山羊Agouti基因外显子1剪接体及启动子活性区研究

发布时间：2018-03-09 00:14

本文选题：山羊　切入点：Agouti基因　出处：《河北农业大学》2015年硕士论文　论文类型：学位论文

【摘要】：本试验以云南黑山羊的Agouti基因序列为模板设计引物,利用黑白两种颜色的山羊基因组作为样品对Agouti基因部分序列进行扩增,获得了Agouti基因的部分序列。根据不同毛色山羊Agouti基因5′非翻译区外显子1剪接体不同类型,设计引物,采用黑白两种颜色的山羊个体进行剪接体验证。试验结果表明:所选山羊个体的剪接体类型均为It It′。山羊基因组序列(登录号:AJPT01136617.1)中有两段序列与牛的预测启动子同源性比较高,因此设计引物扩增目的片段A和B,同时构建两个荧光素酶报告基因质粒PGL3-basic-A和PGL3-basic-B。并用构建的荧光素酶报告基因质粒瞬时转染人皮肤黑素瘤细胞A375(皮肤细胞)和293T细胞(非皮肤细胞)来检测两段目的片段是否具有Agouti基因启动子活性。结果用SPSS19.0软件进行分析,结果表明:所构建的目的片段的质粒与阴性对照相比不存在显著性差异,说明所获得的两段目的片段均不具有启动子活性。
[Abstract]:In this experiment, the Agouti gene sequence of Yunnan black goat was used as template to design primers, and the partial sequence of Agouti gene was amplified by using black and white goat genome as samples. Partial sequence of Agouti gene was obtained. Primers were designed according to different types of splicing of 5 'untranslated region exon 1 of Agouti gene in different wool goats. The experimental results showed that the splicing types of goat individuals were all it It.Goat genome sequence (accession No.: AJPT01136617.1) had high homology with the predicted promoter of cattle. Therefore, primers were designed to amplify the target fragments A and B, and two luciferase reporter gene plasmids PGL3-basic-A and PGL3-basic-Bwere constructed at the same time. The constructed plasmids of luciferase reporter gene were transiently transfected into human skin melanoma cells A375 (skin cells) and 293T thin cells. Cells (non-skin cells) were used to detect whether the two target fragments had the promoter activity of Agouti gene. The results were analyzed by SPSS19.0 software. The results showed that there was no significant difference between the plasmid of the constructed target fragment and that of the negative control, indicating that neither of the two fragments had promoter activity.
【学位授予单位】：河北农业大学
【学位级别】：硕士
【学位授予年份】：2015
【分类号】：S827

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