基因捕获技术在肝癌侵袭和迁移相关基因筛选中的应用

Application of gene trap technique in genetic screening of genes related to the invasion and matastasis of hepatocellular carcinoma

摘要:

目的：用基因捕获技术在肝癌细胞SMMC7721细胞中寻找与肝癌侵袭和迁移相关的基因。方法：首先，将pU21质粒转染到肝癌细胞SMMC7721细胞，用G418筛选出单克隆稳定细胞系；其次，采用Transwell小室实验和划痕实验，检测这些单克隆细胞侵袭和迁移能力的变化；最后，用5’-Full RACE实验、T克隆及基因测序检测这些细胞系中被pU21质粒捕获的基因。结果：成功获得了约300株稳定转染pU21质粒的单克隆细胞系，得到了约30株侵袭和迁移能力增强的细胞株和约40株侵袭和迁移能力减弱的细胞株。通过5’-Full RACE实验、T克隆及基因测序检测，证明了在A554细胞系中，被pU21质粒捕获的基因是long inter-genic non-protein coding RNA 52（LINC00052）。结论：应用基因捕获技术筛选肝癌侵袭和迁移相关基因是可行的。

Abstract:

Objective：To search for genes related to the invasion and matastasis of hepatocellular carcinoma（HCC） with the gene trap－ping technology. Methods：Firstly，SMMC7721 cells were transfected with capture plasmid pU21 vector and then selected by G418 to set up cell lines. Secondly，transwell assay and wound healing assay were used to detect the invasion and migration abilities of these cell lines. Thirdly，5’-Full RACE，gene cloning experiment and gene sequencing experiments were used to identify what the genes were captured by pU21 vector in these cell lines. Results：About 300 cell lines stably transfected with pU21. Invasion and migration abilities of about 70 cell lines were obviously changed. One cell line，A554 was found that the gene captured by pU21 was long inter-genic non-protein coding RNA 52（LINC00052）. Conclusion：Gene trap technology is useful to find genes related to the invasion and matastasis of HCC.