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  1. DNA preparation by pulsed-field gel electrophoresis (isolation of yeast intact DNA) To prepare yeast DNA for electrophoresis, the cell walls of yeast cells are enzymatically disrupted and then suspended in melted low melting point agarose to ma
  2. The cloned PCR product is ligated into a T vector. DNA fragments amplified by Taq DNA polymerase PCR with a 3' prominent A base are efficiently cloned into T vectors that have an unpaired 3' T base complementary
  3. Flat-end cloning of PCR products After the target gene is amplified by PCR and the sample is purified and recovered as necessary, the next step is to use flat ends for ligation and cloning, whi
  4. Oligonucleotides and excess dNTP were removed from DNA amplification products by ultrafiltration. At the end of the PCR reaction, it is essential to remove oligonucleotide primers, primer dimers and dNTP from the amplified DNA product. This experiment is bas
  5. Restriction endonuclease digestion of DNA in agarose gel plugs Chromosomal DNA isolated from mammalian cells, yeast or bacteria can be digested in agarose plugs with the desired endonuclease. The low melting point agarose p
  6. Southern hybridization of radiolabeled probes with membrane-immobilized nucleic acids The signal intensity obtained by Southern hybridization depends on several factors, including the ratio of immobilized DNA complementary to the probe, the size
  7. Isolation of RNA according to size: agarose gel electrophoresis of glyoxalized RNA This method combines glyoxal denaturation and agarose gel electrophoresis (modified from that of McMaster and Carmichael (1977), Thomas (1983)). automated glyox
  8. RNA mapping with S1 nuclease Three different nucleases, S1 nuclease, RNAase, and exonuclease VII, are used to quantify RNA, to identify intron positions, and to characterize the positions o
  9. Dot hybridization and narrow line hybridization of purified RNA The dot and narrow-line hybridization technique (Kafatos et al. 1979) is used to immobilize several nucleic acid samples on the same solid-phase support (usuall
  10. Northern hybridization RNA samples transferred and immobilized to membranes can be hybridized to specific probes that can be used to localize the RNA of interest. Depending on the exp
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