<i>TNNI1</i>、<i>TNNI3</i>在麦洼牦牛横纹肌中的表达分析

doi:10.7668/hbnxb.20192913

摘要/Abstract

摘要：

为探索TNNI1、TNNI3在麦洼牦牛横纹肌中的表达模式,选取不同年龄麦洼牦牛臀肌、心肌组织,观察其形态学变化,开展了不同年龄麦洼牦牛TNNI1基因在臀肌组织中的表达趋势及与肌纤维性状的关联分析,同时对TNNI1、TNNI3在心肌中的mRNA和蛋白差异表达及蛋白定位进行了相关性分析。结果表明,随着牦牛年龄增长,肌纤维直径及密度存在一定特征性变化,其中肌纤维直径随年龄的增长显著增加,肌纤维密度则发生相反的变化。成年牦牛TNNI1基因在臀肌组织中的mRNA表达水平显著高于犊牛及断奶犊牛,变化趋势具体为成年牛>犊牛>断奶犊牛;且臀肌中TNNI1 mRNA表达水平与肌纤维直径极显著正相关,与肌纤维密度显著负相关。TNNI1 在心肌中的mRNA表达水平随牦牛年龄的增大而降低,而TNNI3则表现为先升高后降低的趋势,且TNNI1在犊牛期mRNA表达水平最高,TNNI3则随年龄增长呈上调趋势。TNNI1、TNNI3蛋白在心肌组织中的表达量随牦牛年龄的增长逐渐减小,且TNNI3 mRNA表达量随牦牛年龄的增长逐渐降低,与蛋白表达趋势一致。TNNI1、TNNI3对不同年龄麦洼牦牛肌肉组织的生长发育具有显著调控作用,且在不同年龄牦牛肌肉组织中的表达水平存在差异。

关键词: 牦牛, TNNI1, TNNI3, 横纹肌, Western Blot, 免疫组织化学

Abstract:

In order to explore the expression patterns of TNNI1 and TNNI3 in the striated muscle of Maiwa yak,this study selectd the gluteal muscle and myocardial tissue of Maiwa yak of different ages,morphological changes observed.The expression trend of TNNI1 gene in gluteal muscle tissue of Maiwa yak at different ages and its association with muscle fiber traits were carried out,the mRNA,protein differential expression and protein localization of TNNI1 and TNNI3 in myocardium were analyzed.The results showed that the muscle fiber diameter and density had certain characteristic changes with the increase of yak age.The muscle fiber diameter increased significantly with the increase of age,while the muscle fiber density had the opposite change.The mRNA expression of TNNI1 gene in gluteal muscle tissue of adult yaks was significantly higher than calves and weaned calves. TNNI1 mRNA expression in gluteus muscle was significantly positively correlated with muscle fiber diameter,and significantly negatively correlated with muscle fiber density.The mRNA expression level of TNNI1 in myocardium decreased with the increase of yak age,while the expression level of TNNI3 showed a trend of first increased and then decreased,and the mRNA expression level of TNNI1 was the highest in calf stage,while TNNI3 showed an upward trend with the increase of age.The expression levels of TNNI1 and TNNI3 proteins in myocardium decreased with the increase of yak age.The expression levels of TNNI1 and TNNI3 proteins in myocardium decreased with the increase of yak age.The TNNI3 mRNA expression decreased with the increase of yak age,which was consistent with the trend of protein expression.It provided basic data for exploring the regulation mechanism of TNNI1 and TNNI3 on yak muscle growth and development.TNNI1 and TNNI3 have significant regulatory effects on the growth and development of the muscle tissue of Maiwa yak at different ages,and the expression levels in the muscle tissue of yak at different ages are different.

Key words: Yak, TNNI1, TNNI3, Striated muscle, Western Blot, Immunohistochemistry

向娅, 钟金城, 武志娟, 柴志欣. TNNI1、TNNI3在麦洼牦牛横纹肌中的表达分析[J]. 华北农学报, 2022, 37(6): 210-218. doi: 10.7668/hbnxb.20192913.

