

畜牧兽医学报 ›› 2025, Vol. 56 ›› Issue (11): 5475-5488.doi: 10.11843/j.issn.0366-6964.2025.11.011
李炳志1,2,4(
), 郭俊涛2,3, 王建芳2,3, 于胜晨2,3, 潘月婷2,3, 余横伟2,3, 刘海兵2,3, 张可2,3, 成功2,3, 田万强1,4,*(
), 昝林森2,3,*(
)
收稿日期:2025-03-25
出版日期:2025-11-23
发布日期:2025-11-27
通讯作者:
田万强,昝林森
E-mail:3640910@qq.com;twqiang2003@163.com;zanlinsen@163.com
作者简介:李炳志(1982-),男,山东人,博士,主要从事动物遗传育种与繁殖研究,E-mail: 3640910@qq.com
基金资助:
LI Bingzhi1,2,4(
), GUO Juntao2,3, WANG Jianfang2,3, YU Shengchen2,3, PAN Yueting2,3, YU Hengwei2,3, LIU Haibing2,3, ZHANG Ke2,3, CHENG Gong2,3, TIAN Wanqiang1,4,*(
), ZAN Linsen2,3,*(
)
Received:2025-03-25
Online:2025-11-23
Published:2025-11-27
Contact:
TIAN Wanqiang, ZAN Linsen
E-mail:3640910@qq.com;twqiang2003@163.com;zanlinsen@163.com
摘要:
旨在探究干扰MAPK6(细胞外信号调节激酶3,ERK3)基因对秦川牛成肌细胞分化的影响及其作用机理。本研究通过qRT-PCR技术检测MAPK6基因在秦川牛不同组织中的表达谱。采用siRNA技术干扰MAPK6基因在秦川牛成肌细胞中的表达,通过qRT-PCR和Western blot检测其干扰效率及肌管分化相关基因的表达水平,并观察肌管表型变化。随后对其进行转录组测序,筛选差异表达基因并进行GO功能注释和KEGG通路富集分析。此外,对成肌分化相关通路进行了初步验证。结果表明,MAPK6在秦川牛背最长肌组织中特异性高表达。干扰MAPK6极显著促进了秦川牛成肌细胞的肌管分化融合(P < 0.01),极显著增加了成肌分化相关基因MEF2C、MYH7的表达(P < 0.01)及MEF2C、MYL2的蛋白表达(P < 0.01),显著增加成肌MYH7的蛋白表达(P < 0.05)。转录组结果发现,与对照组相比,干扰MAPK6基因共有3 224个基因差异表达,其中上调1 707个,下调1 517个。KEGG富集结果显示,差异基因主要富集在PI3K-AKT、MAPK信号通路。MAPK6基因的干扰会显著降低MAPK信号通路中ERK1/2、p-ERK1/2蛋白的表达(P < 0.01)。本研究初步阐明了MAPK6基因在牛肌肉生成中的作用和机理,为提高牛肉品质和指导肉牛分子育种策略奠定基础。
中图分类号:
李炳志, 郭俊涛, 王建芳, 于胜晨, 潘月婷, 余横伟, 刘海兵, 张可, 成功, 田万强, 昝林森. 干扰MAPK6基因对秦川牛成肌细胞分化的影响研究[J]. 畜牧兽医学报, 2025, 56(11): 5475-5488.
LI Bingzhi, GUO Juntao, WANG Jianfang, YU Shengchen, PAN Yueting, YU Hengwei, LIU Haibing, ZHANG Ke, CHENG Gong, TIAN Wanqiang, ZAN Linsen. Study on the Effect of Interfering with MAPK6 Gene on the Differentiation of Qinchuan Cattle Myoblast Cells[J]. Acta Veterinaria et Zootechnica Sinica, 2025, 56(11): 5475-5488.
