Establishment of DNA Family Shuffling Method for Hirudin Genes

LONG Yin; LIU Jia-yun; LIU Li; WANG Zong-ren~

doi:10.13422/j.cnki.syfjx.2006.10.014

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Establishment of DNA Family Shuffling Method for Hirudin Genes

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- Establishment of DNA Family Shuffling Method for Hirudin Genes
- Chinese Journal of Experimental Traditional Medical Formulae Issue 10, Pages: 26-28(2006)
- 作者机构：
  
  1. 第四军医大学西京医院
  2. 第四军医大学西京医院陕西西安710032
- 作者简介：
- 基金信息：
- DOI：10.13422/j.cnki.syfjx.2006.10.014
  CLC： R284
- Published：2006
- 稿件说明：
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[1]龙铟,刘家云,刘莉,王宗仁.水蛭素基因DNA家族改组方法的建立[J].中国实验方剂学杂志,2006(10):26-28.

LONG Yin, LIU Jia-yun, LIU Li, et al. Establishment of DNA Family Shuffling Method for Hirudin Genes[J]. Chinese journal of experimental traditional medical formulae, 2006, (10): 26-28.
[1]龙铟,刘家云,刘莉,王宗仁.水蛭素基因DNA家族改组方法的建立[J].中国实验方剂学杂志,2006(10):26-28. DOI： 10.13422/j.cnki.syfjx.2006.10.014.

LONG Yin, LIU Jia-yun, LIU Li, et al. Establishment of DNA Family Shuffling Method for Hirudin Genes[J]. Chinese journal of experimental traditional medical formulae, 2006, (10): 26-28. DOI： 10.13422/j.cnki.syfjx.2006.10.014.

摘要

目的:运用DNA家族改组技术

增加改组分子库的多样性。方法:取水蛭素HV1、HV2、HV3基因等量混合

用DNaseⅠ消化

回收50bp左右的片段

再经无引物和有引物PCR扩增获得与原基因大小相同的改组分子

将其与载体连接获得水蛭素改组文库

DNA测序观察改组效果。结果:随机挑取3个克隆均为3种模板基因片段的杂合体

表明改组分子库多样性较好

改组方法较为成功。结论:水蛭素基因DNA家族改组方法的成功建立为进一步筛选高活性的水蛭素奠定了基础。

Abstract

Objective:To increase the diversity of hirudin-shuffled libraries by DNA family shuffling method.Methods: PCR products of HV1

HV2 and HV3 genes were combined and digested with DNase Ⅰ.Then random fragments about 50bp were purified and reassembled to the same size of hirudin genes by two amplifications of PCR with or without primers. Pools of hirudin mutant genes were ligated into phagemid pCANTAB-5E vector and three randomly picked clones were sequenced to observe the efficiency of shuffling.Results: The hirudin-shuffled libraries contained 1.2×10

different clones and the three clones were hybrids of three parent genes.Conclusion:These results show that a highly shuffled library of hirudin genes has been obtained which lay a substantial foundation for screening hirudin molecules with high bioactivity.

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LIU Li

LIU Jia-yun

LONG Yin

JIA Zhenhua

HAN Ningxin

YIN Yujie

CHANG Liping

Related Institution

Graduate School of Hebei Medical University

National Key Laboratory of Collateral Disease Research and Innovative Traditional Chinese Medicine （TCM）

Key Research Office of State Administration of TCM;Cerebrovascular Diseases

Hebei Academy of Integrated;Traditional Chinese and Western Medicine

School of Pharmacy，Shanxi Medical University

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