Elution of DNA from LMT agarose using organic extraction
- 최초 등록일
- 2014.09.13
- 최종 저작일
- 2014.08
- 2페이지/
한컴오피스
- 가격 1,000원
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목차
1. Introduction
2. Overview
3. Materials
4. Method
본문내용
Introduction
■ Agarose that has been modified by hydroxyethylation, a substitution that reduces the number of intrastrand hydrogen bonds, melts and sets at lower temperatures than standard agaroses.
■ The degree of substitution within the polysaccharide chain determines the exact melting and gelling temperature. Many commercial low-melting-temperature agarose can be held as liquids in the 30-35℃ range.
■ Enzymatic reactions (ligation, digestion, synthesis of radioactive probes) can be held in remelted gel. But, in general, polymerases, ligases, restriction enzymes work less efficiently in the presence of melted gel than in conventional aqueous buffer.
■ Rapid recovery of DNA from gels, bacterial transformation with nucleic acids in the remelted gel.
■ The protocol works best for DNA fragments ranging in size form 0.5 kb to 5.0 kb.
Overview
1. Separation of DNA according to size by electrophoresis through
low-melting-temperature agarose.
2. EtBR staining and UV light illumination -> cut out a slice of agarose with interest band.
3. Melting agarose and phenol:chloroform extraction
참고 자료
Sambrook and Russell, Molecular cloning 3rd, vol. 1: 5.29~32