the effects of age and rice strawinclusion levels in the diet of Yiling cattle on carcass characteristics,and intramuscularfat(IMF)content,tenderness and water-holding capacity of meat.In the current study,we hypothesized that the reduction of the amounts of straw in the finishing die

173、t of cowswould show better beef quality due to the potential enhancement of energy intake.Besides,younger cull cows may have greater finishing performance due to their higher eatingappetite and capacity.The other objective of the current study was to evaluate the tissueantioxidant status of Yiling c

174、ull cows.We hypothesized that older age would induceprotein and lipid oxidation of tissues and consequently enhance its antioxidant capacity.-70-Animals 2021,11,17323 of 13Moreover,dietary forage levels may alter antioxidant status of cull cows under high-energyration conditions.2.Materials and Meth

175、ods2.1.Animals,Management,and SamplingAll experimental procedures were approved by the Animal Welfare and Ethics Com-mittee of China Agricultural University(Permit No.DK1008).Twenty-four Chinese Yilingcows weighing 330.621.1 kg,culled from National Livestock and Poultry Genetic Re-sources Conservati

176、on Farm(Hubei Fulljoywo Agricultural Development Co.Ltd.,Yichang,China),were divided into four groups of six animals each in a 22 factorial design:two age classes consisting of younger cull cows(YCC,appearing with three or four pairsof permanent teeth)and older cull cows(OCC,worn out teeth);two diet

177、ary treatmentsconsisting of lower and higher rice straw levels(LRS and HRS;providing 0.7 kg/d and1.2 kg/d rice straw per head based on air-dry basis,respectively).All cows were rearedin separate pens with ad libitum access to concentrate and water.Feeding frequency ofconcentrate and rice straw were

178、twice a day at 08:00 h and 16:00 h.Rice straw was offeredat half of the predetermined weight each meal;concentrate was separately supplied basedon 5 to 10%orts.The nutrient composition of concentrate and rice straw are shown inTable 1.Table 1.Ingredients and chemical compositions of the experimental

179、 diets.Item1ConcentrateRice StrawIngredient,%(on DM basis)Corn grain53.2Wheat bran29.4Wheat middling15.0Rapeseed meal0.4NaHCO31.0NaCl1.0Chemical composition,%(on DM basis)OM94.386.3CP12.05.1EE6.12.5NDF23.574.8ADF3.539.3DM,%(on air-dry basis)90.693.6ME,Mcal/kg(on DM basis)2.971.681DM,dry matter;OM,or

180、ganic matter;CP,crude protein;EE,ether extract;NDF,neutral detergent fiber;ADF,acid detergent fiber;ME,metabolizable energy.Feed offered and refusals weight were recorded daily to calculate the intake duringthe whole finishing phases.Concentrate and straw samples were collected every month,and faece

181、s were sampled from the rectum daily at 6:00,12:00,18:00,and 24:00 h duringthe last three days of the test.Samples were dried at 65C and smashed by using a mill(Wiley,A.H.Thomas Co.,Philadelphia,PA,USA)with a 40-mesh screen,and then stored at20C until further chemical analysis.Animals were weighed a

182、nd slaughtered at the 300 d of trial.After evisceration,liversamples were immediately collected from its diaphragmatic surface.Meanwhile,musclesamples were taken with a knife in the LM from the left half carcass between the sixthand seventh ribs.All fresh tissue sample was capped in a tube and store

183、d at80C forthe determination of antioxidant status.In the same place,the pH value of carcass wasrecorded using a Testo 205 pH probe(Testo SE&Co.,KGaA,Lenzkirch,BW,Germany)beforeand after agingfor seven daysat 4C.Thehot carcassweight and cold carcass weight(CCW)were recorded to calculate the dressing

184、 percentage and the carcass composition,respectively.After aging,approximately 10-cm-thick LM samples were removed fromthe left half-carcass between the 12th and 13th ribs,and a 5-cm-thick LM sample was-71-Animals 2021,11,17324 of 13immediately separated for physical analysis;the other 5-cm-thick LM

185、 sample was vacuumpacked and stored at20C for chemical analysis and fatty acids analysis.The LM areabetween the 6th and 7th as well as between the 12th and 13th ribs were measured using avegetable parchment with standard grid.The high rib,ribeye,striploin,and tenderloin areconsidered as the top-grad

186、e cuts in China 23.After cutting,deboning,and trimming,allbeef fat and cuts were weighed to calculate the percentage of fat and meat in CCW.2.2.Chemical CompositionsThe dry matter(DM,method 934.01),crude protein(CP,method 990.03),and etherextract(EE,method 920.39)and ash(method 924.05)were determine

187、d according to themethods of the Association of Offical Analytical Chemists(AOAC 2002)24.The organicmatter(OM)was calculated as the difference between DM and ash.Neutral detergent fiber(NDF)and acid detergent fiber(ADF)contents were analyzed following the methods ofVan Soest et al.(1991)25.Chemical

