Technique / Molecular Biology / DNA analysis techniques / DNA extraction purification and quantification

Methods, protocols and troubleshooting on extraction, purification and quantification of high molecular weight genomic DNA, plasmid DNA, PCR products etc from tissue and cells.

DNA extraction and DNA purification methods new recommended protocol
Generally speaking, strategies for DNA extraction and DNA purification depends on your downstream applications, e.g. southern blot hybridization using high molecular weight genomic DNA, DNA fragments for subcloning, or DNA for PCR etc. Some considerations for ...

DNA extraction and DNA purification methods new recommended protocol
Factors affecting quality of DNA purification: 1. DNase: nuclease contamination in DNA preparation is the major concern for high quality DNA purification. So start with fresh tissue and to process tissue samples quickly is the key to lower the risk of DNA d ...

DNA extraction with phenol chloroform and ethanol precipitation new hot recommended protocol
This protocol uses phenol/chloroform (1:1) to extract DNA samples. Phenol/chloroform extraction can be repeated for two or more times until protein interface is invisible. Purified DNA can be concentrated by ethanol precipitation. (Alam lab, University of Hawa ...

Quantification of DNA using the Fluoroimager and Pico Green protocol
Quantification of DNA using the Fluoroimager and Pico Green (Protocol from genome lab at UC Davis). ...

YAC DNA preparation protocol
This protocol is used for YAC DNA extraction. YAC clones are provided as stab cultures in YPD agar. Glycerol stocks of each clone should be made as soon as possible. The clone will remain viable as a stab culture up to one week if stored at 4 c. (Hiroyuki Take ...

Genomic DNA extraction protocol new protocol
High molecular weight genomic DNA extraction protocol: Buffer and reagent: Genomic DNA extraction buffer (250ml): 1M Tris.Cl (pH 8.0) 2.5ml 0.5M EDTA (pH 8.0) 50 ml Pancreatic RNase 5 mg 10% SDS ...

DNA quantification new protocol
DNA quantification using spectometry: A general protocol for spectometry quantification of DNA in pdf. ...

• References
• Related Books
• Discussion
• Related news
• Job search

Preparation of genomic DNA from bacteria.
(Curr Protoc Mol Biol. 2001 Nov;Chapter 2:Unit 2.4.)
Wilson K.

Australian Institute of Marine Science, Townsville, Australia.

Most protocols for the preparation of bacterial genomic DNA consist of lysis,
followed by incubation with a nonspecific protease and a series of extractions
prior to precipitation of the nucleic acids. Such procedures effectively remove
contaminating proteins, but are not effective in removing exopolysaccharides
which can interfere with the activity of enzymes such as restriction
endonucleases and ligases. In this unit, however, the protease incubation is
followed by a CTAB extraction whereby CTAB complexes both with polysaccharides
and with residual protein, effectively removing both in the subsequent
emulsification and extraction. This procedure is effective in producing
digestible chromosomal DNA from a variety of gram-negative bacteria, all of which
normally produce large amounts of polysaccharides. If large amounts of
exceptionally clean DNA are required, the procedure can be scaled up and the DNA
purified on a CsCl gradient, as described in the alternate protocol.

Isolation and purification of large DNA restriction fragments from agarose gels.
(Curr Protoc Mol Biol. 2002 Aug;Chapter 2:Unit 2.6.)
Moore D, Dowhan D, Chory J, Ribaudo RK.

Baylor College of Medicine, Houston, Texas, USA.

This unit describes methods for recovering and purifying DNA restriction
fragments from agarose gels. The first protocol describes electroelution of the
fragment of interest from standard agarose gels using buffer-filled dialysis
bags, followed by concentration and purification using an Elutip column. This
approach can be used effectively for fragments of all sizes from 50 to 20,000 bp.
Electrophoresis directly onto NA-45 paper (second protocol) provides relatively
high yields for fragments <2000 bp. Fragments >1000 bp can also be separated on
low gelling/melting agarose gels and purified by phenol extraction (third
protocol), b-agarase digestion of the gel (first alternate protocol), or via
glass beads extraction (second alternate protocol). Removing linkers from a
fragment using a column rather than a gel is described, followed by a method for
estimating DNA concentrations in solution.

Purification and concentration of DNA from aqueous solutions.
(Curr Protoc Mol Biol. 2002 Aug;Chapter 2:Unit 2.1A.)
Moore D, Dowhan D.

Baylor College of Medicine, Houston, Texas, USA.

This unit presents basic procedures for manipulating solutions of single- or
double-stranded DNA through purification and concentration steps. These
techniques are useful when proteins or solute molecules need to be removed from
aqueous solutions, or when DNA solutions need to be concentrated. The , using
phenol extraction and ethanol (or isopropanol) precipitation, is appropriate for
purification of DNA from small volumes (<0.4 ml) at concentrations lower than 1
mg/ml. Three support protocols outline methods to buffer the phenol used in
extractions, concentrate DNA using butanol, and extract residual organic solvents
with ether. An alternative to these methods is nucleic acid purification using
glass beads and this is also presented. These protocols may also be used for
purifying RNA. The final two alternate protocols are used for concentrating RNA
and extracting and precipitating DNA from larger volumes and from dilute
solutions, and for removing low-molecular-weight oligonucleotides and
triphosphates.

DNA extraction protocol and method books
(Biowww bookshelf)
DNA extraction

Purification of small (100bp) PCR products
(Forum)
Small PCR fragment purification discussion