利用P因子不精确剪切获得Tis11基因敲除果蝇

Using P-element Imprecise Excision to Generate Tis11 Knockout Fly

Tis11是哺乳动物基因TTP在果蝇中的同源物,其基因位于果蝇X染色体11B9区.它含有两个保守的顺式Cys-X8-Cys-X5-Cys-X3-His(CCCH)锌指结构.它与mRNA的稳定性有关.为了进一步从动物水平研究Tis11的生物学功能,作者利用P因子不精确剪切的特性来获得Tis11基因敲除果蝇.以P因子插入果蝇19949为起始材料,经过一系列杂交后,挑取了700株P因子被切除的果蝇.通过PCR筛选和鉴定,从这700株果蝇中获得一株果蝇279,它的Tis11基因的第2个外显子中包括翻译起始位点的区域被删除.

Tis11 is a mammalian TTP homologue in Drosophila. It locates to region 11B9 on X chromosome in Drosophila. It consists of two conserved tandem Cys-Xs-Cys-Xs-Cys-X3-His（CCCH）zinc fingers. Tisll can be bound to target mRNA through its zinc finger domain. In mammalian,TTP destabilize the target mRNA by binding to the AU-rich element which locates on the mRNA 3＇ UTR. However,in fly,the physiological function of Tis11 is largely unknown. In order to further elucidate the physiological function of Tis1 1, we generated the Tis1 1 knockout fly. In this study, we generate Tis1 1 knockout fly by using P-element imprecise excision. P-element is one group of elements in fly genome which are comparable to bacterial transposons. P-elements can not only be inserted into genome,but also be precisely and imprecisely excised from genome, when transposase is present in cells. Since their discovery, Pelements have become a powerful tool of molecular biology to manipulate fly genome. We generated 700 P-element excision lines from P-element insertion line 19949. After screening and identification by PCR,we only get one line 279 containing partial sequence deletion in the second exon of Tis11 among the 700 lines. The start codon locates in the deletion part.