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Products >  cDNA_Synthesis_and_Library_Construction >  Suppression_Subtractive_Hybridization >  PCR-Select_Kits

Subtractive Hybridization Kits

The PCR-Select cDNA Subtraction Kit offers an efficient method for selectively amplifying differentially expressed genes—those genes expressed in one mRNA population but reduced or absent in another (1–4). This method is particularly well-suited for the identification of target cDNAs that correspond to rare transcripts, typically the most difficult to obtain. In contrast to other methods that require physically separating single-stranded and double-stranded cDNAs, Clontech's PCR-Select method allows the exponential amplification of only the desired sequences. This method offers many significant advantages:

  • Straightforward method with only a few steps. With the PCR-Select method, subtraction occurs by one round of subtractive hybridization and selective amplification of differentially expressed genes, not by physical separation.
  • Over 1,000-fold enrichment of rare transcripts. This kit allows you to equalize transcript abundance and subtract in the same procedure, dramatically increasing the probability of obtaining differentially expressed rare transcripts.
  • Subtraction can be performed with just 2 µg of poly A+ RNA. This feature is especially useful when working with RNA samples that are difficult to obtain. If you have only nanograms of total RNA, generate high-quality cDNA for use in PCR-Select cDNA subtraction with the SMARTer Pico PCR cDNA Synthesis Kit.

Bacterial Genome Subtraction Kit

The Clontech PCR-Select Bacterial Genome Subtraction Kit offers an effective method for comparing bacterial genomes. In a matter of days, you can obtain a subtracted library of genomic sequences that are present in one bacterial strain but absent in another. This kit allows you to identify pathogenicity islands or other genomic DNA differences between two strains.

With the Clontech PCR-Select method, subtraction occurs in one round of subtractive hybridization and by selective amplification, not by physical separation of single-stranded DNA (1–3, 5). The Clontech PCR-Select Kit requires as little as 2 µg of each bacterial genomic DNA sample, with the procedure requiring only 2–3 days. It can be readily adapted to high-throughput sampling.

PCR-Select Differential Screening Kit

The Clontech PCR-Select Differential Screening Kit allows you to identify differentially expressed clones in a subtracted library (6–9). After generating pools of differentially expressed genes with the Clontech PCR-Select cDNA Subtraction Kit, use this kit to quickly confirm differential expression.

With the Clontech PCR-Select Differential Screening Kit, the subtracted library is hybridized with probes synthesized directly from tester and driver populations; a probe made from the subtracted cDNA, as well as a probe made from reverse-subtracted cDNA (a second subtraction performed in reverse). Clones that hybridize to tester but not driver probes are differentially expressed; however, nonsubtracted probes are not sensitive enough to detect rare messages. Subtracted probes are greatly enriched for differentially expressed cDNAs, but may give false positive results. Using both subtracted and non-subtracted probes provides the most effective way to identify differentially expressed genes.

  At-A-Glance   Documents   Images & Data   Resources

Features

  • Ideal for isolating novel, differentially expressed genes
  • Provides greater than 1,000-fold enrichment of rare transcripts
  • Requires only 2 µg of poly A+ RNA
  • cDNA subtraction process takes only 3–4 days

Applications

  • Subtractive hybridization
  • Gene identification
  • Differential gene expression analysis
  • Gene expression analysis
  • cDNA subtraction
  • Differential screening

References

  1. Diatchenko, L., et al. (1996) Proc. Natl. Acad. Sci. USA 93:6025–6030.
  2. Gurskaya, N. G., et al. (1996) Anal. Biochem. 240:90–97.
  3. PCR-Select cDNA Subtraction Kit (October 1995) Clontechniques X(4):2–5.
  4. Diatchenko, L., et al. (1998) In RT-PCR Methods for Gene Cloning and Analysis, Eds. Siebert, P. D., et al. (BioTechniques Books, MA), pp. 213–239.
  5. Akopyants, N. S., et al. (1998) Proc. Natl. Acad. Sci. USA 95:13108–13113.
  6. Wang, Z. & Brown, D. D. (1991) Proc. Natl. Acad. Sci. USA 88:11505–11509.
  7. Jin, H., et al. (1997) BioTechniques 23:1084–1086.
  8. Diatchenko, L., et al. (1998) In RT-PCR Methods for Gene Cloning and Analysis, Eds. Siebert, P. D., et al. (BioTechniques Books, MA), pp.213–239.
  9. Diatchenko, L., et al. (1998) In Methods in Enzymology, Ed. Weissman, S. M., 303:349–380.

Additional Information

Please see the product's Certificate of Analysis for information about storage conditions, product components, and technical specifications. Please see the Kit Components List to determine kit components. Certificates of Analysis and Kit Components Lists are located under the Documents tab.


 
 
Products
Cat. # Product Contents Size Price License Units Select
637404 Clontech PCR-Select™ Bacterial Genome Subtraction Kit 7 Rxns $929.00 License Statements
637403 Clontech PCR-Select™ Differential Screening Kit Each $470.00  
637401 Clontech PCR-Select™ cDNA Subtraction Kit 7 Rxns $929.00 License Statements
637402 PCR-Select™ Differential Screening Blocking Solution 1 mL $256.00  
 

 

Required Products
Cat. # Product Contents Size Price # of Units Select
639201 Advantage® 2 Polymerase Mix 100 Rxns $245.00
639206 Advantage® 2 PCR Kit 100 Rxns $425.00
639250 TaqStart® Antibody 200 Rxns $190.00
 

 

 
 

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