Cloning and Tissue Expression PHB Gene in Meiren Yak

doi:10.7668/hbnxb.20194168

Abstract

Abstract:

In order to study the structure and function of PHB gene in Meiren yak,and to explore its expression in different tissues at 7-day-old and adult cattle.This study cloned the coding region of PHB gene from Meiren yak kidney tissue cDNA as a template.The gene sequence was analyzed by bioinformatics and the physical and chemical properties of the protein were predicted.The relative expression levels of PHB gene in 8 tissues of Meiren yak at two ages were detected by Real-time Fluorescence Quantitative (RT-qPCR).The results showed that the CDS region of the PHB gene of Meiren yak was 819 bp in length,encoding 272 amino acids.The results of homology analysis showed that Meiren yak had the highest homology with common cattle (99.1%).The analysis of PHB protein showed that there was one potential N-glycosylation site and two potential O-glycosylation sites in the PHB protein.Its amino acid isoelectric point was 5.55,the instability coefficient (Ⅱ) was 46.07,and the total average hydrophobicity was 0.045,which was predicted to be an unstable hydrophobic protein.Subcellular localization showed that PHB protein was located in the mitochondria (30.4%),and there were 15 potential phosphorylation sites in the protein.The high-level structure of PHB protein was interconnected by α-helix (56.25%),random coil (22.43%),extended chain (16.54%) and β-corner (4.78%).The results of RT-qPCR showed that the PHB gene was expressed in eight tissues of Meiren yak at two developmental stages.The expression of PHB gene in 7 tissues decreased with the growth and development of Meiren yak.The expression of PHB gene in liver,spleen,lung,fat and testis decreased significantly with age (P<0.05).

Key words: Meiren yak, PHB genes, Cloning, Bioinformatics, Expression profile

YANG Guowu, WANG Tong, MA Xiaoyong, ZHA Lao, LU Jianwei, ZHAO Xue, YAN Ping, LIANG Chunnian. Cloning and Tissue Expression PHB Gene in Meiren Yak[J]. Acta Agriculturae Boreali-Sinica, 2023, 38(5): 218-226. doi: 10.7668/hbnxb.20194168.

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Figures/Tables 15

References 33

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基因 Genes	引物序列(5'-3') Primer sequences(5'-3')	长度/bp Length	退火温度/℃ Annealing temperature	用途 Application
PHB	F:CAGAGTGGAAGCAGGTGTGAG	1 241	58.2	PCR
	R:AAGGGTTGGTTGGAGGCTAC
PHB	F:AGTCAGCTTCCTCGCATCTTC	116	59.0	RT-qPCR
	R:AGTTCTCCAGCATCAAAGCGA
GAPDH	F:AATGAAAGGGCCATCACCATC	204	59.0	RT-qPCR
	R:GTGGTTCACGCCCATCACA

基因 Genes	引物序列(5'-3') Primer sequences(5'-3')	长度/bp Length	退火温度/℃ Annealing temperature	用途 Application
PHB	F:CAGAGTGGAAGCAGGTGTGAG	1 241	58.2	PCR
	R:AAGGGTTGGTTGGAGGCTAC
PHB	F:AGTCAGCTTCCTCGCATCTTC	116	59.0	RT-qPCR
	R:AGTTCTCCAGCATCAAAGCGA
GAPDH	F:AATGAAAGGGCCATCACCATC	204	59.0	RT-qPCR
	R:GTGGTTCACGCCCATCACA

软件 Software	功能 Function
ORF Finder	开放阅读框分析
Blast	同源性比对分析
ProtParam	蛋白质理化性质分析
ProtScale	蛋白质亲疏水性分析
SignalP 5.0	蛋白质信号肽分析
TMHMM 2.0	蛋白质跨膜结构预测
NetPhos 3.1	蛋白磷酸化位点预测
NetNGlyc	蛋白糖基化位点预测
TargetP 2.0	蛋白亚细胞定位预测
PSORT Ⅱ	蛋白亚细胞定位预测
SOPMA	蛋白质二级结构预测
SWISS-MODEL	蛋白质三级结构预测
STRING	蛋白互作网络预测

软件 Software	功能 Function
ORF Finder	开放阅读框分析
Blast	同源性比对分析
ProtParam	蛋白质理化性质分析
ProtScale	蛋白质亲疏水性分析
SignalP 5.0	蛋白质信号肽分析
TMHMM 2.0	蛋白质跨膜结构预测
NetPhos 3.1	蛋白磷酸化位点预测
NetNGlyc	蛋白糖基化位点预测
TargetP 2.0	蛋白亚细胞定位预测
PSORT Ⅱ	蛋白亚细胞定位预测
SOPMA	蛋白质二级结构预测
SWISS-MODEL	蛋白质三级结构预测
STRING	蛋白互作网络预测

氨基酸 Amino acid	数量/个 Amount	含量/% Content	氨基酸 Amino acid	数量/个 Amount	含量/% Content
丙氨酸Ala(A)	32	11.8	亮氨酸Leu(L)	27	9.9
精氨酸Arg(R)	18	6.6	赖氨酸Lys(K)	12	4.4
天冬酰胺Asn(N)	7	2.6	蛋氨酸Met(M)	1	0.4
天冬氨酸Asp(D)	15	5.5	苯丙氨酸Phe(F)	13	4.8
半胱氨酸Cys(C)	1	0.4	脯氨酸Pro(P)	9	3.3
谷氨酰胺Gln(Q)	15	5.5	丝氨酸Ser(S)	17	6.2
谷氨酸Glu(E)	19	7.0	苏氨酸Thr(T)	14	5.1
甘氨酸Gly(G)	18	6.6	色氨酸Trp(W)	1	0.4
组氨酸His(H)	3	1.1	酪氨酸Tyr(Y)	4	1.5
异亮氨酸Ile(I)	21	7.7	缬氨酸Val(V)	25	9.2

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