关键词: 逆转录病毒, 增强型绿色荧光蛋白, 胚盘细胞, 嵌合体, 鸡

Abstract:

【Objective】 Chicken chimeras were produced using chicken stage X blastoderm cells, which were infected by recombinant retrovirus in vivo. 【Method】 The recombinant expression vector pour-pBabe-EGFP was established by connecting expression vector and EGFP gene. The recombinant vector and packaging vector pVSVG were transfected into 293 10A1 cells together using calcium phosphate transfection to produce retrovirus. The concentrated retroviruses (200×) were injected into the central part of the subgerminal cavity to transfect the PGCs in eggs (not incubated). These eggs were incubated and then selected at random when they were hatched at 5.5 days. The genomic DNA were extracted and identified by polymerase chain reaction (PCR) and Southern blot analysis. 【Result】 The recombinant expression vector pour-pBabe-EGFP, which contained the full ORF, was established Successfully. The positive ratio was 75% by PCR analysis and 66.7% by Southern blot analysis. 【Conclusion】 The chicken chimeras were obtained successfully. Chicken chimeras can be produced using chicken blastoderm cells infected by recombinant retrovirus in vivo.

Key words: retrovirus, EGFP, blastoderm cells, chimeras, chicken