[Cloning associated genes using microdissection-cDNA PCR-SSH in gastric dysplasia].
OBJECTIVE
To construct cDNA subtracted libraries from gastric dysplasia and further screen differentially expressed genes.


METHODS
Relatively pure dysplasia and normal tissue were procured by manual microdissection, and amplified by cDNA-PCR, which was used to carry on for suppression subtractive hybridization (SSH). Subtracted cDNA fragments were linked with vector, cloned, screened, sequenced, and made homologous search. Differentially expressed fragments were verified by dot hybridization.


RESULTS
Two subtracted cDNA libraries were constructed. Among 26 sequenced clones, 15 fragments corresponded to known genes, 3 fragments were known EST and 8 fragments were unknown EST (GenBank BQ164614-BQ164616, BQ291516-BQ291520). Fifteen fragments were verified to be differentially expressed in gastric dysplasia.


CONCLUSIONS
Subtracted cDNA libraries from gastric dysplasia are constructed using combination of microdissection-cDNA PCR and SSH setup in our laboratory. Some fragments have been screened and verified to help to search for novel associated genes with gastric carcinogenesis.
Hao
Dong-mei
DM
Department of Medical Genetics, China Medical University, Shenyang 110001, China.
Sun
Xiu-ju
XJ
Zheng
Zhi-hong
ZH
He
Guang
G
Ma
Ming-chao
MC
Xu
Hui-mian
HM
Wang
Mei-xian
MX
Sun
Kai-lai
KL
D003001:Cloning, Molecular; D018076:DNA, Complementary / Q000235:genetics; D020869:Gene Expression Profiling; D015972:Gene Expression Regulation, Neoplastic; D015723:Gene Library; D006801:Humans; D042282:Microdissection; D009693:Nucleic Acid Hybridization / Q000379:methods; D016133:Polymerase Chain Reaction; D011230:Precancerous Conditions / Q000235:genetics* / Q000473:pathology; D017422:Sequence Analysis, DNA; D013274:Stomach Neoplasms / Q000235:genetics* / Q000473:pathology
D018076:DNA, Complementary