【目的】基于高叶酸小鼠睾丸转录组数据,筛选叶酸调控牛和小鼠繁殖力的同源基因,以期为通过叶酸改善雄性动物繁殖力奠定理论基础。【方法】选取24只健康、生长状况相近的11周龄C57BL/6雄性小鼠,随机分为3组(n=8):CON、A和B组,分别饲喂2...【目的】基于高叶酸小鼠睾丸转录组数据,筛选叶酸调控牛和小鼠繁殖力的同源基因,以期为通过叶酸改善雄性动物繁殖力奠定理论基础。【方法】选取24只健康、生长状况相近的11周龄C57BL/6雄性小鼠,随机分为3组(n=8):CON、A和B组,分别饲喂2、7和15 mg/kg叶酸,连续饲喂57 d。饲喂结束后分别测定3组小鼠左、右睾丸和精囊的重量,精子密度以及缓慢前进的精子比例,通过Illumina NovaseqTM6000高通量测序进行小鼠睾丸转录组测序分析,对测序结果进行差异表达基因(DEGs)、GO功能注释和蛋白互作网络(PPI)分析,筛选叶酸调控精液品质和繁殖力的关键基因,并结合荷斯坦种公牛繁殖力相关同源基因和数量性状基因座(QTLs)数据,进一步筛选叶酸调控牛和小鼠雄性繁殖力的同源基因。【结果】相较于CON组,B组小鼠左、右精囊重量均极显著增加(P<0.01),A和B组小鼠精液中缓慢前进的精子比例均极显著降低(P<0.01)。经RNA-Seq分析发现,在A vs CON组中筛选出423个DEGs,在B vs CON组中筛选出661个DEGs,A vs CON和B vs CON组中共有的上调DEGs 85个、下调DEGs 40个。通过GO功能注释分析发现,A vs CON组的DEGs显著富集在叶酸代谢和精液品质调控相关的GO条目,B vs CON组的DEGs显著富集在叶酸代谢和生殖器官发育相关的GO条目,A vs CON和B vs CON两个组共有的上调DEGs显著富集在叶酸代谢和精液品质调控相关等功能,共有的下调DEGs显著富集在一碳代谢过程和单精受精作用功能。通过DEGs的PPI网络分析发现,筛选出的核心节点(hub)基因可能通过影响精液品质而影响雄性动物繁殖力。同时本研究中筛选出的hub基因均为奶牛的同源基因,并找出了不同浓度叶酸影响荷斯坦种公牛繁殖力的同源基因及其附近的QTLs,包括每次妊娠的授精次数QTL、精子活力QTL等。将hub基因进一步与前人荷斯坦种公牛精子转录组差异表达基因进行比对,�展开更多
Sperm quality is evaluated for the calculation of sperm dosage in artificial reproductive programs. The most common parameter used is motility, but morphology has a higher potential as a predictor of genetic quality. ...Sperm quality is evaluated for the calculation of sperm dosage in artificial reproductive programs. The most common parameter used is motility, but morphology has a higher potential as a predictor of genetic quality. Morphometry calculations from CASA-Morph technology improve morphological evaluation and allow mathematical approaches to the problem. Semen from 28 Holstein bulls was collected by artificial vagina, and several ejaculates were studied. After general evaluation, samples were diluted, packaged in 0.25 ml straws, and stored in liquid nitrogen. Two straws per sample were thawed, and slides were processed and stained with Diff-Quik. Samples were analyzed by a CASA-Morph system for eight morphometric parameters. In addition to the ＂classical＂ statistical approach, based on variance analysis （revealing differences between animals, ejaculates, and straws）, principal component （PC） analysis showed that the variables were grouped into PC1, related to size, and PC2 to shape. Subpopulation structure analysis showed four groups, namely, big, small, short, and narrow from their dominant characteristics, representing 31.0%, 27.3%, 24.1%, and 17.7% of the total population, respectively. The distributions varied between animals and ejaculates, but between straws, there were no differences in only four animals. This modern approach of considering an ejaculate sperm population as divided into subpopulations reflecting quantifiable parameters generated by CASA-Morph systems technology opens a new view on sperm function. This is the first study applying this approach to evaluate different ejaculates and straws from the same individual. More work must be done to improve seminal dose calculations in assisted reproductive programs.展开更多
