水貂DCT基因5'UTR克隆分析与启动子活性探究

doi:10.11843/j.issn.0366-6964.2019.03.007

Abstract

Abstract:

The research aimed to get 5'UTR sequence of mink DCT gene, characterize its structure, predict the transcriptional regulation elements, detect its promoter activity, and provide a theoretical basis for exploring the role of DCT gene in regulating the fur color formation of mink. 5'UTR of DCT gene of black mink, white mink and coffee mink were amplified and compared. Serial fragments deleted in 5'UTR, pGL3-1-pGL3-7 for coffee mink and pGL3-4-pGL3-6 for black mink, were amplified to construct the recombined luciferase reporter gene plasmid and to measure the promoter activity. The three kinds of minks with different coat colors were detected for the methylation level of CpG island in promoter of DCT gene by bisulfite sequencing PCR. 8 203 bp sequence of 5'UTR in DCT gene was cloned and a transposon with 204 bp in length in g.7133-7336 region was discovered. Among the 100 items with high similarity, one was from Soboliphyme baturini in Ecdysozoa and the others were all from Caniformia. P3 and P4 fragments had significant promoter activity (P<0.05). The methylation level in CpG island of coffee mink was significantly higher than that of black mink and white mink (P<0.05), and the promoter activity of individuals with CC haplotype in coffee mink was significantly lower than those with TT haplotype in black mink (P<0.05). The 204 bp transposable element in 5'UTR of mink DCT gene, special Can-SINEs of Caniformia, was inserted into the genome from Soboliphyme baturini into Ecdysozoa. The 32 bp element and the proximal domain in DCT co-activated the promoter, while the GC-box and CpG islands silenced the promoter activity of mink DCT gene. The CC haplotype, formed from the 2 SNPs of T > C mutation at locus of g.-684 and g.-621, resulted in the higher methylation level and the lower promoter activity in coffee mink, and might ultimately decrease the synthesis of eumelanin and form the coffee coat character.

CLC Number:

S829.9

LI Lanhui, DU Xiaolong, WANG Qi, GE Linhan, ZHANG Lechao, LI Xuemei, LI Xianglong. Exploration of Promoter Activity and Analysis of 5'UTR Sequence of Mink DCT Gene[J]. ACTA VETERINARIA ET ZOOTECHNICA SINICA, 2019, 50(3): 524-533.

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Exploration of Promoter Activity and Analysis of 5'UTR Sequence of Mink DCT Gene

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