188、analyses were performed on each sample in duplicate.Apparent digestibility was calculated following the endogenous tracer acid-insoluble ash(AIA)method described in our previous trial 26.2.3.Meat QualityExternal fat and thick connective tissue were removed and then the LM sampleswere freeze dried in

189、 a freeze-drier(FD-1-50,Biocool,Beijing,China)for 7 days at50Cto determine the DM contents.The freeze-dried samples were analyzed for CP and EEaccording to the AOAC methods 24.Cooking loss was determined by weight loss duringthe immersing LM sample(333 cm)in a resealable pack bag in a water bath at

190、80Cuntil its internal temperature reached 70C.Six meat cores(1.27-cm-diameters)parallelto the muscle fiber were removed from each cooked sample,and sheared in the centerperpendicular to the muscle fibers using a texture analyzer(TA.XT plus,Godalming,Surrey,UK)for measurement of the Warner-Bratzler s

191、hear force(WBSF).Three meat samples(241 cm)were pressed using the texture analyzer under 25 kg pressure for 5 minto measure the pressing loss.Drip loss was measured by the weight loss during thesuspension of three standardized LM sample(235 cm)in a foam box at 4C for 24 h.2.4.Fatty Acid ProfileFatty

192、 acid profile in the meat was measured according to the fatty acid methyl ester(FAME)analysis method described in our previous trial 27 with slight modification ininternal standard matter.This involved the combination of 100 mg freeze-dried and groundmeat with 4 mL of a mixed solution of ethyl chlor

193、ide and methanol(1:10;v/v)and 2 mL ofan internal standard hexane solution(nonadecanoic acid,1 mg/mL).After hydrolyzation,neutralization(4 mL of 100 g/L potassium carbonate solution),centrifugation(3000g for10 min)and filtration(0.2m pore size),FAME profile in the supernatant was analyzedusing a gas

194、chromatograph(GC-2014,Shimadzu Corporation,Kyoto,Japan)installedwith a hydrogen flame detector and a 100 m0.25 mm inner diameter0.20m filmthickness capillary column(HP-88;Agilent Technologies,Santa Clara,CA,USA).Nitrogenwas used as carrier gas at a flow rate of 1 mL/min and the splitting ratio was c

195、ontrolledat 1:40.The initial oven temperature was 140C and held for 5 min,then increased to230C at 3.5C/min and held for 15 min.The injector and detector temperatures wereset at 280C.The injection volume was 1L.The chromatographic peak was identified bycomparing the relative retention times with tho

196、se of the standard mixtures of 37 FAME(18919-1AMP,Sigma Chemical Co.,Shanghai,China).The relative proportions werecalculated as percentages of summed peak areas.2.5.Antioxidant StatusMuscle and liver sample was homogenized with saline solution to determine theconcentrations of total antioxidant capa

197、city(T-AOC),catalase(CAT),superoxide dismutase(SOD),malondialdehyde(MDA),protein carbonyl(PC),and ROS.All antioxidant parame-72-Animals 2021,11,17325 of 13ters were examined in triplicate using commercial test kits(Nanjing Jian Chen Instituteof Biological Technology,Nanjing,China)except for ROS prod

198、uced by Shanghai MlbioInstitute of Biological Technology(Shanghai,China).2.6.Statistical AnalysisAll statistical analyses were carried out using the GLM procedure(SAS Inst.Inc.,Cary,NC,USA).The model included the fixed effects of age(YCC and OCC),rice strawlevels(LRS and HRS),and the interaction bet

199、ween age and rice straw levels.Treatmentmeans were determined using the LSMEANS option and separated using F-test protectedLSD(p0.05).Differences were considered statistically significant when p0.05,while0.05 p0.10 was identified as a tendency.3.Results3.1.Growth PerformanceThe effects of age and st

200、raw levels on total intake,apparent digestibility of nutrientsand body weight results are shown in Table 2.Initial body weight was similar for theexperimental animals.Final BW,total BW gain,average daily gain(ADG)and gain:feedintake(G:F)were higher(p 0.05)in the YCC group than in the OCC group.Total

201、 digestiblenutrients(TDN)intake,final BW and ADG were tended to be higher(p 0.10)in the HRSgroup than in the LRS group,but the HRS group had slightly lower(p=0.06)OM apparentdigestibility than the LRS group.Total DMI was higher(p=0.03)in the HRS group than inthe LRS group.However,the NDF apparent di

202、gestibility was negatively affected(p=0.01)by increased straw.Table 2.Effects of age and straw inclusion levels on growth performance of Yiling cull cows1.Items2YCCOCCp-ValueLRSHRSLRSHRSSEMAgeDietAgeDietIntake(kg/d,on DM basis)Concentrate3.734.443.443.540.320.100.230.36DM4.365.554.094.620.330.110.03

203、0.35TDN3.354.163.123.410.270.100.070.35Apparent digestibility of nutrients(%)OM70.964.370.662.93.560.820.060.88CP60.958.459.653.94.590.540.390.74EE85.181.278.575.93.430.110.360.85NDF62.454.864.652.93.350.980.010.55Body weight(BW,kg)Intial BW3303333313299.40.85Final BW39042934836213.80.0020.080.36Tot

