山羊CSRP3基因对成肌细胞分化的功能研究

doi:10.3864/j.issn.0578-1752.2026.09.012

Abstract

Abstract:

【Objective】Skeletal muscle growth and development is one of the important factors affecting meat quality of goat. This study aimed to clone the coding sequence (CDS) of the goat CSRP3 gene (Cysteine and Glycine Rich Protein 3), conduct bioinformatics analysis using the online software, investigate the expression patterns of the goat CSRP3 gene in different tissues and at different differentiation stages of goat myoblasts, and clarify the regulatory role of this gene in myoblast differentiation.【Method】Tissues including the heart, liver, spleen, lung, kidney, and longissimus dorsi muscle were collected from goats. Total RNA was extracted from these tissues using the Trizol method and was reverse-transcribed to cDNA. Using the cDNA from the longissimus dorsi muscle as the template, RT-PCR was performed to amplify the CDS region of the goat CSRP3 gene, and online software was used for bioinformatics analysis. cDNA from different goat tissues and myoblasts at different differentiation stages was used as templates to detect the CSRP3 gene expression level by quantitative Real-Time PCR (qPCR). Through the construction of an overexpression vector, siRNA synthesis, and cell transfection experiments, qPCR and immunofluorescence staining were employed to determine the relative mRNA expression level of myoblast differentiation marker genes and the formation of myotubes after overexpression or interference of the goat CSRP3 gene.【Result】The full-length of the goat CSRP3 gene is 1 103 bp, with a CDS sequence of 585 bp. Phylogenetic tree analysis showed that the goat CSRP3 gene has the closest genetic relationship with sheep and cattle, indicating high conservation of this gene among closely related species. The goat CSRP3 gene encodes 194 amino acids, and the encoded protein is a hydrophilic and stable basic protein. Protein-protein interaction analysis revealed that the goat CSRP3 protein may be involved in muscle growth and development through interactions with muscle function-related proteins. The relative mRNA expression level of the CSRP3 gene in goat longissimus dorsi muscle and heart was significantly higher than that in other tissues (P < 0.05). During myoblast differentiation, the goat CSRP3 gene expression was low at the initial stage, increased gradually with differentiation time, peaked on day 3 (P < 0.01), and then decreased, showing a dynamic change of first increasing and then decreasing. This suggested that the goat CSRP3 gene may play an important regulatory role in the transition of myoblasts from proliferation to differentiation. Overexpression of the goat CSRP3 gene significantly increased the relative mRNA expression levels of myoblast differentiation marker genes and promoted myotube formation, indicating myoblast differentiation. Conversely, interference with the CSRP3 gene significantly reduced the relative mRNA expression levels of myoblast differentiation marker genes and inhibited myotube formation, impairing the differentiation ability of myoblasts.【Conclusion】The CDS of CSRP3 was successfully cloned (the cloned sequence is 603 bp in length, including a 585 bp CDS region), and its sequence characteristics and physicochemical properties were clarified. The CSRP3 tissue and temporal expression profiles in goats were established. It was confirmed that the CSRP3 gene exerts a positive regulatory effect on goat myoblast differentiation. These results preliminarily indicate that the CSRP3 gene plays an important biological function in the muscle growth and development of meat goats.

Key words: goat, CSRP3 gene, tissue expression, temporal expression, myoblast differentiation

MA HuiFeng, WEN ShuLiang, LIU WanYan, LIN YaQiu, LI Xin, LIU KeHan, ZHU HuiYing, XUE FuLai, XING JiaNi. Functional Study of Goat CSRP3 Gene on Myoblast Differentiation[J].Scientia Agricultura Sinica, 2026, 59(9): 2002-2015.

Figures/Tables 11

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基因 Gene	序列 Sequence	退火温度 Tm (℃)	产物长度 Product length (bp)	用途 Uses
CSRP3	F: tagCTCGAGatgccaaactggggtggag R: tacGGATCCtcaatctttcttttccacttgg	59	603	RT-PCR
CSRP3	F: CCGTCTACCATGCGGAAGAA R: GATCTCTGACTCGTGAGCGG	60	122	qPCR
UXT	F: GCAAGTGGATTTGGGCTGTAAC R: ATGGAGTCCTTGGTGAGGTTGT	60	180	qPCR
MYHC	F: CTCTTCCCGCTTTGGTAAGTT R: CAGGAGCATTTCGATTAGATCCG	60	187	qPCR
MYOG	F: GCAGGCTCAAGAAAGTGAATGA R: TAGGCGCTCAATGTACTGGAT	60	122	qPCR
MYOD1	F: GTGCAAACGCAAGACGACTA R: GCTGGTTTGGGTTGCTAGAC	59	128	qPCR

软件Software	网址 Websites	功能预测Function prediction
ProParam	https://web.expasy.org/protparam/	理化性质physicochemical properties
ProScale	https://web.expasy.org/protscale/	亲/疏水性hydrophilicity/hydrophobicity
SOPMA	https://npsa.lyon.inserm.fr/cgi-bin/npsa_automat.plpage=/NPSA/npsa_sopma.html	二级结构secondary structure
SWISS-MODEL	https://swissmodel.expasy.org/	三级结构tertiary structure
STRING	http://string-db.org/	蛋白互作protein-protein interaction

基因 Gene	正义链5′—3′ Sense 5′-3′	反义链5′—3′ Antisense 5′-3′
CSRP3-231	AGUGCAAUGGAAGGAGUUUTT	AAACUCCUUCCAUUGCACUTT
CSRP3-311	GCUCACGAGUCAGAGAUCUTT	AGAUCUCUGACUCGUGAGCTT
CSRP3-521	CCUCGAUGCGGAAAGUCAGTT	CUGACUUUCCGCAUCGAGGTT
CSRP3-612	GGAAGAGUCUAGAGUCCACTT	GUGGACUCUAGACUCUUCCTT
Negative Control FAM	UUCUCCGAACGUGUCACGUTT	ACGUGACACGUUCGGAGAATT

Functional Study of Goat CSRP3 Gene on Myoblast Differentiation

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