Application

For random-primed labeling of DNA probes with DIG-11-dUTP, alkali-labile and color detection of DIG-labeled hybrids. DIG-labeled probes are used in a variety of hybridization techniques: ??Southern blots
??Northern blots
??Dot blots
??Colony and plaque screening
DIG-labeled hybrids are detected with an anti-DIG-alkaline phosphatase conjugate and the substrates NBT (nitroblue tetrazolium salt) and BCIP (5-bromo-4-chloro-3-indolyl phosphate, toluidinium salt), which give a light blue precipitate.

General description

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Convenient kit for random-primed labeling of DNA with digoxigenin-dUTP, alkali-labile, and color detection of hybrids by enzyme immunoassay. In this method, the complementary strand of denatured DNA is synthesized by Klenow polymerase using the 3′-OH termini of random oligonucleotides as primers.

Contents:
??Unlabeled Control DNA 1, 100 μg/ml
??Unlabeled Control DNA 2, 200 μg/ml
??DNA Dilution Buffer
??DIG-labeled Control DNA, 5.2 μg/ml
??10x Hexanucleotide Mix
??10x dNTP Labeling Mixture
??Klenow Enzyme, Labeling grade, 2 U/μl
??Anti-digoxigenin-AP-conjugate, 750 U/ml
??NBT/BCIP Concentrated Stock Solution
??Blocking Reagent?

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 10 components.

Preparation Note

Working concentration:?Dilution of Antibody 1:5,000

Specificity

Sensitivity and specificity: The standard labeling reaction with 1 μg template will give a yield of 260 ng DIG-labeled DNA after a 1-hour incubation at +37°C, or 780 ng after a 20-hour incubation. The sensitivity of DIG detection depends on the concentration of the DIG-labeled probe in the hybridization reaction, and the duration of the color reaction.