204、al BWgain55.290.422.133.013.70.0060.120.39ADG(g/d)182.0281.874.4113.834.70.0020.070.41G:F(g/kg)42.649.914.023.47.20.0050.280.891YCC,younger cull cows;OCC,older cull cows;LRS,providing 0.7 kg/d rice straw per head;HRS,providing1.2 kg/d rice straw per head.2DM,dry matter;TDN,total digestible nutrients

205、;TDN(kg)=ME(Mcal)/3.62;OM,organic matter;CP,crude protein;EE,ether extract;NDF,neutral detergent fiber;ADG,average daily gain;G:F,gain:feed intake.3.2.Carcass CharacteristicsThe effects of age and straw levels on carcass characteristics of Yiling cull cows areshown in Table 3.Both YCC group and HRS

206、group had higher(p 0.05)differences were observedbetween both age classes and straw levels for dressing percentage and carcass composition.The area of LM between the 6th and 7th as well as between the 12th and 13th ribs were notaffected(p 0.05)by straw levels.Both age and diet had no effect(p 0.05)o

207、n the LM areabetween the 6th and 7th ribs,but the LM area between the 12th and 13th ribs tended to be-73-Animals 2021,11,17326 of 13higher(p=0.07)in YCC than in OCC.High rib and tenderloin were heavier(p 0.05)betweenexperimental groups(Table 4).Cooking loss was higher(p=0.02)in the LRS group than in

208、the HRS group,but not affected(p 0.05)by age.Table 4.Effects of age and straw inclusion levels on meat quality of LM1from Yiling cull cows2.Items3YCCOCCp-ValueLRSHRSLRSHRSSEMAgeDietAgeDietpH,day 06.226.266.116.370.141.000.330.47pH,day 75.665.625.775.740.110.320.740.99DM(%)30.934.334.232.21.430.680.6

209、10.08IMF(%)21.931.028.528.83.960.600.250.28CP(%)72.562.566.366.24.400.780.270.28WBSF(N)26.931.631.229.73.230.710.630.36Drip loss(%)6.565.516.455.871.330.930.560.86Pressing loss(%)30.532.431.431.31.600.980.570.53Cooking loss(%)26.924.527.525.20.860.480.021.001LM,longissimus dorsi muscle.2YCC,younger

210、cull cows;OCC,older cull cows;LRS,providing 0.7 kg/d ricestrawper head;HRS,providing 1.2 kg/d rice straw per head.3DM,dry matter;IMF,intramuscular fat,based onDM basis;CP,crude protein,based on DM basis;WBSF,Warner-Bratzler shear force.3.4.Fatty Acid ProfileFatty acid profile results are in Table 5.

211、The interactions between age and straw levels(p 0.05)were observed for the proportions of C15:0,C16:0,C17:0,C18:1n9c,C20:5n3c,SFAs,and MUFAs.Decreased C15:0,C17:0,C20:5n3c,and SFAs proportion as well asincreased C18:1n9c and MUFAs proportion were observed in YCC with HRS diet ascompared to YCC with

212、LRS diet(p 0.05)in older cows between the diet groups.The proportion of C18:3n3cwas higher(p=0.04)in the HRS group than in the LRS group.Table 5.Effects of age and straw inclusion levels on fatty acid profile(%,of total fatty acids)of LM1from Yiling cull cows2.Items3YCCOCCp-ValueLRSHRSLRSHRSSEMAgeDi

213、etAgeDietSaturatedC14:03.043.283.283.660.290.310.300.81C15:00.29 a0.21 b0.21 b0.23 b0.0160.130.100.006C16:028.9526.4826.7728.640.940.990.750.04C17:00.80 a0.59 b0.59 b0.62 b0.0360.030.030.006C18:012.2510.0811.2210.800.790.850.120.29C20:00.110.100.100.100.0060.670.180.48C21:00.050.050.060.040.0080.930

214、.100.12C22:00.060.030.040.040.0100.700.160.32MonounsaturatedC14:1n5c1.191.441.361.350.160.800.470.42C15:1n5c0.220.150.140.150.0450.340.520.40C16:1n7c4.014.844.874.980.370.200.220.35C17:1n7c0.690.620.570.610.0460.160.720.27C18:1n9t0.470.480.740.380.0950.390.080.07C18:1n9c43.66 b48.19 a46.33 ab44.19 a

215、b1.350.630.390.03C22:1n9c0.820.380.450.530.140.480.220.09C24:1n9c0.090.050.050.070.0170.570.650.15PolyunsaturatedC18:2n6c2.492.332.512.890.370.440.760.47C18:3n6c0.080.060.060.070.0140.880.560.36C18:3n3c0.190.250.230.230.0150.670.040.08C20:2n6c0.040.050.070.040.0110.480.460.16C20:3n6c0.220.170.180.17

