Amersham Rediprime II DNA Labeling System
Provides individually dispensed reaction mixes which are dried in the presence of a stabilizer and a dye to make labeling probes easier for the user.
Amersham Rediprime II DNA Labeling System
Provides individually dispensed reaction mixes which are dried in the presence of a stabilizer and a dye to make labeling probes easier for the user.
Overview
Rediprime II DNA Labeling System provides individually dispensed reaction mixes which are dried in the presence of a stabilizer and a dye to make labeling probes easier for the user. Rediprime reaction mixes have been formulated using an improved exonuclease-free Klenow to give probes with specific activity of 1.9 × 109 dpm/μg or greater after 10 minutes incubation at 37°C with the majority of DNA substrates. By replacing a nonradioactive nucleotide with the radiolabeled equivalent in the reaction mixture, newly synthesised DNA is made radioactive. Very small amounts of input DNA can be labeled, enabling very high specific activity probes to be produced with relatively small quantities of added nucleotides. These radioactive labeled fragments can then be used as sensitive hybridization probes for a wide range of filter-based applications.
- Long shelf-life and simple, fast protocol.
- Premixed, ambient-temperature-stable reactions with a color check for the labeling of DNA to specific activities of > 10⁹ dpm/µg.
- No need to remove unincorporated nucleotides before hybridization.
- DNA can be labeled in solution or in the presence of low melting point agarose.
- Labeling reaction complete in 10 min.
Rediprime II consists of premixed, individually dispensed, random-prime reactions, freeze-dried for ambient stability. Simply add substrate and [α-32P]dCTP. As little as 25 ng substrate DNA can be used. Suitable for use with dCTP only. Each system includes the following reagents, sufficient for 30 or 60 reactions (depending on product ordered): premixed, predispensed tubes containing 30× labeling mix and control DNA.
Feinberg and Vogelstein introduced the use of random sequence hexanucleotides to prime DNA synthesis on denatured template DNA at numerous sites along its length. The primer-template complex is a substrate for the Klenow fragment of DNA polymerase I. By replacing a nonradioactive nucleotide with the radiolabeled equivalent in the reaction mixture, newly synthesised DNA is made radioactive. Very small amounts of input DNA can be labeled, enabling very high specific activity probes to be produced with relatively small quantities of added nucleotides. These radioactive labeled fragments can then be used as sensitive hybridization probes for a wide range of filter-based applications.
