One-tube post-PCR fluorescent labeling of DNA fragments.

One-tube post-PCR fluorescent labeling of DNA fragments.

A method for fluorescent postlabeling of PCR products has been developed. The method uses Klenow fragment of DNA polymerase I that exchanges the 3'-terminal residue of PCR-amplified DNA fragment for fluorescent nucleotides. All reactions, including PCR, are performed in one tube simply by successive addition of reagents. The products can be applied directly to fluorescence-based automated DNA s...

Multicolor post-PCR labeling of DNA fragments with fluorescent ddNTPs.

1.Meldrum, D. 2000. Automation for genomics, part two: sequencers, microarrays, and future trends. Genome Res. 10:12881303. 2.Orlando, C., P. Pinzani, and M. Pazzagli. 1998. Developments in quantitative PCR. Clin. Chem. Lab. Med. 36:255-269. 3.Rudi, K., S.L. Flateland, J.F. Hanssen, G. Bengtsson, and H. Nissen. 2002. Development and evaluation of a 16S rDNA array approach for describing complex...

Fluorescent Labeling of Antibody Fragments Using Split GFP

Antibody fragments are easily isolated from in vitro selection systems, such as phage and yeast display. Lacking the Fc portion of the antibody, they are usually labeled using small peptide tags recognized by antibodies. In this paper we present an efficient method to fluorescently label single chain Fvs (scFvs) using the split green fluorescent protein (GFP) system. A 13 amino acid tag, derive...

One-tube restriction enzyme digest and fluorescent labeling for restriction endonuclease fingerprinting single-strand conformational polymorphism.

PCR amplification of long DNA fragments.

The polymerase chain reaction (PCR) has recently evolved as a standard laboratory technique, popular in all areas of molecular biology research. However, the technique still has two limitations: the relatively low fidelity of Taq polymerase when compared with other polymerases (1), and its inability to efficiently amplify fragments higher than 3 kbp (2, 3). Although these two issues are irrelev...