216、0.0460.680.520.69C20:3n3c0.020.020.020.020.0030.770.450.30C20:4n6c0.030.020.020.030.0050.480.660.09C20:5n3c0.06a0.03b0.03b0.05ab0.0080.360.510.02C22:6n3c0.160.100.090.110.0270.230.480.11SFA45.57 a40.82 b42.28 ab44.14 ab1.460.990.340.04MUFA51.14 b56.15 a54.52 ab52.25 b1.310.850.320.02PUFA3.293.033.20

217、3.610.450.600.870.47PUFA/MUFA0.060.050.060.070.0090.540.920.24n-6 PUFA2.852.632.833.200.410.520.870.49n-3 PUFA0.440.400.360.410.0420.440.890.32n-6/n-3PUFA6.506.507.877.570.630.070.810.81ab Means followed by different letters in the same row are significant at the p 0.05)between the experimental grou

218、ps(Table 6).TheCAT activity of liver tissue was higher(p=0.045)in the YCC group than in the OCC group,but was unaffected(p 0.05)by straw levels.Moreover,the SOD activity of muscle tissuewas higher(p=0.01)in the LRS group than in the HRS group,but similar(p 0.05)betweenage groups.-75-Animals 2021,11,

219、17328 of 13Table 6.Effects of age and straw inclusion levels on antioxidant status of tissues from Yilingcull cows1.Items2YCCOCCSEMp-ValueLRSHRSLRSHRSAgeDietAgeDietT-AOC(umol Trolox/Mgprot)Liver0.280.260.260.210.020.110.150.61Muscle1.171.251.121.180.100.550.480.91CAT(U/mgprot)Liver4.783.853.293.280.

220、450.0450.330.34Muscle3.164.594.286.531.140.200.130.73SOD(U/mgprot)Liver39638237038017.00.420.920.50Muscle136511891315117852.50.570.010.71ROS(U/mgprot)Liver13.212.913.613.20.620.570.630.99Muscle122.7116.4124.3119.75.070.630.300.87MDA(nmol/mgprot)Liver2.502.272.362.430.130.930.530.25Muscle17.216.715.9

221、16.50.990.430.970.59PC(nmol/mgprot)Liver12.211.410.211.41.230.430.870.43Muscle53.466.957.245.37.370.250.920.111YCC,younger cull cows;OCC,older cull cows;LRS,providing 0.7 kg/d rice straw per head;HRS,providing1.2 kg/d rice straw per head.2T-AOC:total antioxidative capacity;CAT,catalase;SOD:superoxid

222、e dismutase;ROS,reactive oxygen species.MDA:malondialdehyde;PC:protein carbonyl.4.DiscussionOne of the most important parameters of beef production is the growth ability ofanimals.Growth in young cattle is unequal and influenced by age and diet.In the currentstudy,younger cows showed higher final BW

223、,total BW gain,and average daily gain,which is supported by the study of Sawyer et al.(2004)28,who found that DMI andADG decreased linearly with the increased age of the cull cows.However,the results ingrowth performance are opposite to the finding of Galli et al.(2008)29,who reportedthat younger co

224、ws finished under grazing conditions had lower the final BW than oldercows.The increased DMI with forage inclusion levels in the diet could be supported byGalyean and Defoor(2003)30.They reported a positive linear relationship(R2=0.92)between forage NDF and DMI(%BW),and also claimed that rumen and g

225、ut fill doesnot limit intake when ruminants are fed high concentrate diets and the mechanism forenhanced intake is energy dilution and to maintain energy intake.However,we believethe changed intake is more likely due to the more balanced ruminal environment andthe greater ruminal pH value,particular

226、ly in high concentrate diet,caused by enhancedchewing activity and saliva flow with increasing forage inclusion 7.In the present study,calculated by the proportions of total concentrate and straw intake in total DMI duringfinishing period,the percentages of forage in diet groups were 15.2%and 21.7%(

227、LRS vs.HRS),respectively.Hales et al.(2013)31 compared different levels of forage inclusion(2%,6%,10%,or 14%of alfalfa hay)and showed a quadratic effect of increasing forageproportion on DMI during the whole finishing period in steers.They further reported thatan increase in DMI with alfalfa hay inc

228、lusion of up to 10%,and then a decrease in DMIwith 14%alfalfa hay.A previous study by Swanson et al.(2017)32 reported that ADGand DMI decreased linearly with increasing forage inclusion.The inconsistency amongstudies may be due to the differences in breed or forage type.The apparent digestibility of

229、OM(slightly)and NDF decreased with straw levels,which is consistent with the result ofSalinas-Chavira et al.(2013)33.Generally,Chinese south native cattle were used as draftanimals and had not undergone long-term commercial selection.Thus,we speculated thatYiling cattle may be intolerant to forage r

230、estriction diet that could cause intake inhibition.-76-Animals 2021,11,17329 of 13In terms of feed intake,daily gain and feed conversion efficiency,this fattening strategyseems less efficient in resource utilization.The inefficiency may be mainly due to the factthat the experimental cattle were matu