表 1
实时定量PCR(qRT-PCR)引物序列"
| 基因 Gene | GenBank号 GenBank number | 引物 Primer | 引物序列(5′→3′) Primer sequence | 产物长度/bp Product length |
| GAPDH | NM_001034034.2 | GAPDH-F | AGTTCAACGGCACAGTCAAGG | 124 |
| GAPDH-R | ACCACATACTCAGCACCAGCA | |||
| MAPK6 | XM_059890855.1 | MAPK6-F | ACTTGGTGCTGAAGATAGGTGA | 206 |
| MAPK6-R | AAAGAGGGTTTTACCAGTCAGC | |||
| MEF2C | XM_059888100.1 | MEF2C-F | GCACCAACAAGCTGTTCCAG | 117 |
| MEF2C-R | GAACAGCCTCCACGATGTCT | |||
| MYH7 | XM_024997290.2 | MYH7-F | TCCAGTCTTCTCTCCTTGCTG | 187 |
| MYH7-R | AGGTCGAAAGGCCTGGTCTG | |||
| MYL2 | NM_001035025.2 | MYL2-F | CTTTCCACCATGTCACCTAAGA | 114 |
| MYL2-R | GATGGTGAAGGCCTCCTTAAA | |||
| CEND1 | NM_001080222.1 | CEND1-F | GAGCAGCAGCGCCCATA | 125 |
| CEND1-R | CTTCCCTCTGGACTCCATGC | |||
| SORD | NM_001037320.1 | SORD-F | CTGGCAGCATGGTCGAATTG | 102 |
| SORD-R | TGCCTGACCAATGATCCCAC | |||
| WASF3 | XM_005213794.5 | WASF3-F | CCAGGTTATCACAGAGCGCA | 102 |
| WASF3-R | CCGGGTACGAGTAATCCGTG | |||
| VPS50 | NM_001105413.2 | VPS50-F | ATGAGGGTCGTGCCTTGATG | 162 |
| VPS50-R | TGTGCTCTTTGATCCAGCGT | |||
| ZFP90 | XM_059877474.1 | ZFP90-F | CGAGTGAAGACGGCACAGAA | 85 |
| ZFP90-R | CCAGGGACAGGGTTGACTTC | |||
| CCNG1 | NM_001013364.1 | CCNG1-F | GCTGTGAAGCCTCAAAACATC | 170 |
| CCNG1-R | TCCAGGCTTGTCATTTGCATT | |||
| NR1D2 | XM_059882241.1 | NR1D2-F | GTCTAATTACCTGGTTTCCCCC | 127 |
| NR1D2-R | GACGCGGACTGGAAGCTATT | |||
| GCSH | NM_174844.1 | GCSH-F | TATTGAGGAGTGAAACTGGAACCC | 188 |
| GCSH-R | AGATGCAGTGTTTATCTGAAGTCAT | |||
| ABCB10 | XM_024986813.2 | ABCB10-F | CTCATGCAGACGTCAGGTCA | 107 |
| ABCB10-R | TCTCCTGTGCGTGTCTTGTC |
表 2
测序数据质控分析"
| 样品 Sample | 原始数据Raw data | 过滤数据Clean data | 有效比对率/% Valid ratio (reads) | Q20% | Q30% | GC含量/% GC content | |||
| 读数 Read | 碱基/G Base | 读数 Read | 碱基/G Base | ||||||
| MAPK6S1 | 40 273 634 | 6.04 | 38 754 750 | 5.81 | 96.23 | 99.93 | 98.04 | 50.50 | |
| MAPK6S2 | 40 890 852 | 6.13 | 39 414 348 | 5.91 | 96.39 | 99.94 | 97.97 | 50.50 | |
| MAPK6S3 | 40 659 854 | 6.10 | 39 237 220 | 5.89 | 96.50 | 99.94 | 98.04 | 49.50 | |
| SINC1 | 39 899 744 | 5.98 | 38 249 602 | 5.74 | 95.86 | 99.93 | 97.97 | 50.50 | |
| SINC2 | 38 885 912 | 5.83 | 37 072 804 | 5.56 | 95.34 | 99.92 | 97.89 | 51.50 | |
| SINC3 | 39 628 850 | 5.94 | 38 159 344 | 5.70 | 96.29 | 99.93 | 97.97 | 51.50 | |
表 3
参考基因组比对结果"
| 样品 Sample | 过滤数据 Clean reads | 比对读数 Mapped reads | 唯一比对读数 Unique mapped reads | 多比对读数 Multi mapped reads |
| MAPK6S1 | 38 754 750 | 36 246 759(93.53%) | 35 413 870(91.38%) | 832 889(2.15%) |
| MAPK6S2 | 39 414 348 | 36 937 794(93.72%) | 36 105 382(91.60%) | 832 412(2.11%) |
| MAPK6S3 | 39 237 220 | 36 907 942(94.06%) | 36 079 312(91.95%) | 828 630(2.11%) |
| SINC1 | 38 249 602 | 35 839 948(93.70%) | 34 934 213(91.33%) | 905 735(2.37%) |
| SINC2 | 37 072 804 | 34 373 945(92.72%) | 33 513 528(90.40%) | 860 417(2.32%) |
| SINC3 | 38 159 344 | 35 644 691(93.41%) | 34 805 597(91.21%) | 839 094(2.20%) |
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