Application

DIG-High Prime is used for the highly efficient random-primed labeling of DNA probes with DIG-11-dUTP, alkali-labile. DIG-labeled probes are generated at high yield within one hour or after overnight incubation. DIG-High Prime-labeled DNA probes are used in a variety of hybridization techniques: ??Southern blots
??Northern blots
??Dot blots
??Colony and plaque hybridizations
??For all types of filter hybridization
??For single-copy gene detection in total genomic DNA, even from organisms with high complexity, for example, human, barley, and wheat

The DIG High Prime DNA Labeling and Detection Starter Kit I uses digoxigenin (DIG), a steroid hapten, to label DNA probes for hybridization and subsequent color detection by enzyme immunoassay. The "random primed" DNA labeling method originally developed by Feinberg and Vogelstein is based on the hybridization of oligonucleotides of all possible sequences to the denatured DNA to be labeled. The input DNA serves as a template for the synthesis of labeled DNA, and is not degraded during the reaction, making it possible to label minimal amounts of DNA (10 ng) with this method. The complementary DNA strand is synthesized by Klenow polymerase using the 3′-OH termini of the random oligonucleotides as primers. Modified deoxyribonucleoside triphosphates, labeled with digoxigenin present in the reaction are incorporated into the newly synthesized complementary DNA strand.

General description

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Convenient kit for random-primed labeling of DNA templates with DIG-11-dUTP, alkali-labile, and color detection of the DIG-labeled hybrids. 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.This kit was assembled with convenience in mind, offering a ready-made blocking solution, combined stock solution of NBT/BCIP, and DIG Easy Hyb granules. The DIG-High Prime mixture includes stabilized Klenow Enzyme, nucleotides, primers, and reaction buffer, all in one convenient reagent.

Sample materials

??DNA fragments of at least 100 bp
??Linearized plasmid, cosmid or ?DNA
??Supercoiled DNA

Sigma Life Science is committed to bringing you Greener Alternative Products, which adhere to one or more of The 12 Principles of Greener Chemistry. This product is designed as a safer chemical.? The DIG System was established as a sensitive and cost-effective alternative to using radioactivity for the labeling and detection of nucleic acids. There are many available publications that prove the versatility of the DIG System, so use of radio-labeling is no longer the only option for labeling of DNA for hybridization.

Other Notes

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

Packaging

1 kit containing 7 components.

Quality

Function tested in a dot blot.