Detection of expression of mouse MN/CA9 gene with reverse transcriptase-polymerase chain reaction / 吉林大学学报(医学版)

ABSTRACT

Objective To clone and analyze the MN/CA9 gene sequence in ICR mice and detect the expressions of MN/CA9 gene in various tissues of ICR mice.Methods The fresh tissues of small intesine, uterrus, skin, musle, liver, pancreas, heart, lung, thymus, spleen, kidney, ovary, stomach ,urinary bladder from ICR mice were obtained , the total RNA was extracted by GIT method, the 1st strand and 2nd strand of cDNA were synthesized, the EcoRⅠ adapters were lingated,the EcoRⅠ ends were phosphorylated, digested with XhoⅠ ,cDNA was ligated into the ZAP expression vector, packaged, planted, screened.The expressions of MN/CA 9 gene in various tissues in mice were detected by RT-PCR.Results A fragment of human MN/CA9 gene was used as probe, and 1.47?10~3 clones were screened with radioactive isotopic ~ 32 P labeled probe, after hybridizations, one positive signal of cDNA was detected and the complete nucleotide sequence contained 1 671 bp was determined (GenBank:AB086322), The nucleotide similarity between mouse and human cDNA (GenBank:Z54349) was 69.1%.The MN/CA9 gene detected by RT-PCR assay (primer: P521-P1193) strongly expressed in small intesine,uterus, musle, pancreas, heart, lung, thymus, spleen, kidney, ovary, stomach,and urinary bladder,meanwhile did not express in skin and liver. Conclusion The expressions of MN/CA9 gene in some tissues of ICR mice are similar to that of human, it can be used to further functional analysis of MN/CA9 gene.