Cloning, Identification, Computor-based Location And Expression Of A Novel CCR-L2/FXYD6 Gene Related To Cholangiocarcinoma

This study was designed to clone a novel Cholangiocarcinoma related and FXYD6 homologous gene- containing L2 cDNA (CCR-L2/FXYD6) for the first time .A novel database of fresh Cholangiocarcinoma tissues was made in the PLA hospital. Sequence 4 (144bp) was cloned which contains the part sequence (120bp) of L2 cDNA according to the designed primers (Pl and P2).The genomic sequence of the candidate unigene on Chromosome 11 was found and downloaded by homology searching of L2 cDNA in public databases. Bioinformational methods were used to analyze the sequence of candidate unigene by GOLDKEY Sequence Analyzing Software, and two pairs of primers (P3/P4 and P5/P6) were designed. The reverse transcription-coupled polymerase chain reaction (RT-PCR), molecular cloning, restriction endonuclease digestion, andDNA sequencing technique were performed on total RNA isolated from normal bile duet and cholangiocarcinoma tissues. The expressional difference of CCR-L2/FXYD6 between nonnal and malignant bile duet tissues was also identified by RT-PCR method. A 540bp product was acquired and cloned in pGEM-T plasmid vector. The sequence of the clone(WT5) represents one part of CCR-L2/FXYD6 gene as predicted, which was located on chromosome llq. So a novel sequence of Unigene CCR-L2/FXYD6 was cloned, and the correlationship between the gene and cholangiocarcinoma was observed, which are high expressed in the cholangiocarcinoma samples and low expressed in nonnal bile duet tissue.Further study should be performed on the novel gene sequence and the mechanism of cholangiocarcinoma.