摘要
【目的】通过开展绵羊Dlx3基因启动子结构、活性、多态性及其与羊毛品质性状的关联等分析,揭示Dlx3基因在绵羊毛囊发育中的作用及其作用机制。【方法】采用PCR扩增Dlx3基因起始密码子上游1.5 kb区域,利用荧光素酶报告基因技术分析Dlx3基因启动子活性,采用测序方法寻找Dlx3基因启动子区SNP,并利用PCR-RFLP技术进行SNP分型。【结果】①Dlx3基因启动子近端序列的保守性较高,该区域内人、鼠及绵羊都具有23个保守的转录因子结合位点和一个CpG岛,而启动子的远端序列的保守性较低;②Dlx3基因的启动子在绵羊胚胎成纤维细胞中具有启动子活性;③Dlx3基因启动子的-1 551—-1 108 bp与-1 108—-707 bp区域对启动子活性影响较大;④Dlx3基因启动子区SNP位点(G-1166A)多态性与羊毛卷曲度显著相关。【结论】①Dlx3基因启动子在绵羊胚胎成纤维细胞中有活性;②Dlx3基因启动子的近端序列组成在人、鼠和绵羊中较为保守,而其远端序列的保守性较低,但是Dlx3基因启动子的远端序列对启动子活性影响较大;③Dlx3基因启动子区G-1166A位点是羊毛卷曲度的一个分子标记。
[Objective] Dlx3 gene is an important candidate gene for wool quality in sheep, but its function and transcriptional regulation are not fully understood. The aim of the present study was to characterize sheep Dlx3 gene promoter and analyze the association of Dlx3 gene promoter polymorphisms with wool quality traits. [ Method ] The 5＇ flanking fragment of the sheep Dlx3 gene was amplified by PCR and cloned. Promoter activity was analyzed using the luciferase reporter gene system, and SNP was genotyped by PCR-RFLP. [Result] Sequence analysis showed that the proximal promoter region of Dlx3 gene shared 23 transcription factor binding sites and a CpG island among human, mouse and sheep. Promoter activity analysis showed that sheep Dlx3 gene promoter was active in sheep embryonic fibroblasts. Promoter truncation mutation analysis showed that the -1 551 to - 1 108 and - 1 108 to -707 promoter regions had stronger promoter activity in vitro. Association analysis showed that SNP （G- 1166A） was significantly associated with wool crimp. [Conclusion] Dlx3 gene promoter is active in sheep embryonic fibroblasts, and its proximal promoter region, not the distal promoter region, is conserved among human, mouse and sheep. The distal promoter regionof Dlx3 gene has stronger promoter activity. The G-1166A is a potential marker of wool crimp for molecular marker-assisted selection in sheep.
出处
《中国农业科学》
CAS
CSCD
北大核心
2013年第3期614-622,共9页
Scientia Agricultura Sinica
基金
转基因生物新品种培育重大专项(2009ZX08009-160B
2011ZX08009-002)
