鳞头犬牙南极鱼atp6v0c基因在HeLa细胞中抗寒功能的研究

Cloning of atp6v0c gene from Antarctic notothenioid fish Dissostichus mawsoni and identification of its cold tolerance in He La cell

为探求南极鱼ATP酶类在低温适应下的作用,从鳞头犬牙南极鱼Dissostichus mawsoni的c DNA文库中克隆atp6v0c基因(dmatp6v0c),并转染到He La细胞中,通过流式细胞仪检测细胞在低温胁迫下(10℃)的死亡率,获得了dmatp6v0c基因的全部编码序列,其长度为462 bp,并将其插入到真核表达载体pc DNA3.1(-)上用于体外表达。结果表明,与对照组相比,在低温胁迫下,过表达dmatp6v0c基因的He La细胞死亡率从(19.23±1.87)%显著降低到(8.13±0.04)%(P<0.05)。研究表明,在低温胁迫下dmatp6v0c基因过表达能显著降低He La细胞的死亡率,试验结果可为dmatp6v0c基因作为鱼类耐寒育种的候选基因提供理论依据。

The atpev0c gene of Antaratic notothenioid fish Dissostichus mawsoni was cloned from Antarctic notothe- nioid fish eDNA library, and then transfected the dmatp6vOc gene obtained into the HeLa cell to detect cell viability under cold stress （ 10 ℃ ） by flow cytometry to investigate the ATPase responsive for the cold adaptation in Antarc- tic notothenioid fish. The complete coding sequence with 462 bp of dmatp6vOc gene was cloned and sequenced, and the dmatp6vOc coding sequence was inserted into the pcDNA3.1 （-） vector by XhoI and BamHI restriction sites for gene expression in HeLa cell. The results showed that over-expression of dmatp6vOc gene in HeLa cell was found to reduce the cell mortality from 19.23% ±1.87% to 8.13% ±0.04% （P〈0.05） under cold stress com- pared with the control group. The findings indicated that the expression level of dmatp6vOc was heavily involved in cell viability under cold stress and that dmatp6vOc could be considered as a candidate for breeding cold resistant fish.