Huang Hui-Huang, Ji Dong, Wang Si-Yu, Xu Wan-Zhen, Jia Zhi-Yuan, Li Ke
Liver Failure Treatment Center, 302 hospital of PLA, Beijing 100039, China.
Zhonghua Shi Yan He Lin Chuang Bing Du Xue Za Zhi. 2011 Jun;25(3):179-81.
To construct a subtractive cDNA library of genes transactivated by PS1TP1 protein with suppression subtractive hybridization (SSH) technique.
Suppression subtractive hybridization technique and bioinformatics technique were used, the mRNA from HepG2 cells transfected with pcDNA3.1 (-) -PS1TP1 and pcDNA3. 1 (-) empty vector was isolated, respectively; cDNA underwent two times of nested PCR, amplified cDNA fragments were subcloned into pGEM-Teasy vectors to set up the subtractive library.
The subtractive library of genes transactivated by PS1TP1 was constructed successfully. Sequence analysis was performed in 43 clones randomly, and the full length sequences were obtained with bioinformatics method and searched for homologous DNA sequence from GenBank, altogether 12 coding sequences were gotten, which consisted of 10 known and 2 unknown ones.
The obtained sequences may be target genes transactivated by PS1TP1 protein among which some genes coding proteins involved in cell cycle regulation, metabolism, immunity and cell apoptosis. This finding brought some clues for studying the biological functions of PS1T1.
应用抑制性消减杂交（SSH）技术构建PS1TP1蛋白反式激活基因的消减cDNA文库。
采用抑制性消减杂交技术和生物信息学技术，分别提取转染pcDNA3.1（-）-PS1TP1和pcDNA3.1（-）空载体的HepG2细胞的mRNA；cDNA经两次巢式PCR，将扩增的cDNA片段亚克隆至pGEM-Teasy载体构建消减文库。
成功构建了PS1TP1蛋白反式激活基因的消减文库。随机挑选43个克隆进行序列分析，通过生物信息学方法获得全长序列并在GenBank中搜索同源DNA序列，共得到12个编码序列，其中10个为已知序列，2个为未知序列。
所获得的序列可能是PS1TP1蛋白反式激活的靶基因，其中一些基因编码的蛋白参与细胞周期调控、代谢、免疫及细胞凋亡等过程。这一发现为研究PS1T1的生物学功能提供了线索。
