泥蚶转录因子c-Myc与下游ATP结合盒转运蛋白基因ABCA3启动子的结合鉴定

doi:10.16178/j.issn.0528-9017.20221192

摘要/Abstract

摘要：

为了确定泥蚶转录因子c-Myc(Tgc-Myc)与下游ATP结合盒转运蛋白ABCA3(TgABCA3)基因启动子的结合关系。利用在线生物信息学网站预测以及凝胶电泳迁移(electrophoretic mobility shift assay,EMSA)和染色质免疫共沉淀(chromatin immunoprecipitation assay,ChIP)技术,对Tgc-Myc与TgABCA3启动子的结合位点进行分析。结果显示,TgABCA3启动子上的E-box顺式作用元件位于-40~-35 bp,Tgc-Myc能够与该序列特异性结合。转录因子Tgc-Myc通过调控TgABCA3基因转录,从而参与泥蚶应对重金属Cd胁迫的响应机制。

关键词: 泥蚶, 转录因子c-Myc, ATP结合盒转运蛋白, E-box, 凝胶电泳迁移, 染色质免疫共沉淀

中图分类号:

S-03

赵德风, 陈然, 肖国强, 滕爽爽. 泥蚶转录因子c-Myc与下游ATP结合盒转运蛋白基因ABCA3启动子的结合鉴定[J]. 浙江农业科学, 2023, 64(6): 1317-1322.

图/表 8

参考文献 63

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反应组分	阴性对照反应组	结合反应组	探针冷竞争反应组	突变探针的冷竞争反应组	特异性抗体组
无菌水	11	9	7	7	7
10×binding buffer	2	2	2	2	2
1 g·L^-1 Poly(dI.dC)	1	1	1	1	1
50%甘油	1	1	1	1	1
1% NP-40	1	1	1	1	1
1 mol·L^-1 KCl	1	1	1	1	1
100 mmol·L^-1 MgCl₂	1	1	1	1	1
200 mmol·L^-1 EDTA	1	1	1	1	1
未标记特异性探针	0	0	2	0	0
未标记突变探针	0	0	0	2	0
核蛋白	0	2	2	2	2
目标特异性抗体	0	0	0	0	2
生物素标记特异性探针	1	1	1	1	1
总体积	20	20	20	20	20

反应组分	阴性对照反应组	结合反应组	探针冷竞争反应组	突变探针的冷竞争反应组	特异性抗体组
无菌水	11	9	7	7	7
10×binding buffer	2	2	2	2	2
1 g·L^-1 Poly(dI.dC)	1	1	1	1	1
50%甘油	1	1	1	1	1
1% NP-40	1	1	1	1	1
1 mol·L^-1 KCl	1	1	1	1	1
100 mmol·L^-1 MgCl₂	1	1	1	1	1
200 mmol·L^-1 EDTA	1	1	1	1	1
未标记特异性探针	0	0	2	0	0
未标记突变探针	0	0	0	2	0
核蛋白	0	2	2	2	2
目标特异性抗体	0	0	0	0	2
生物素标记特异性探针	1	1	1	1	1
总体积	20	20	20	20	20

引物名称	引物序列(5'-3')	扩增位置/bp
ABCA3-1F	GTCTCGCTCCTCTACCCAC	-1 955~-1 415
ABCA3-1R	TAACTTTACCGATCTCCTGTTTGTG
ABCA3-2F	CCTCTACCCACACAGATTC	-1 235~-815
ABCA3-2R	AAATGGTAGAGGAGAGGGAC
ABCA3-3F	CAGGTAACCAACACAGG	-450~-10
ABCA3-3R	TTGGGATGTCGAATATGC

引物名称	引物序列(5'-3')	扩增位置/bp
ABCA3-1F	GTCTCGCTCCTCTACCCAC	-1 955~-1 415
ABCA3-1R	TAACTTTACCGATCTCCTGTTTGTG
ABCA3-2F	CCTCTACCCACACAGATTC	-1 235~-815
ABCA3-2R	AAATGGTAGAGGAGAGGGAC
ABCA3-3F	CAGGTAACCAACACAGG	-450~-10
ABCA3-3R	TTGGGATGTCGAATATGC

软件	起始位点	终止位点	E值	评分
PROMO	-43(+)	-35(+)	0.51	68.55
LASAGNA-Search2.0	-40(+)	-35(+)	0.49	67.75

泥蚶转录因子c-Myc与下游ATP结合盒转运蛋白基因ABCA3启动子的结合鉴定

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