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Screening and characterization of aptamers of chronic myelognous leukemia K562 cells

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Author:
No author available
Journal Title:
JOURNAL OF THE FOURTH MILITARY MEDICAL UNIVERSITY
Issue:
13
DOI:
10.3321/j.issn:1000-2790.2009.13.002
Key Word:
白血病,髓样,慢性;K562细胞;SELEX;适体;leukemia,myeloid,chronic;K562 cell;SELEX;aptamer

Abstract: AIM: To screen and characterize oligonucleotide aptamers of chronic myelognous leukemia K562 cells. METHODS: Oligonucleotide aptamers specifically binding to chronic myelog-nous leukemia K562 cells were screened from 88 nt random ssDNA library in vitro syntbesis by SELEX method, sub-library was prepared by biotin-streptavidin magnetic beads and neutro-phils from blood of normal humans were used as anti-sieve cells. The screened aptamters were purified and connected to pGEM-T plasmid vector and 24 clones of random aelection were sequenced after screening by the blue and white. The affinities of the screened aptamers binding to chronic myelognous leukemia K562 cells were detected by fluorescent primers. Homology analyses of the primary structure and secondary structure prediction to the screened aptamters were conducted with Clustal 2.05 and DNA sis V 2.5 software. RESULTS: After 13 rounds of screening cycles, the absorbance (A) of the screened aptamers to chronic myelog-nous leukemia K562 cells rose from 0.12 up to 1.25 and the ahaorhance(A) of the screened aptamers did not significantly increase until the 13 th round of the screening. No homologous sequences were found by analysis of the primary structure but they could be divided into 6 families. Respective conserved series was found in 5 of the 6 families and no conserved series was found in only one family. Secondary structure analysis showed that the stem-loops and bulges structure of the aptamters might be the structure foundation of specific binding to chronic myelognous leu-kemia K562 cells. CONCLUSION: We have successfully screened out the oligonucleotide aptamers of high-affinity binding to chronic myelognous leukemia K562 cells by SELEX method.

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