[Construction and identification of the cDNA phage expression library for human colorectal cancer antigens].
OBJECTIVE
To construct a cDNA phage expression library for human colorectal carcinoma antigens.


METHODS
After the total RNA was extracted from human colorectal cancer tissues, the single-strand and double-strand cDNA were synthesized through reverse transcriptase PCR and long-distance PCR, with the cDNA fragments smaller than 500 bp removed and the remaining cDNA combined with the right and left arms of dephosphorylated lambdaTriplEx2 phage vector. The recombinant phage were then packaged in vitro by MaxPlax Packaging extract, and a small portion of the packaged phage was used to infect E.coli XL1-Blue. Titer measurement was performed so as to determine the capacity of the library. SfiI restriction endonucleases was used to cut the recombined phage DNA in order to identify the size of inserted cDNA.


RESULTS
The constructed cDNA phage expression library for human colorectal cancer antigens consisted of 2.39 x 10(6) pfu/ml bacteriophages with a recombination rate of 97.5% and the length of the inserted cDNA fragment ranged from 600 to 4,000 bp with an average of 1,400 bp.


CONCLUSIONS
The cDNA phage expression library of human colorectal cancer antigens is successfully constructed to meet the currently recognized standards, and can be well applicable in screening cDNA-cloned genes of human colorectal cancer-associated antigens by immunoscreening.
Liu
Yu-hu
YH
Institute for Digestive Diseases of PLA, Nanfang Hospital, First Military Medical University, Guangzhou 510515, China. liuyuhu@fimmu.edu.cn
Zhang
Zhen-shu
ZS
Zhong
Dong
D
Wu
Jin-bao
JB
Dan
Han-lei
HL
Yang
Xing-long
XL
Yang
Xiao-qiang
XQ
Chen
Xue-qing
XQ
Lai
Zhuo-sheng
ZS
Xiao
Bing
B
D000951:Antigens, Neoplasm / Q000235:genetics*; D001435:Bacteriophages / Q000235:genetics*; D015179:Colorectal Neoplasms / Q000276:immunology*; D015723:Gene Library; D006801:Humans
D000951:Antigens, Neoplasm