斑马鱼(Danio rerio)mapk1基因对tp53基因调控研究  

Regulation of Gene mapk1 in Danio rerio on Gene tp53

作　　者：包林珠 时灿 卢玲儿 徐行 周泽斌[1,2] 任建峰 李伟明[3] 张庆华 BAO Lin-zhu;SHI Can;LU Ling-er;XU Xing;ZHOU Ze-bin;REN Jian-feng;LI Wei-ming;ZHANG Qing-hua(Key Laboratory of Exploration and Utilization of Aquatic Genetic Resources,Ministry of Education,Shanghai Ocean University,Shanghai 201306;International Research Center for Marine Biosciences,Ministry of Science and Technology,Shanghai Ocean University,Shanghai 201306;Department of Fisheries and Wildlife,Michigan State University,Michigan 48824,USA)

机构地区：[1]上海海洋大学水产种质资源发掘与利用教育部重点实验室,上海201306 [2]上海海洋大学中国科学技术部海洋生物科学国际联合研究中心,上海201306 [3]密歇根州立大学渔业与野生生物系,美国密歇根48824

出　　处：《生物技术通报》2021年第12期160-168,共9页Biotechnology Bulletin

基　　金：教育部留学回国人员科研启动基金(D-8002-15-0042);中美海洋研究中心基金(A1-0209-15-0806);水产动物疾病与基因编辑育种的平台建设和前沿科学研究(A1-3201-19-3013)。

摘　　要：旨在研究斑马鱼(Danio rerio)mapk1基因对tp53基因的调控作用。通过生信网站分析斑马鱼tp53启动子序列与mapk1基因CDS序列,构建斑马鱼tp53启动子荧光素酶报告基因质粒pGL3-tp53-Luc和mapk1表达质粒pCMV-Tag2B-mapk1。利用Luciferase实验验证tp53启动子报告基因的活性,以及mapk1基因对tp53基因的活性影响。结果显示,斑马鱼tp53基因启动子区无CpG岛位点,包含Oct-1(TATGTAAAGC)、Sp-1(AGATCCCGCC)、c-Jun(CTGACGTCAC)、CREB(CTGACGTCAC)、CREBP1(CTGACGTCAC)、NF-kappa B1(AGGGGAATCC)和RAR-alpha1(TTGAACTTTT)共7个转录因子结合位点;斑马鱼与人和小鼠MAPK1氨基酸的一致性分别为91.33%和90.79%。pGL3-tp53-Luc在哺乳动物细胞系和鱼类细胞系中均具有很高的活性,分别是对照组的17倍和9倍。在HEK293T细胞中过表达pCMV-Tag2B-mapk1质粒后pGL3-tp53-Luc的luciferase活性是对照组的3倍左右。实验验证了斑马鱼mapk1基因可促进tp53基因的表达。The objective is to investigate the regulation of zebrafish(Danio rerio)mapk1 gene on tp53 gene.The tp53 promoter sequence and mapk1 gene CDS sequence of zebrafish were analyzed by bioinformatics websites,and then the luciferase reporter gene plasmid pGL3-tp53-Luc and mapk1 expression plasmid pCMV-Tag2B-mapk1 were constructed.Finally,luciferase assay was used to verify the activity of tp53 promoter reporter gene and the effect of mapk1 gene on the activity of tp53 gene.The results showed that there was no CpG island site in the promoter region of tp53 gene in zebrafish,and there were 7 transcription factors binding sites,including Oct-1(TATGTAAAGC),Sp-1(AGATCCCGCC),c-Jun(CTGACGTCAC),CREB(CTGACGTCAC),CRE-BP1(CTGACGTCAC),NF-KappaB1(AGGGGAATCC),and RAR-alpha1(TTGAACTTTT).The amino acid consistency of MAPL1 between zebrafish and human and mouse was 91.33%and 90.79%,respectively.The tp53 promoter fragment sequence had high activity in both mammalian cell lines and fish cell lines,which were 17 times and 9 times that of the control group,respectively.After overexpression of pCMV-Tag2B-mapk1 plasmid in the HEK293T cells,the luciferase activity of pGL3-tp53-Luc was about 3 times that of the control group.This experiment verifies that zebrafish mapk1 may promote the expression of tp53.

关 键 词：斑马鱼 TP53 mapk1 报告基因 

分 类 号：Q78[生物学—分子生物学]