231、re cows and the increase in BW comes from fatdeposition rather than the growth and development of bones and muscles.Although thereis no control group(no fattening)in the current study,the production experience of thefarm has proved that long-term fattening can increase the marbling richness of beef

232、fromYiling cull cows.High marbled beef,especially from native cattle,can fetch high prices atthe market,which can support the reasonableness of this fattening strategy.Younger cull cows and cows fed higher straw levels had heavier carcass that couldbe justified by their higher final BW.A recent stud

233、y reported that the proportion of fat incarcasses increased,and the relative proportion of muscles decreased as animals becomeolder 34.However,fat percentage in carcass was similar between age groups in thecurrent study.Dressing percentage is an important indicator in the evaluation of carcasscharac

234、teristics,and the rise in dressing percentage is a direct result of increasing fatnesswith slaughter weight 35.Thus,similar dressing percentage between experimental groupswas due to unchanged fat contents in carcass in the current study.In addition,heavierstriploin found in cows fed with higher stra

235、w levels and high rib found in younger cowswere related to higher carcass weight.Taken together,the present results may not only beuseful for further studies,but also for the sustainable and profitable beef production fromcull cows.In the current study,no significant significance,including the effec

236、ts of age and dietand their interaction,was observed for IMF deposition.However,the mean value of IMFcontent in younger cows fed higher straw levels was 41.92%higher than in younger cowsfed lower straw levels(31.0%vs.21.9%).By contrast,the numerical difference in IMFcontent of older cows between die

237、t groups was small(28.8%vs.28.5%),which may indicatethat IMF deposition is insensitive to the change of energy intake for older cows.It is knownthat the tenderness of beef from cull cows decreased with increasing age 1,2.However,theshear force was unaffected by age in this study.A recent study 36 re

238、ported that the shearforce of beef from cull cows was lower than that from heifers when fed with a high-energyration for 150 d,which indicates that the high IMF content induced by long-term fatteningcould weaken the effect of connective tissue properties on tenderness.Drip loss,pressingloss as well

239、as cooking loss can describe the water-holding capacity of beef and reflectdifferent characteristics.Galli et al.(2008)29 reported that cooking loss was unchangedwith the increasing age of cull cows,which is consistent with our results.However,anotherrecent study 37 showed that age altered the cooki

240、ng loss of beef from cull cows and thisinfluence depends on different beef cuts.In the current study,dietary treatment has noeffect on the physical properties of beef except for cooking loss.The decreased cookingloss caused by high-level straw diet may lead to inferior juiciness.Fatty acid compositi

241、on can vary dramatically in beef depending on several factors,such as breed,sex,age,and diet 38.Fruet et al.(2018)39 reported that higher C18:1n9cand MUFAs proportion as well as lower C20:5n3c and SFAs proportion were found inthe high concentrate diet treatment.Moreover,Wang et al.(2019)27 reported

242、thatthe proportions of C18:1n9c and MUFAs rose with dietary energy.In the current study,younger cows fed higher straw levels had higher C18:1n9c and MUFAs proportion as wellas lower C20:5n3c and SFAs proportion than younger cows fed lower straw levels,whichcould be associated with the enhancement of

243、 energy intake.Although total TDN intake oftwo age classes both increased with straw levels,the effects of diet on the proportions ofC15:0,C17:0,C18:1n9c,C20:5n3c,SFAs,and MUFAs were more pronounced in youngercows than in older cows.Several studies have demonstrated that the fatty acid profilewas st

244、rongly affected by IMF content 40,41.Thus,the interaction between age and dietfor those fatty acids may partially come from their numerical difference in IMF content.Cho et al.(2013)42 reported that the proportions of C18:3n3c,C18:3n6c,SFA,and n-3PUFAs increased and MUFAs decreased in striploin as t

245、he cow age increased.However,inthis study,differences were only detected for the slightly higher n-6/n-3 PUFAs ratio and-77-Animals 2021,11,173210 of 13greater C18:3n3c proportion in older cows.Reasons for the inconsistency in these resultsmay be related to the differences in age classes.Oxidative s

246、tress caused by the increased production of ROS or a decrease in antiox-idant capacity,results in damage to biological macromolecules as well as disruption ofnormal metabolism and physiology 43.Moreover,oxidative stress plays a key role inthe pathogenesis of diverse diseases in cattle 44,45.Antioxid

247、ases CAT,SOD,and glu-tathione peroxidase(GSH-Px)are part of enzymatic antioxidant systems that protect tissuecomponents against oxidative stress caused by ROS.The concentration of MDA reflectsthe extent of lipid oxidation,which has a negative impact on meat freshness and quality,including undesirabl

248、e off-flavor,toxic substances,and discoloration 46.Protein carbonyl(PC)is a biomarker of protein oxidative damage,which could decrease the water-holdingcapacity and tenderness of the meat 47.Halliwell(1994)48 reported that animal age in-duced the oxidative damage to cellular macromolecules,such as l

