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Process for purification preparing DNA optical repairase

A photorepair enzyme and chromatographic purification technology, which is applied in the field of protein purification and preparation, can solve the problems of many steps, difficult activity detection, complicated post-purification treatment, etc., and achieve the effect of reducing separation steps and overcoming complicated steps

Inactive Publication Date: 2005-06-08
UNIV OF SCI & TECH OF CHINA
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Problems solved by technology

[0007] The present invention proposes a method for purifying and preparing DNA photorepair enzymes, in order to overcome the existing method with many steps, high price of purification chromatographic column matrix, complicated post-purification treatment, difficulty in activity detection, and unsuitability for large-scale batch preparation of DNA photorepair enzymes Defects

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  • Process for purification preparing DNA optical repairase
  • Process for purification preparing DNA optical repairase
  • Process for purification preparing DNA optical repairase

Examples

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Embodiment 1

[0029] Example 1: Purification and preparation of Escherichia coli DNA photorepair enzyme

[0030] 1. Obtaining Escherichia coli DNA Photorepair Enzyme with Histidine-Tag

[0031] Using the genomic DNA of Escherichia coli k12 strain as a template, and according to the general principle of DNA polymerase chain reaction primer design, the polymerase chain reaction primers of Escherichia coli DNA photorepair enzyme gene were designed.

[0032] Class 1 primers:

[0033] 5'cg cat atg aca act cac cta gtt tgg ttc cgc 3'

[0034] 5'cg cat atg acg aca cat ctc gta tgg ttt cgc 3'

[0035] 5'cg cat atg act acc cat ctg gtc tgg ttt cgc 3'

[0036] 5'cg cat atg acc acg cac ctt gtg tgg ttc cgc 3'

[0037] 2 types of primers:

[0038] 5'gtg ctc gag ttt ccc ctt ccg cgc cgc ctc a 3'

[0039] 5'gtg ctc gag ttc cct tta cca cgt cgt tta g 3'

[0040] 5'gtg ctc gag ttc cca cta ccc cgg cgt ttc t 3'

[0041] 5'gtg ctc gag ttt ccg tta cct cga cgc cta c 3'

[0042] Utilize each of these two type...

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Abstract

A process for preparing high-purity DNA photorepairase includes such steps as sing gentic engineering method to obtain the DNA photorepairase with His marker of colibacillus, and affinity chromatography by Ni-ion affinity chromatographic column. Its advantages are high purity (90%) short period, and low cost.

Description

Technical field: [0001] The invention belongs to the technical field of protein purification and preparation, and in particular relates to the purification and preparation of DNA photorepair enzyme by affinity chromatography and its activity detection technology. Background technique: [0002] The American "Journal of Biochemistry" (J.Biol.Chem.1984, 259, 6028-6032) pointed out that DNA photorepair enzyme is a protein with a molecular weight of about 55kD, and its main function is to repair the genome DNA of organisms formed by ultraviolet radiation. pyrimidine dimer. The United States "Nucleic Acids Res. 1994, 22, 4119-4124" pointed out that DNA photorepair enzymes were first discovered in some lower animals such as bacteria and insects, and then in higher plants, vertebrates and lower animals. Photorepair enzymes and their genes have been found in marsupial mammals, but so far no DNA photorepair enzymes have been found in higher mammals, including humans. [0003] The pu...

Claims

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Application Information

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IPC IPC(8): C07K1/22C12N9/00C12N15/52C12N15/63C12Q1/25C12Q1/68
Inventor 王玉珍章东方沐万孟俞书勤宋钦华罗昭锋于宙娄阳徐蕾
Owner UNIV OF SCI & TECH OF CHINA
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