Gastric adenocarcinoma is one of the most common gastrointestinal malignant tumors. Due to the lack of efficient method for diagnosis in early gastric adenocarcinoma, doctors undergo operations on middle and terminal patients, which results in a poor prognosis. In this study, we employed cell-SELEX to screen aptamers in early gastric adenocarcinoma primary cells, which could provide new ideas for diagnosis of early gastric adenocacinoma. Early gastric adenocarcinoma primary cells were isolated from early gastric adenocarcinoma tissue, the ssDNA library with 88 bases in length containing a 52 base random sequence was synthesized in vitro, PCR conditions for ssDNA amplification were optimized, the products of PCR were dissociated by biotin-streptavidin magnetic beads system. After the repeative screenings, the aptamers specially bound to early gastric adenocarcinoma were obtained by cell-SELEX. After 12 cycles, the binding affinity between ssDNA library and early gastric adenocarcinoma has increased from 1560 to 4 336, indicating that aptamers with high affinity were enriched progressively. Then PCR products of the last round were cloned and sequenced. By analysis with DNAMAN70, we found that there were two sequences of C17 and C27 in the 30 clones utterly identified, which means a homology. Secondary structures prediction revealed that the stem-loop structures formed by ssDNA are possibly the action basis between aptamers and early gastric adenocarcinoma primary cells. In specificity, the fluorescence intensity of gastric adenocarcinoma primary cells were significantly different from that of normal gastric mucosal epithelium and the blank group (P<001). In affinity, the dissociation coefficient between aptamers and early gastric adenocaicinoma reached nmol level, indicating a high affinity. The aptamers specific to early gastric adenocarcinoma screened by cell-SELEX established the foundation for early diagnosis of gastric adenocarcinoma and the study of drug targets.
Key words
tomach neoplasms /
edenocarcinoma /
cell-SELEX /
aptamers /
random ssDNA library
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