Objective       To construct  the differentially expressed cDNA libraries of aldehyde dehydrogenase with high and low activity (ALDH

high

/ALDH

low

) from tongue squamous carcinoma Tca8113 cell line.       Methods     

   Expression of stem cell marker ALDH was detected

and ALDH

high

and ALDH

low

cells were collected by Aldefluor assay combined with flow cytometry. Differentially expressed genes of total RNA that was extracted from the two cell subpopulations by Trizol were screened and amplified by suppressing subtractive hybridization (SSH)

and the PCR products were connected with pMD18-T vector and then transfected into E.coli DH5α for amplification. Enzyme digestion

gene sequencing and homology analysis were performed in 24 positive clones that were randomly picked from each library.       Results       Two subproportions of ALDH

high

and ALDH

low

were screened out

and ALDH

high

cells in Tca8113 cells accounted for 2.5%. RNA D(260)/D(280) of ALDH

high

and ALDH

low

were 1.93 and 1.92

respectively. Two-directional subtractive cDNA libraries of ALDH

high

and ALDH

low

were constructed

and each library comprised 500 clones. PCR analysis of 24 clones randomly picked from each library showed that insert-fragments distributed in 200-700 bp

and no false positive clones were detected. Gene sequencing result that was analyzed and indexed by PubMed showed that cancer related genes included SLC25A13

KLHL2

NPC1

WAPL

BARD1

Notch2 and EEF2K.       Conclusion        Two-directional subtractive cDNA libraries of ALDH

high

and ALDH

low

cells were successfully constructed.