衣藻的核基因组转化及外源基因转录分析

Nuclear genome transformation and transcription analysis of exogenous gene in Chlamydomonas reinhardtii

通过珠磨法将衣藻表达载体pSP108转入到莱茵衣藻细胞壁缺陷型CC-400藻株中,经抗性筛选及PCR鉴定,获得了24个转化藻株.在抗性基因ble编码序列的中间部位和末端部位分别设计两对不同的探针引物,运用实时荧光定量PCR对转基因藻株进行外源基因转录水平的分析.结果发现:不同转化藻株外源基因的转录水平有明显差异;同一转化藻株两对探针引物,外源基因的转录水平也存在差异.说明莱茵衣藻在转化过程中,外源基因整合到基因组上的片段数量及片段长度和完整性具有不确定性.

Transformation of Chlamydomonas reinhardtii strain CC-400 with the plasmid pSP108 was carried out by glass beads method and twenty four transgenic monoclonal algae strains were successfully achieved by resistance selection and PCR identification.Two pairs of primers were designed at the middle and end part of the coding sequence of the exogenous gene ble.The transcription level of the transgenic monoclonal algae strains were detected by fluorescence quantitative PCR.The results showed that the transcription level of the exogenous gene ble in different transgenic monoclonal algae strains have obvious difference.In addition,the transcription level of exogenous gene ble in the same transgenic monoclonal algae strain was different though using two pairs of primers.That showed the copies,length and integrity of exogenous gene integrated into the genome were uncertainty.