文摘【目的】基于高叶酸小鼠睾丸转录组数据,筛选叶酸调控牛和小鼠繁殖力的同源基因,以期为通过叶酸改善雄性动物繁殖力奠定理论基础。【方法】选取24只健康、生长状况相近的11周龄C57BL/6雄性小鼠,随机分为3组(n=8):CON、A和B组,分别饲喂2、7和15 mg/kg叶酸,连续饲喂57 d。饲喂结束后分别测定3组小鼠左、右睾丸和精囊的重量,精子密度以及缓慢前进的精子比例,通过Illumina NovaseqTM6000高通量测序进行小鼠睾丸转录组测序分析,对测序结果进行差异表达基因(DEGs)、GO功能注释和蛋白互作网络(PPI)分析,筛选叶酸调控精液品质和繁殖力的关键基因,并结合荷斯坦种公牛繁殖力相关同源基因和数量性状基因座(QTLs)数据,进一步筛选叶酸调控牛和小鼠雄性繁殖力的同源基因。【结果】相较于CON组,B组小鼠左、右精囊重量均极显著增加(P<0.01),A和B组小鼠精液中缓慢前进的精子比例均极显著降低(P<0.01)。经RNA-Seq分析发现,在A vs CON组中筛选出423个DEGs,在B vs CON组中筛选出661个DEGs,A vs CON和B vs CON组中共有的上调DEGs 85个、下调DEGs 40个。通过GO功能注释分析发现,A vs CON组的DEGs显著富集在叶酸代谢和精液品质调控相关的GO条目,B vs CON组的DEGs显著富集在叶酸代谢和生殖器官发育相关的GO条目,A vs CON和B vs CON两个组共有的上调DEGs显著富集在叶酸代谢和精液品质调控相关等功能,共有的下调DEGs显著富集在一碳代谢过程和单精受精作用功能。通过DEGs的PPI网络分析发现,筛选出的核心节点(hub)基因可能通过影响精液品质而影响雄性动物繁殖力。同时本研究中筛选出的hub基因均为奶牛的同源基因,并找出了不同浓度叶酸影响荷斯坦种公牛繁殖力的同源基因及其附近的QTLs,包括每次妊娠的授精次数QTL、精子活力QTL等。将hub基因进一步与前人荷斯坦种公牛精子转录组差异表达基因进行比对,�
文摘Sperm quality is evaluated for the calculation of sperm dosage in artificial reproductive programs. The most common parameter used is motility, but morphology has a higher potential as a predictor of genetic quality. Morphometry calculations from CASA-Morph technology improve morphological evaluation and allow mathematical approaches to the problem. Semen from 28 Holstein bulls was collected by artificial vagina, and several ejaculates were studied. After general evaluation, samples were diluted, packaged in 0.25 ml straws, and stored in liquid nitrogen. Two straws per sample were thawed, and slides were processed and stained with Diff-Quik. Samples were analyzed by a CASA-Morph system for eight morphometric parameters. In addition to the ＂classical＂ statistical approach, based on variance analysis （revealing differences between animals, ejaculates, and straws）, principal component （PC） analysis showed that the variables were grouped into PC1, related to size, and PC2 to shape. Subpopulation structure analysis showed four groups, namely, big, small, short, and narrow from their dominant characteristics, representing 31.0%, 27.3%, 24.1%, and 17.7% of the total population, respectively. The distributions varied between animals and ejaculates, but between straws, there were no differences in only four animals. This modern approach of considering an ejaculate sperm population as divided into subpopulations reflecting quantifiable parameters generated by CASA-Morph systems technology opens a new view on sperm function. This is the first study applying this approach to evaluate different ejaculates and straws from the same individual. More work must be done to improve seminal dose calculations in assisted reproductive programs.