249、ipids,proteins,and DNA.Moreover,Cho et al.(2015)20 reported that oxidative deterioration on d2 post-slaughterwas accelerated with older age,despite the increased activity of antioxidant enzymes.En-hanced activities of antioxidant enzymes during aging were the consequences of increasedexpression of t

250、he mRNA of antioxidant enzymes 49.However,only the CAT of livertissue was different between age groups in this study,and its activities were higher inyounger cows than in older cows.In addition,the antioxidant parameters were unaffectedby straw amounts except for SOD.Antioxidase CAT and SOD are coup

251、led enzymes 50,but the activity of CAT and SOD in both liver and muscle tissues did not exhibit the samepattern in our results,most likely due to the additional effect of dietary stress in our study,including high energy intake and abnormal level of dietary forage.A high concentration ofunsaturated

252、fatty acids,particularly PUFAs,accelerates the lipid oxidation process 51.Inthe current study,the unsaturation degree in meat was unaffected by animal age and strawamounts,which may be related to the similarity of MDA concentration in muscle betweenexperimental groups.The results that T-AOC,ROS,PC a

253、nd MDA remain unchangedindicate that the antioxidant status under high-concentration diets may be less affected byage and diet.5.ConclusionsBoth age and rice straw inclusion levels had impact on the growth performanceand carcass traits of Yiling cull cow.Younger age increased ADG,BW gain,and feedcon

254、version efficiency,while higher straw amounts increased DM intake and decreasedNDF digestibility.The change of fatty acid composition caused by dietary rice straw levelswas more pronounced in younger cows than in older cows.Younger cows fed higherrice straw levels had lower C15:0,C17:0,C20:5n3c,and

255、SFAs proportion as well as higherC18:1n9c and MUFAs proportion compared to younger cows fed lower rice straw levels.Older age decreased the CAT activity of liver tissue,while higher straw amounts reducedthe SOD activity of muscle tissue.Thus,selecting according to their age and providingadequate for

256、age feed for Yiling cull cows would be of greater finishing benefit.Author Contributions:Conceptualization,X.Q.(Xinjun Qiu),X.Q.(Xiaoli Qin),B.C.and H.S.;resources,L.C.;investigation,L.C.;data curation,X.Q.(Xinjun Qiu),Q.Q.and H.W.writingreviewand editing,Q.Q.,M.A.u.R.,B.C.and H.S.;project administr

257、ation,B.C.and H.S.;funding acquisition,B.C.and H.S.All authors have read and agreed to the published version of the manuscript.Funding:This research was funded by the National Key R&D Program of China(2018YFD05018000);the Key Technology R&D Program of Ningxia(2017BY078);Demonstration Project of Expl

258、oitationand Utilization of High-Quality Silage and Roughage Resources(16200157);and China AgricultureResearch System(CARS-37).Institutional Review Board Statement:The study was conducted according to the guidelines ofthe Declaration of Helsinki,and approved by the Animal Welfare and Ethics Committee

259、 of ChinaAgricultural University(Permit No.DK1008).-78-Animals 2021,11,173211 of 13Acknowledgments:We are thankful for the help from staff at the Hubei Fulljoywo DevelopmentCompany Limited during the feeding period.We are also grateful to Xiaojie Su,farm departmentmanager,Xiuzhong Wu,slaughter depar

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297、at Sci.2016,111,18.CrossRef-81-4.926ArticleSerum Biochemical Parameters,Rumen Fermentation,and RumenBacterial Communities Are PartlyDriven by the Breed and Sex ofCattle When Fed High-Grain DietXinjun Qiu,Xiaoli Qin,Liming Chen,Zhiming Chen,Rikang Hao,Siyu Zhang,Shunran Yang,Lina Wang,Yafang Cui,Ying

298、qi Li et al.https:/doi.org/10.3390/microorganisms10020323-82-?Citation:Qiu,X.;Qin,X.;Chen,L.;Chen,Z.;Hao,R.;Zhang,S.;Yang,S.;Wang,L.;Cui,Y.;Li,Y.;et al.SerumBiochemical Parameters,RumenFermentation,and Rumen BacterialCommunities Are Partly Driven bythe Breed and Sex of Cattle When FedHigh-Grain Diet

299、.Microorganisms2022,10,323.https:/doi.org/10.3390/microorganisms10020323Academic Editor:Francois BlachierReceived:19 November 2021Accepted:6 January 2022Published:30 January 2022Publishers Note:MDPI stays neutralwith regard to jurisdictional claims inpublished maps and institutional affil-iations.Co

300、pyright:2022 by the authors.Licensee MDPI,Basel,Switzerland.This article is an open access articledistributedunderthetermsandconditions of the Creative CommonsAttribution(CC BY)license(https:/creativecommons.org/licenses/by/4.0/).microorganismsArticleSerum Biochemical Parameters,Rumen Fermentation,a

