[Application of oligonucleotide array in single nucleotide polymorphism typing of cytokines and cytokine receptors].
OBJECTIVE
To develop an oligonucleotide array for single nucleotide polymorphism (SNP) typing of cytokines, such as tumor necrosis factor (TNF)-a, interleukin (IL)-10, tumor growth factor (TGF)-betal, IL-4, and IL-6, and their receptors and evaluate its function by direct sequencing.


METHODS
According to relevant literature, SNP database of NCBI and SNP500 Cancer database of NCI, SNP loci and sequences of cytokines of clinical importance, TNF-a, IL-10, TGF-bl, IL-4 and IL-6, and matched cytokine receptors were chosen and 59 synthesized oligonucleotide probes were immobilized on a glass support, then the primer and Cy5-dCTP were used in multi-PCR, thus the products were labeled with Cy5. The labeled PCR products were hybridized with the probes in the array, and the signals were scanned by Scanner and then analyzed by Image software. Peripheral blood samples were collected from 80 healthy donors and 50 patients with uremia to isolate lymphocytes and DNA so as to undergo typing by this array, and the results were validated with direct sequencing.


RESULTS
All the samples with PCR products except for 10 from uremia patients had been genotyped by cytokine array successfully. No diversity was found in genotyping for three times. Incorrect locus was not found with direct sequencing.


CONCLUSIONS
With high specificity, sensitivity, repetitiveness, and throughout, and easy to manipulate, oligonucleotide array technique is an ideal molecular method for SNP genotyping of cytokine and cytokine receptor.
Guo
Yi-feng
YF
Shanghai First People's Hospital, Shanghai Organ Transplant and Research Center, Shanghai 200080, China.
Tan
Jian-ming
JM
Li
Rong-yu
RY
Liu
San-zhen
SZ
Li
Yao
Y
Ying
Kang
K
Xie
Yi
Y
Mao
Yu-min
YM
D016207:Cytokines / Q000235:genetics*; D006801:Humans; D020411:Oligonucleotide Array Sequence Analysis; D020641:Polymorphism, Single Nucleotide; D018121:Receptors, Cytokine / Q000235:genetics*
D016207:Cytokines; D018121:Receptors, Cytokine