11585592001

Roche

High Prime

form

solution

Quality Level

usage

sufficient for 50 labeling reactions

packaging

pkg of 200 μL

manufacturer/tradename

Roche

shipped in

dry ice

storage temp.

−20°C

General description

High Prime, a novel labeling mixture, contains optimal concentrations of nucleotides and primers in a highly efficient reaction mix with Klenow enzyme. This convenient "all-in-one" principle of High Prime reduces pipetting steps to a minimum, and increases accuracy and reproducibility of labeling reactions.
Enzyme and nucleotide mixture for rapid random-primed labeling of DNA with [32P], [35S], or [3H] dCTP. In this method, the complementary DNA strand of denatured DNA is synthesized by Klenow polymerase using the 3′-OH termini of the random oligonucleotides as primers.
Contents:
5x concentrated random primer mix: 1 U/μl Klenow polymerase, labeling grade, 0.125 mM dATP, 0.125 mM dGTP, 0.125 mM dTTP in 50% (v/v) glycerol.
High Prime is used for rapid random-primed labeling of DNA with [32P], [35S], or [3H]dCTP to a guaranteed specific activity of >2 x 109 dpm/μg. Incubation time is only 10 minutes at +37°C.
High Prime guarantees efficient labeling of:
  • DNA amounts ranging from 10 ng to 3 μg in a standard reaction
  • DNA of different lengths ranging from small restriction fragments to l or cosmid DNA
  • DNA, supercoiled or linearized
  • DNA in low melting-point agarose.
High Prime-labeled DNA probes are suitable for single-copy gene detection in mammalian DNA in Southern, northern, and dot blots, as well as in colony- or plaque hybridization.

Specificity

Heat inactivation: Stop the reaction by adding 2 μl 0.2 M EDTA (pH 8.0) and/or by heating to 65 °C for 10 minutes.

Application

For the labelling of DNA with radioactive dCTP using random oligonucleotides as primers.
High Prime-labelled probes are used in a variety of hybridization techniques: Southern blots
  • Northern blots
  • Dot/slot blots
  • Screening of gene libraries
  • In situ hybridizations

High Prime has been used to label the probes by random hexamer labelling and labelling of probes with 32P by random priming.

Quality

Function tested in the standard assay.

Principle

Labeled probes are generated with High Prime according to the random-primed labeling technique. High Prime is a specifically developed reaction mixture containing random oligonucleotides, Klenow polymerase, labeling grade, dATP, dGTP, and dTTP in an optimized reaction buffer concentrate in 50% glycerol for rapid and efficient labeling of DNA with [32P]-, [35S]-, or [3H]-labeled dCTP.

Analysis Note

Specific Activity: The standard assay routinely yields a specific activity of 2 x 109 dpm/μg, using different substrate DNAs after 10 minutes of incubation.
Assay Time: 30 minutes
Sample Materials
  • DNA fragments of at least 100 bp
  • Linearized plasmid, cosmid or λDNA
  • Supercoiled DNA
  • Or minimal amounts of DNA (10 ng), e.g., DNA restriction fragments isolated from gels or in molten agarose
Note: The length of the DNA to be labeled does not influence the reaction. Maximal incorporation may require a prolonged incubation period of 30 to 60 minutes.

Other Notes

For life science research only. Not for use in diagnostic procedures.

RIDADR

NONH for all modes of transport

WGK Germany

WGK 1

Flash Point(F)

No data available

Flash Point(C)

No data available

Certificate of Analysis

Certificate of Origin

Network plasticity of pluripotency transcription factors in embryonic stem cells
Filipczyk A, et al.
Nature Cell Biology, 17(10), 1235-1235 (2015)
Pof8 is a La-related protein and a constitutive component of telomerase in fission yeast
Paez-Moscoso DJ, et al.
Nature Communications, 9(1), 587-587 (2018)

Our team of scientists has experience in all areas of research including Life Science, Material Science, Chemical Synthesis, Chromatography, Analytical and many others.

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