301、ndRumen Bacterial Communities Are Partly Driven by the Breedand Sex of Cattle When Fed High-Grain DietXinjun Qiu1,Xiaoli Qin1,Liming Chen2,Zhiming Chen1,Rikang Hao1,Siyu Zhang1,Shunran Yang1,Lina Wang1,Yafang Cui1,Yingqi Li1,Yiheng Ma1,Binghai Cao1,*and Huawei Su1,*1State Key Laboratory of Animal Nu

302、trition,College of Animal Science and Technology,China Agricultural University,Beijing 100000,China;(X.Q.);(X.Q.);(Z.C.);(R.H.);(S.Z.);(S.Y.);(L.W.);(Y.C.);(Y.L.);(Y.M.)2Hubei Fulljoywo Agricultural Development Company Limited,Yichang 443000,China;*Correspondence:(B.C.);(H.S.);Tel.:+86-010-6273-3850

303、(B.C.);+86-010-6281-4346(H.S.)Abstract:Hybridization in bovines is practiced with the main aim of improving production perfor-mance,which may imply the microbial variations in the rumen from the parental breed cross totheir progeny.Besides,the interactions of offspring breed with sex in terms of rum

304、en bacteria arenot clear.This study aims to evaluate the variations in rumen bacterial communities in differentbreeds and sexes,and the correlations among fattening performance,serum biochemical parame-ters,and rumen fermentation.Forty-two 19.20.67-month-old beef cattle(39095 kg of initialbody weigh

305、t)comprising two genetic lines(Yiling and AngusYiling)and two sexes(heifers andsteers)were raised under the same high-grain diet for 120 d.On the last two days,blood sampleswere collected from each animal via the jugular vein before morning feeding for analyzing serumbiochemical parameters;rumen flu

306、id samples were obtained via esophageal intubation 2 h aftermorning feeding for analyzing rumen fermentation parameters and bacterial communities.Theresults show that both breed and sex had a certain impact on fattening performance,serum bio-chemical parameters,and rumen fermentation.No differences

307、in the diversity and structure ofrumen bacterial communities were observed.Significant interactions(p 0.05)of breed and sex wereobserved forSuccinivibrionaceae UCG-002and Prevotellaceae UCG-001.The relative abundances of theRikenellaceae RC9 gut group,Prevotellaceae UCG-003,and Succinivibrio were di

308、fferent(p 0.05)betweenbreeds.Heifers had a higher(p=0.008)relative abundance of the Rikenellaceae RC9 gut group thansteers.Correlation analysis showed a significant relationship(p 0.05)of rumen bacteria with serumbiochemical parameters,rumen pH,and rumen fermentation patterns.Additionally,only two g

309、enera,Prevotellaceae UCG-003 and Prevotellaceae UCG-001,had positive correlations with feed efficiency.Inconclusion,serum biochemical parameters,rumen fermentation,and rumen bacterial communitiesare partly driven by the breed and sex of cattle fed a high-grain diet.Keywords:breed;fattening performan

310、ce;rumen bacteria;rumen fermentation;serum parame-ters;sex1.IntroductionFifty-five indigenous bovine breeds with nearly 30 million animals have been observedin China 1.In general,they are characterized by a small size,slow growth,inferiordressing percentage,etc.These characteristics have hindered th

311、e current beef market.TheYiling(YL)breed is typically raised in the Yichang district,Hubei province.It was formerlyselected as a draught animal,but now this breed and its hybrids are bred only for beefMicroorganisms 2022,10,323.https:/doi.org/10.3390/microorganisms10020323https:/ 2022,10,3232 of 13p

312、roduction,especially for high marbling beef.A previous study 2 evaluated its geneticbackground.Furthermore,the finishing performance of YL cull cows was evaluated inour previous feeding experiment 3.As expected,it presented an inferior performance indaily gain and feed efficiency.In order to protect

313、 and utilize this genetic resource,YL cattleand its hybrids need to be further evaluated to provide a scientific basis for its breedingand industrialization.Ruminants have evolved a complex and diverse symbiotic microbiota consisting ofbacteria,archaea,protozoa,fungi,and viruses in their rumen 4.In

314、particular,rumen bacte-ria are the most abundant microbiota in terms of diversity and account for the vast majorityof the microbiome.Additionally,they have been the focus of most quantitative studies onrumen microbial composition.These microbes of ruminants can typically degrade plantfibers and util

315、ize non-protein nitrogen to produce volatile fatty acids(VFAs)and microbialproteins,further meeting the hosts nutrient requirements for maintenance and production.Our recent efforts indicated that the differences in rumen bacterial communities wereparticularly associated with diet,including forage i

316、nclusion 5,6,energy levels 7,proteinlevels 8,and even nutrient density 9.These studies also confirmed the correlations ofrumen bacterial abundances with the hosts phenotypic characteristics,such as nutrientintake 6,9,rumen fermentation products 6,7,9,nutrient apparent digestibility 6,9,bloodmetaboli

317、tes 6,9,and meat fatty acids 7.Rumen microbiota contributes to the hostsnutrient availability and subsequently exerts a potential impact on production performance.In this sense,it is effective to establish the interactions among diet,rumen microbiota,andphenotypic characteristics in ruminants.Recent

