Cloning and expression of ornithine decarboxylase gene from human colorectal carcinoma

AIM:To construct and express ODC recombinant gene forfurther exploring its potential use in early diagnosis ofcolorectal carcinoma.METHODS:Total RNA was extracted from colon cancertissues and amplified by reverse-transcription PCR with twoprimers,which span the whole coding region of ODC.Thesynthesized ODC cDNA was cloned into vector pQE-30 atrestriction sites BamH I and Sal I which constitutedrecombinant expression plasmid pQE30-ODC.The sequenceof inserted fragment was confirmed by DNA sequencing,the fusion protein including 6His-tag was facilitated forpurification by Ni-NTA chromatographic column.RESULTS:ODC expression vector was constructed andconfirmed with restriction enzyme digestion and subsequentDNA sequencing.The DNA sequence matching on NCBI Blastshowed 99%affinity.The vector was transformed into E.coli M15 and expressed.The expressed ODC protein wasverified with Western blotting.CONCLUSION:The ODC prokaryote expression vector isconstructed and thus greatly facilitates to study the role ofODC in colorectal carcinoma.