XIANG Ya, ZHONG Jincheng, WU Zhijuan, CHAI Zhixin. Expression Analysis of TNNI1 and TNNI3 in Striated Muscle of Maiwa Yak[J]. Acta Agriculturae Boreali-Sinica, 2022, 37(6): 210-218. doi: 10.7668/hbnxb.20192913.

http://www.hbnxb.net/CN/Y2022/V37/I6/210

图/表 14

表1 PCR扩增引物

Tab.1 Primers for PCR

基因名称 Genes	引物序列(5'—3') Oligo sequence(5'—3')	片段长度/bp Fragment length	退火温度/℃ Annealing temperature
TNNI1	F:CTCCAAGCACAAGGTGTCTATG	158	56
	R:TGGCGGCATCAAACATCT
TNNI3	F:CTCGTGTGGACAAGGTGGA	137	58
	R:GCCCCGAAGGTCAAAGAT
GAPDH	F:CCACGAGAAGTATAACAACACC	120	55
	R:GTCATAAGTCCCTCCACGAT

图1 麦洼牦牛臀肌组织TNNI1 mRNA表达水平不同小写字母表示差异显著(P<0.05)。图2,5,6,9—11同。

Fig.1 Expression level of TNNI1 in gluteal muscle tissues of Maiwa yak Different lowercase letters indicate significant difference (P<0.05).The same as Fig.2,5,6,9—11.

图2 麦洼牦牛心肌组织TNNI1、TNNI3 mRNA表达量不同大写字母表示不同基因表达量之间差异显著(P<0.05)。

Fig.2 Expression of TNNI1 and TNNI3 in Maiwa yak myocardium Different capital letters indicate significant difference between two genes(P<0.05).

图3 蛋白质吸光度标准曲线

Fig.3 Protein absorbance standard curve

图4 心肌蛋白聚丙烯酰胺凝胶电泳 1—3.成年牛;4—6.断奶犊牛;7—9.犊牛。

Fig.4 SDS-PAGE electrophoresis results of myocardial protein 1—3.Adult yak;4—6.Weaned calf;7—9.Calf.

图5 TNNI1蛋白印迹分析

Fig.5 Western Blot analysis of TNNI1

图6 TNNI3蛋白WB分析

Fig.6 Western Blot analysis of TNNI3

图7 不同年龄麦洼牦牛臀肌纤维切片(10×10) A.犊牛;B.断奶犊牛;C.成年牛。图8—11同。

Fig.7 Gluteal muscle fibers slices of Maiwa yak at different ages(10×10) A.Calf;B.Weaned calf;C.Adult yak.The same as Fig.8—11.

表2 各年龄段麦洼牦牛臀肌纤维组织学性状测定

Tab.2 Gluteal muscle fiber traits comparison of Maiwa yak at different ages

年龄段 Age	肌纤维直径/μm Muscle fiber diameter	肌纤维密度 /(N/mm²) Muscle fiber density
犊牛Calf	65.26±9.79b	4 437.45±280.29a
断奶犊牛Weaned calf	69.79±6.00b	4 207.25±118.11a
成年牛Adult yak	142.31±13.09a	2 880.91±69.83b

图8 不同年龄麦洼牦牛心肌纤维切片

Fig.8 Myocardial muscle fibers slices of Maiwa yak at different ages

表3 TNNI1 mRNA表达量与肌纤维性状的相关性

Tab.3 Correlations between TNNI1 mRNA expression and muscle fiber traits

mRNA表达 mRNA expression	项目 Items	相关系数 Correlation coefficient	P值 P value
TNNI1	肌纤维直径	0.95	0.004
	肌纤维密度	-0.90	0.016

图9 不同年龄麦洼牦牛臀肌组织TNNI1蛋白表达特征(10×10)

Fig.9 Protein expression characteristics of TNNI1 in gluteal muscle(10×10)

图10 不同年龄麦洼牦牛心肌组织TNNI1蛋白表达特征(10×10)

Fig.10 Protein expression characteristics of TNNI1 in myocardial(10×10)

图11 不同年龄麦洼牦牛心肌组织TNNI3蛋白表达特征(10×10)

Fig.11 Protein expression characteristics of TNNI3 in myocardial(10×10)

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