Pages 30-35, Language: EnglishZhang, Ping / Xu, Qin / Chen, Wan Tao / Zhang, Zhi Yuan / Li, Jin Zhong / Zhou, Xiao JianObjective: To construct a cDNA library of human tongue squamous cell carcinoma cell line (Tca8113) for further study of protein-protein interaction in oral squamous cell carcinoma cells for exploring the mechanisms of carcinogenesis.
Methods: Total RNA was extracted and reverse-transcribed into cDNA, followed by long distance PCR. The product was co-transformed into the competent AH109 with Sma I-linearised pGADT7-Rec. After growing on SD/-Leu plates, these transformants were harvested and the quality of the constructed library was analysed.
Results: The transformation efficiency was 1 × 106 transformants/3 µg pGADT7-Rec. The library size was 4 × 107 cfu/ml. The inserts were from 500 bp to 3 kb. With the antibody of HAtag of pGADT7-Rec, the corresponding fusion proteins expressing in AH109 were detected.
Conclusion: A two-hybrid cDNA library from Tca8113 has been constructed. Analysis of the quality of this two-hybrid cDNA library indicated that this library would be useful for identifying protein-protein interactions in oral squamous cell carcinoma.
Keywords: cDNA library, squamous cell carcinoma cell line, yeast two-hybrid
