Storage: -20C
UNSPSC Code: 41106000
General description: 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.-á 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.
General description: Assay Time: 145minutesSample Materials. Linearized plasmid DNA. PCR product PrincipleThe DIG RNA Labeling Kit produces DIG-labeled, single-stranded RNA probes of known length. Either SP6 or T7 RNA polymerase transcribes these probes in vitro from template DNA (in the presence of digoxigenin-UTP).RNA Labeling by in vitro TranscriptionThe DNA to be transcribed is cloned into the polylinker site of appropriate transcription vectors (e.g., pSPT 18 or 19), which contain promoters for SP6 and T7 RNA polymerases. Adjacent template DNA is linearized at a suitable site and the RNA polymerases are used to produce "run off" transcripts. DIG-UTP is incorporated into the transcript. Every 20 to 25th nucleotide of the newly synthesized RNA is a DIG-UTP. Since the nucleotide concentration does not become limiting in the standard transcription reaction, this reaction can generate large amounts of labeled RNA.
General description: Kit for the labeling of RNA with digoxigenin-UTP by in vitro transcription with SP6 and T7 polymerases. By this method, single-stranded RNA probes of known length are produced, which can be used in a variety of hybridization techniques.
Application: For RNA labeling with DIG-11-UTP by in vitro transcription with SP6 and T7 RNA Polymerases. DIG-labeled "run off" transcripts are synthesized with high efficiency and can be used in a variety of hybridization techniques: . Northern blots. Southern blots. In situ hybridizations. Plaque or colony lifts. RNase protection experimentsDue to highly specific and sensitive detection systems, DIG-labeled probes can be used for single-copy gene detection in 1++g total human DNA.Note: Since the linkage between DIG and UTP is resistant to alkali, DIG-labeled RNA can be fragmented by alkaline treatment. Slightly reducing the size of the DIG-labeled RNA probe may make it more suitable for certain applications in in situ hybridization.
Quality: Test principle: The DNA template to be transcribed is cloned into the polylinker site of appropriate transcription vectors, which contain promoters for SP6 and/or T7 RNA Polymerases. After linearization at a suitable site, RNA is transcribed in the presence of DIG-UTP. Under standard conditions, approximately 10mug of full-length DIG-labeled RNA is transcribed from 1mug template.
Specificity: Sensitivity and SpecificityDIG-labeled RNA probes can detect single-copy genes in as little as 1 mug of mammalian DNA under the following assay conditions: The hybridization mix contains 20 to 100 ng labeled probe/ ml, and the bound probe is detected with anti-DIG-AP and visualized with the chemiluminescent substrate CDP-Star.Heat inactivation: Stop the reaction by adding 2 mul 0.2 M EDTA (pH 8.0).
Other Notes: For life science research only. Not for use in diagnostic procedures.
Components: pSPT18 DNA 0.25 mg/ml; pSPT19 DNA 0.25 mg/ml; Control DNA 1, pSPT18-Neo, cleaved with Pvu II 0.25 mg/ml; Control DNA 2, pSPT19-Neo, cleaved with Pvu II 0.25 mg/ml; DIG-labeled Control RNA, DIG-labeled "antisense" neo RNA 100 ng/µl; Unlabeled Control RNA, neo poly (A) "sense" RNA 200 µg/ml; NTP Labeling Mixture 10x concentrated; Transcription Buffer 10x concentrated; DNase I, RNase-free 10 U/µl; Protector RNase Inhibitor 20 U/µl; SP6 RNA Polymerase 20 U/µl; T7 RNA Polymerase 20 U/µl
RIDADR: NONH for all modes of transport