5 μL of the RT-qPCR reaction was digested with 1 μL FastDigest PstI restriction enzyme (#FD0614, ThermoFisher Scientific) for 1 h at 37°C. Full-length amplicons and digestion products were separated on 1.5% agarose gel with ethidium bromide.
Fastdigest psti restriction enzyme
Manufactured by Thermo Fisher Scientific
FastDigest PstI is a restriction enzyme that recognizes and cleaves the DNA sequence 5'-CTGCAG-3'. It is a fast-acting enzyme designed to provide rapid DNA digestion for various molecular biology applications.
Automatically generated - may contain errors
Lab products found in correlation
3 protocols using fastdigest psti restriction enzyme
1
RT-qPCR Amplicon Restriction Digest
2
Plasmid Digestion and Sequencing
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Two micrograms of plasmid was cut with the FastDigest PstI restriction enzyme (Thermo Fisher), and the 900 bp piece was gel purified using a gel DNA extraction kit (Zymo). This piece was then recirculized in a 20 µL reaction containing 60 ng of gel purified DNA and 1 µL T4 DNA Ligase (Thermo Fisher) in buffer. Cutting and recircularizing was necessary to increase efficiency of later sequencing steps for unknown reasons. The ligation product was then amplified with primers containing the appropriate overhangs for high-throughput sequencing and then gel purified. This piece was then sequenced through paired-end sequencing using custom primers on an Illumina (San Diego, CA) sequencing machine (Miseq or Nextseq).
3
Plasmid Linearization and Sequencing
Check if the same lab product or an alternative is used in the 5 most similar protocols
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Two µ g of plasmid was cut with the FastDigest PstI restriction enzyme (Thermo Fisher) and the 900 bp piece was gel purified using a gel DNA extraction kit (Zymo). This piece was then recirculized in a 20 µL reaction containing 60 ng of gel purified DNA and 1 µ L T4 DNA Ligase (Thermo Fisher) in buffer. Cutting and recircularizing was necessary to increase efficiency of later sequencing steps for unknown reasons. The ligation product was then amplified with primers containing the appropriate overhangs for high throughput sequencing and then gel purified. This piece was then sequenced through paired end sequencing using custom primers on an Illumina (San Diego, CA) sequencing machine (Miseq or Nextseq).
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