牦牛GLUT1、GLUT3和GLUT8基因的克隆及其在乳腺上皮细胞中的表达

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Abstract Glucose uptake in the mammary gland mainly depends on the glucose transporter (GLUT), and the different GLUT has different effects on glucose uptake. The aim of this study was to investigate the molecular mechanism of glucose transporter that participate in lactation regulation of yak (Bos grunniens). Mammary epithelial cells were cultured in vitro using pancreatic enzyme digestion method, and the immunocytochemistry method was used to appraise mammary epithelial cells by detecting the expression of cytokeratin 18, β-casein and vimentin; GLUT1, GLUT3 and GLUT8 gene were cloned by PCR from purified cells and its biological characteristics were also analyzed. The mRNA and protein expression of GLUT1, GLUT3 and GLUT8 was detected by qRT-PCR and Western blot, and their location in yak mammary epithelial cells were detected by indirect immunofluorescence. The results showed that cytokeratin 18 and β-casein could be detected in the isolated cells, and vimentin was negative, which indicated the cells were mammary epithelial cells. The GLUT1, GLUT3 and GLUT8 (GenBank accession number: KU902419, KX094556 and KX268646) gene were successfully cloned from mammary epithelial cells of yak, which contained a complete coding sequence, the length of nucleotide were 1 479, 1 485 and 1 437 bp, respectively, and encoded 492, 494 and 478 amino acids, respectively. Nucleotide sequence analysis revealed that GLUT1, GLUT3 and GLUT8 genes were conserved. The physical and chemical properties of protein encoded by 3 genes were similar, all of them were hydrophobic membrane proteins including 12 transmembrane helical region. The expression of GLUT1, GLUT3 and GLUT8 in mammary epithelial cells of yak were extremely significant difference (P＜0.01), GLUT1 was the highest, followed by GLUT8, while GLUT3 was the lowest. GLUT1, GLUT3 and GLUT8 protein were mainly located in the nucleus of in mammary epithelial cells of yak. The results provide important information for understanding the physiological function of yak glucose transporters and a new theoretical basis for further studying the biological function of yak lactation.

Key words： Yak Glucose transporter 1 (GLUT1) GLUT3 GLUT8 Mammary epithelial cells

Received: 02 September 2016 Published: 24 December 2016

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	LI Ru-Qin
	PAN Yang-Yang
	TIAN Kui-Li
	DIAO Tian
	ZHANG Yi-Fu
	FAN Jiang-Feng
	CUI Yan
	YU Si-Jiu

Cite this article:

LI Ru-Qin,PAN Yang-Yang,TIAN Kui-Li, et al. Cloning of GLUT1, GLUT3 and GLUT8 Genes and Their Expression in Mammary Epithelial Cells of Yak (Bos grunniens) [J]. , 2017, 25(1): 21-31.

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http://journal05.magtech.org.cn/Jwk_ny/EN/ OR http://journal05.magtech.org.cn/Jwk_ny/EN/Y2017/V25/I1/21

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