Construction and significance of directional expression cDNA library from human NB4 cells

Gang Chen, Wanggang Zhang, Jie Fu, Xingmei Cao, Wanhong Zhao, Yueheng Han, Aizhi Zhao, Fuyang Li, Xinping Liu, Libo Yao

Research output: Contribution to journalArticle

Abstract

Human acute premyeloid leukemia cell cDNA expression library was constructed to screen acute premyeloid leukemia tumor antigen. Total RNA and purified mRNA were extracted from human premyeloid cell line NB4. First and second strands of cDNA were synthesized by reverse transcription. After blunting, the cDNA fragments were ligated with EcoR I adapters. Then the cDNAs were digested with Xho I, and less than 400 bp cDNA fragment was removed by Sephacryl-S400 spin column, the remaining were ligated with λZAP vector. The recombinants were packaged in vitro, and a small portion of packaged phage was used to infect E. coli XL1-Blue-MRF' for titration. The recombinants were examined by color selection. In order to evaluate the size of cDNA inserts and the diversity of library, the pBK-CMV phagemid was excised from the ZAP express vector by using ExAssist helper phage with XLOLR strain, and then the pBK-CMV phagemid was digested by Xho I and EcoR I. The results showed that the NB4 cell line cDNA library consisting of 1. 65 × 10 6 recombinant bacteriophages was constructed with the recombinant ratio of 99.6 %. The average length of the recombinant exogenous inserts was about 1.7 kb. It was concluded that the constructed cDNA library are deserved to screen target clones.

Original languageEnglish (US)
JournalJournal of Huazhong University of Science and Technology - Medical Science
Volume24
Issue number1
StatePublished - 2004
Externally publishedYes

Fingerprint

Bacteriophages
Gene Library
Complementary DNA
Cells
Transcription
Antigens
RNA
Titration
Leukemia
Escherichia coli
Tumors
Cell Line
Color
Neoplasm Antigens
Libraries
Reverse Transcription
Clone Cells
cDNA
Messenger RNA
Recombinant

Keywords

  • Acute premyeloid leukemia
  • cDNA library
  • NB4 cells

ASJC Scopus subject areas

  • Management of Technology and Innovation
  • Medicine(all)

Cite this

Construction and significance of directional expression cDNA library from human NB4 cells. / Chen, Gang; Zhang, Wanggang; Fu, Jie; Cao, Xingmei; Zhao, Wanhong; Han, Yueheng; Zhao, Aizhi; Li, Fuyang; Liu, Xinping; Yao, Libo.

In: Journal of Huazhong University of Science and Technology - Medical Science, Vol. 24, No. 1, 2004.

Research output: Contribution to journalArticle

Chen, G, Zhang, W, Fu, J, Cao, X, Zhao, W, Han, Y, Zhao, A, Li, F, Liu, X & Yao, L 2004, 'Construction and significance of directional expression cDNA library from human NB4 cells', Journal of Huazhong University of Science and Technology - Medical Science, vol. 24, no. 1.
Chen, Gang ; Zhang, Wanggang ; Fu, Jie ; Cao, Xingmei ; Zhao, Wanhong ; Han, Yueheng ; Zhao, Aizhi ; Li, Fuyang ; Liu, Xinping ; Yao, Libo. / Construction and significance of directional expression cDNA library from human NB4 cells. In: Journal of Huazhong University of Science and Technology - Medical Science. 2004 ; Vol. 24, No. 1.
@article{5edb4c95bd5046ecb43057fb46786a7d,
title = "Construction and significance of directional expression cDNA library from human NB4 cells",
abstract = "Human acute premyeloid leukemia cell cDNA expression library was constructed to screen acute premyeloid leukemia tumor antigen. Total RNA and purified mRNA were extracted from human premyeloid cell line NB4. First and second strands of cDNA were synthesized by reverse transcription. After blunting, the cDNA fragments were ligated with EcoR I adapters. Then the cDNAs were digested with Xho I, and less than 400 bp cDNA fragment was removed by Sephacryl-S400 spin column, the remaining were ligated with λZAP vector. The recombinants were packaged in vitro, and a small portion of packaged phage was used to infect E. coli XL1-Blue-MRF' for titration. The recombinants were examined by color selection. In order to evaluate the size of cDNA inserts and the diversity of library, the pBK-CMV phagemid was excised from the ZAP express vector by using ExAssist helper phage with XLOLR strain, and then the pBK-CMV phagemid was digested by Xho I and EcoR I. The results showed that the NB4 cell line cDNA library consisting of 1. 65 × 10 6 recombinant bacteriophages was constructed with the recombinant ratio of 99.6 {\%}. The average length of the recombinant exogenous inserts was about 1.7 kb. It was concluded that the constructed cDNA library are deserved to screen target clones.",
keywords = "Acute premyeloid leukemia, cDNA library, NB4 cells",
author = "Gang Chen and Wanggang Zhang and Jie Fu and Xingmei Cao and Wanhong Zhao and Yueheng Han and Aizhi Zhao and Fuyang Li and Xinping Liu and Libo Yao",
year = "2004",
language = "English (US)",
volume = "24",
journal = "Current Medical Science",
issn = "2096-5230",
publisher = "Huazhong University of Science and Technology",
number = "1",

