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| 不同抗冻保护剂对鸡精液冷冻效果及基因表达的影响 |
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李博宇1,郭勇1,倪和民1,盛熙晖1,陈余2,王梁2,王相国1,肖龙菲1,邢凯1*,齐晓龙1
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| (1.北京农学院 动物科学技术学院, 北京 102206;2.北京市畜牧总站, 北京 100107) |
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| 摘要: |
| 旨在研究不同抗冻保护剂对鸡精液的冷冻保存效果，并通过比较差异表达基因，以解释不同保护剂的抗冻机制。以海兰褐种公鸡为研究对象，检测了不同抗冻保护剂(保护剂Ⅰ和保护剂Ⅱ)冷冻保存后的精子活性，并利用Affymetrix表达谱芯片检测技术，对差异表达相关基因进行筛选。结果表明:1)保护剂Ⅰ组的精液冻后活力活率(27.05±1.22 vs 47.65±5.89)显著高于保护剂Ⅱ组(20.65±1.50 vs 35.15±5.15)(P<0.05)；2)鸡精液冷冻后，2个不同抗冻保护剂处理组共发现1 604个差异表达基因。保护剂Ⅰ组中显著上调的基因1 380个，显著下调的基因224个；3)基因本体(GO)分析表明，包括核糖体亚基、线粒体呼吸链、泛素蛋白连接酶结合和微管蛋白结合，翻译起始以及RNA分解代谢等过程都与精液抗冻性相关；4)KEGG通路富集分析发现，其主要富集在内质网应激通路、氧化磷酸化、蛋白质泛素化、核糖体调控以及线粒体呼吸调控等相关信号通路上；5)挑选CCND1、HSP70、RHOA、HSP90和CIRBP 5个差异表达基因并利用荧光定量PCR对芯片结果进行验证，二者吻合。综上，初步研究比较了2种精液保护剂的冷冻效果，并筛选出差异表达基因，发现抗冻保护剂Ⅰ更适合海兰褐种公鸡冷冻精液的保存，为鸡精液冷冻技术提供参考，并为鸡精液抗冻分子机制解析提供方向。 |
| 关键词: 鸡精液 抗冻保护剂 表达谱芯片 差异表达基因 |
| DOI：10.11841/j.issn.1007-4333.2021.08.13 |
| 投稿时间：2020-10-15 |
| 基金项目:“十三五”国家重点研发计划(2016YFD0700201)；现代农业产业技术体系北京市家禽创新团队项目(BAIC04-2020)；2020年科技创新服务能力建设-农业科技新星计划(20200210) |
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| Effects of different cryoprotectants on the freezing effect and gene expression of chicken sperm |
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LI Boyu1,GUO Yong1,NI Hemin1,SHENG Xihui1,CHEN Yu2,WANG Liang2,WANG Xiangguo1,XIAO Longfei1,XING Kai1*,QI Xiaolong1
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| (1.Animal Science and Technology College, Beijing University of Agriculture, Beijing 102206, China;2.Beijing General Station of Animal Husbandry Service, Beijing 100107, China) |
| Abstract: |
| To explore the antifreezing effect of different cryoprotectants on chicken semen, and explain the antifreeze mechanism of different cryoprotectants by comparing the differentially expressed genes, Hy-line Breeder Cocks were taken as study objects to invesitgate the activity of sperm preserved in different cryoprotectants(cryoprotectant Ⅰ and cryoprotectant Ⅱ), and the differentially expressed genes were screened by Affymetrix microarray. The results showed that: 1)The sperm viability of group Ⅰ(27. 05±1. 22 vs 47. 65±5. 89)was significantly higher than that of group Ⅱ(20. 65±1. 50 vs 35. 15±5. 15)(P<0. 05); 2)A total of 1 604 differentially expressed genes were found in the two treatment groups. In group Ⅰ, 1 380 genes were significantly up-regulated and 224 genes were significantly down-regulated; 3)Gene ontology analysis showed that the processes including ribosomal subunits, mitochondrial respiratory chain, ubiquitin protein ligase binding, and tubulin binding, translation initiation and RNA catabolism were related to semen antifreeze; 4)KEGG pathway enrichment analysis found that it was mainly enriched in endoplasmic reticulum stress pathway, oxidative phosphorylation, protein ubiquitination, ribosome regulation and mitochondrial respiratory regulation and other related signal pathways. 5)Four differentially expressed genes of CCND1, HSP70, HSP90 and CIRBP were selected and verified by fluorescence quantitative PCR. To sum up, the preliminary study compared the freezing effects of two cryoprotectants and screened out differentially expressed genes, and found that the cryoprotectant I was more suitable for the preservation of frozen semen of Hy-line Breeder Cocks, which provided references for chicken semen freezing technology and direction for the molecular mechanism analysis of chicken sperm antifreeze. |
| Key words: chicken sperm cryoprotectants differentially expressed gene expression profiling chip |