318、 genome-wide association studies revealedthat the composition of rumen microbiota can be affected by host additive genetics orgenotypes at multiple taxonomic levels 1012.Furthermore,heritable rumen microbialfeatures are associated with rumen metabolites 10,12,13,feed efficiency 10,and milkquality 12

319、,13.These observations more strongly confirm another notion of a triangularrelationship among the host genetics,rumen microbiota,and phenotypic characteristics.Several studies have explored the rumen microbial differences driven by differentcross combinations.For instance,Li et al.14 reported that t

320、he microbiota and metabolitesin the rumen were largely affected by different hybrid crosses between sika deer andelk;Bainbridge et al.15 reported that rumen bacterial communities were less affectedby dairy breeds(Holstein,Jersey,and HolsteinJersey)when compared with lactationdays;Hernandez-Sanabria

321、et al.16 and Roehe et al.17 found a significant effect ofsired beef breed on rumen bacterial and archaeal communities.These results imply thepotential differences in rumen bacterial communities between purebred and crossbredbreeds.However,the interaction with sex was not explored in these studies.He

322、re,we compared the differences in rumen bacterial communities driven by thebreed(YL vs.AngusYL(AY)and sex(steers vs.heifers)of cattle fed the same high-graindiet.We also analyzed the correlations of rumen bacterial communities with fatteningperformance,serum biochemical parameters,and rumen fermenta

323、tion.We hypothesizedthat breed may impact the fattening performance and bacterial communities of cattleregardless of sex.It should be noted that this breed factor can also be further defined asa different sired breed(YL vs.Angus)in the current study.Thus,the improvement infattening performance and e

324、ven the differences in the rumen bacterial communities ofcrossing progenies mainly derive from the transmission of superior traits from sires 16.Inthis sense,the particular rumen bacterial communities of crossing progenies could be usedas a reference for improving the productivity of Chinese indigen

325、ous cattle.2.Materials and Methods2.1.Ethics StatementAnimal studies were performed in accordance with institutional guidelines and theapproval of the Animal Care and Use Committee of China Agricultural University(PermitNo.AW09059102-3,6 September 2017).-84-Microorganisms 2022,10,3233 of 132.2.Anima

326、ls,Management,and SamplingBefore the trial,all cattle were weaned at 4 months old and castrated at 5 monthsold and subsequently received the same diet and management.Forty-two cattle aged19.20.67 months were selected and fed the same total mixed ration(TMR)for the 120 dfattening trial.These cattle c

327、omprised two genetic lines and two sexes:YL heifers(n=10)and steers(n=10);AngusYL(AY,siredam)heifers(n=9),and steers(n=13).Allanimals were reared in separate pens with ad libitum access to TMR based on 5 to 10%orts.Experimental TMR(11.4 MJ/kg metabolic energy,12.0%crude protein)was formed by20.0%cor

328、n silage,6.63%rice straw,and 73.3%concentrate composed of corn grain,wheatbran,rapeseed cake,soybean meal,limestone,premix,NaHCO3,and NaCl.The feedingfrequency of TMR was twice a day at 08:00 h and 16:00 h.Feed provided and residue were recorded daily to calculate the average dry matterintake(DMI)du

329、ring the whole fattening period.Body weight(BW)was recorded beforemorning feeding for 3 consecutive days.Average daily gain(ADG)was calculated basedon the difference between initial body weight and final body weight.The last 7 days of thefattening period were the sampling phase.Blood samples were co

330、llected from each animalvia the jugular vein before morning feeding and then centrifuged at 3500g for 10 minto obtain serum and subsequently stored at80C until serum biochemical parametersanalysis.A total of 41 rumen fluid samples were collected via esophageal intubation 2 hafter morning feeding(dis

331、carded one sample polluted by saliva).The first 200 mL of rumenfluid samples was discarded to minimize contamination from the saliva.The pH value ofrumen fluid samples was measured immediately.Then,rumen fluid samples were filteredusing four layers of sterile cheesecloth.Two aliquots were stored at8

332、0C for VFA andammonia-N analysis,respectively.Another two aliquots were stored at80C for 16SrRNA pyrosequencing.2.3.Chemical AnalysisGlucose(GLU),triglyceride(TG),cholesterol(CHO),non-esterified fatty acid(NEFA),beta-hydroxybutyrate(BHB),high-density lipoprotein cholesterol(HDL-C),low-densitylipopro

333、tein cholesterol(LDL-C),creatinine(CREA),urea(UREA),aspartate aminotrans-ferase(AST),alanine aminotransferase(ALT),alkaline phosphatase(ALP),total protein(TP),and albumin(ALB)concentrations were determined using commercial test kits(Bei-jing Jiuqiang Bio-Technique Co.Ltd.,Beijing,China)with an automated biochemistryanalyzer(Hitachi 7020;Hitachi Ltd.,Tokyo,Japan).The ammonia-N concentration ofrumen