}

TY - JOUR

T1 - Construction and significance of directional expression cDNA library from human NB4 cells

AU - Chen, Gang

AU - Zhang, Wanggang

AU - Fu, Jie

AU - Cao, Xingmei

AU - Zhao, Wanhong

AU - Han, Yueheng

AU - Zhao, Aizhi

AU - Li, Fuyang

AU - Liu, Xinping

AU - Yao, Libo

PY - 2004

Y1 - 2004

N2 - Human acute premyeloid leukemia cell cDNA expression library was constructed to screen acute premyeloid leukemia tumor antigen. Total RNA and purified mRNA were extracted from human premyeloid cell line NB4. First and second strands of cDNA were synthesized by reverse transcription. After blunting, the cDNA fragments were ligated with EcoR I adapters. Then the cDNAs were digested with Xho I, and less than 400 bp cDNA fragment was removed by Sephacryl-S400 spin column, the remaining were ligated with λZAP vector. The recombinants were packaged in vitro, and a small portion of packaged phage was used to infect E. coli XL1-Blue-MRF' for titration. The recombinants were examined by color selection. In order to evaluate the size of cDNA inserts and the diversity of library, the pBK-CMV phagemid was excised from the ZAP express vector by using ExAssist helper phage with XLOLR strain, and then the pBK-CMV phagemid was digested by Xho I and EcoR I. The results showed that the NB4 cell line cDNA library consisting of 1. 65 × 10 6 recombinant bacteriophages was constructed with the recombinant ratio of 99.6 %. The average length of the recombinant exogenous inserts was about 1.7 kb. It was concluded that the constructed cDNA library are deserved to screen target clones.

AB - Human acute premyeloid leukemia cell cDNA expression library was constructed to screen acute premyeloid leukemia tumor antigen. Total RNA and purified mRNA were extracted from human premyeloid cell line NB4. First and second strands of cDNA were synthesized by reverse transcription. After blunting, the cDNA fragments were ligated with EcoR I adapters. Then the cDNAs were digested with Xho I, and less than 400 bp cDNA fragment was removed by Sephacryl-S400 spin column, the remaining were ligated with λZAP vector. The recombinants were packaged in vitro, and a small portion of packaged phage was used to infect E. coli XL1-Blue-MRF' for titration. The recombinants were examined by color selection. In order to evaluate the size of cDNA inserts and the diversity of library, the pBK-CMV phagemid was excised from the ZAP express vector by using ExAssist helper phage with XLOLR strain, and then the pBK-CMV phagemid was digested by Xho I and EcoR I. The results showed that the NB4 cell line cDNA library consisting of 1. 65 × 10 6 recombinant bacteriophages was constructed with the recombinant ratio of 99.6 %. The average length of the recombinant exogenous inserts was about 1.7 kb. It was concluded that the constructed cDNA library are deserved to screen target clones.

KW - Acute premyeloid leukemia

KW - cDNA library

KW - NB4 cells

UR - http://www.scopus.com/inward/record.url?scp=16544392742&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=16544392742&partnerID=8YFLogxK

M3 - Article

C2 - 15165115

AN - SCOPUS:16544392742

VL - 24

JO - Current Medical Science

JF - Current Medical Science

SN - 2096-5230

IS - 1

ER -