Construction of cDNA libraries of renal papillary adenocarcinoma tissue and its side tissue with SMART technology

    CHENG Yu, LI Xu, CHEN Wei, YANG Yongª²Zong

    Medical Laboratory Center, First Hospital, Xi¡¯an Jiaotong University, Xi¡¯an 710061, China

    ¡¾Abstract¡¿ AIM: To construct the cDNA libraries of renal papillary adenocarcinoma tissue and its side tissue by SMART technology. METHODS: Switch mechanism at 5¡äend of mRNA template technology, SMART technology, was used to construct the libraries. The purified Poly(A)+RNA was used as template, Powerscript reverse transcriptase was used to transcribe and 5¡äª²oligo as a short extended template was added to the 5¡ä end of mRNA to enrich the fullª²length cDNAs. After amplification, the ds cDNA was digested by sfiI, sizeª²fractionated by columns (CHROMA SPINª²400) and then recombined to the ¦ËTripIEx2 vector. After package, the libraries were tittered, the rate of recombination ( blue/white) was determined and then the cDNA libraries were amplified. RESULTS: The titer of the cDNA libraries of renal papillary adenocarcinoma tissue and its side tissue were 1.4¡Á106 pfu¡¤mL-1 and 2.6¡Á106 pfu¡¤mL-1, respectively. After amplification, the rates increased to 6¡Á1011 pfu¡¤mL-1 and 9¡Á1011 pfu¡¤mL-1, respectively. CONCLUSION: Our successfully constructed cDNA libraries are fullª²length libraries with high efficiency and can be screened by probes and antibodies to find the associated genes of the renal papillary adenocarcinoma. ......