一个抗旱/抗寒基因过表达载体构建及转基因植株的筛选鉴定

Construction and Characterization of a Drought / Cold Tolerant Gene Overexpression Lines in Arabidopsis

[目的]进一步研究LWT1基因在调控低温和干旱应答中的功能,构建拟南芥LWT1基因过表达载体,获得相应的转基因株系。[方法]以野生型拟南芥的c DNA为模板,利用PCR扩增LWT1基因全长,将该基因连接到p ART27载体上。将获得的重组载体转化至农杆菌菌株GV3101,通过浸花转化法将LWT1重组载体转化到拟南芥野生型植株中,利用转基因筛选与遗传鉴定获得LWT1转基因阳性植株。[结果]LWT1基因CDS全长990 bp,PCR电泳结果一致,测序比对结果正确。通过抗性筛选和分子鉴定获得转基因植株。[结论]LWT1过表达载体构建成功并获得相应的转基因株系,为进一步研究该基因的分子机制奠定基础。

[ Objective J in order to further confirm that LWT1 gene is involved in the regulation of low temperature and drought stress responses, the transgenic lines of LWTI overexpression were generated. [ Method] The full-length LWT1 coding sequence was amplified through PCR from eDNA of Arabidopsis thaliana （Col-0） using specific primers and cloned into the pART27 vector. The resulting construct was transformed into Agrobacterium strain GV3101 for transformation into the Col-0 plants by the floral dip method. The transgenic lines were isolated through antibiot- ic screening and genetic identification. [ Result] The PCR product is 990 bp which is the same as the full-length LWTI CDS. The transgenic plants is gained and verified by genetic identification. [ Conclusion ] The vector was constructed successfully, and transgenic plants were ob- tained. The study will lay a foundation for further study.