Phenol chloroform isoamyl alcohol
Phenol/chloroform/isoamyl alcohol is a liquid solution used for the extraction and purification of nucleic acids, such as DNA and RNA, from biological samples. It is a common reagent used in molecular biology and genomics applications.
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18 protocols using phenol chloroform isoamyl alcohol
Genomic DNA Extraction and PCR Genotyping
DNA Extraction from Seaweed Samples
DNA Purification from Enzyme-Treated Mixtures
Mitochondrial DNA Mutation Analysis
In vitro Reconstitution of m1A947 in Human Mitochondrial 16S rRNA
The 114-mer RNA segment including Helix 71 (G866-U979) of human mitochondrial 16S rRNA was transcribed in vitro using T7 RNA polymerase. Template DNA with T7 class III promoter for the 114-mer RNA segment was prepared by assembling the following DNA sequences:
5’-gctaatacgactcactataggcaccgcctgcccagtgacacatgtttaacggc-3’,
5’-gtgacacatgtttaacggccgcggtaccctaaccgtgcaaaggtagcataatcac-3’,
5’-gtgcaaaggtagcataatcacttgttccttaaatagggacctgtatgaatggctccacgagggtt-3’,
and 5’-aaccctcgtggagccattc-3’. In vitro transcription by T7 RNA polymerase was performed as described [40 (link)]. The transcript was purified by denaturing PAGE and quantified by measuring the optical density at 260 nm. The reaction mix (50 μL), consisting of 25 mM Hepes-KOH (7.5), 100 mM KCl, 2.5 mM MgCl2, 1 mM DTT, 1 mM Ado-Met, 1.5 μg total RNA, and 1 μM His-tagged TRMT61B, was incubated for 2 h at 37°C, followed by adding phenol-chloroform isoamylalcohol (Nacalai) to terminate the reaction. Total RNA was recovered by ethanol precipitation and subjected to primer extension as described above.
In Vitro Reconstitution of tRNA t6A37
For kinetic analyses, initial velocity of t6A37 formation was measured at 37 °C in a 10-μL reaction mixture (for each data point) containing 100 mM Tricine-NaOH (pH 8.0), 2 mM DTT, 15 mM MgCl2, 18.75–150 μM ATP, 14.275–114.2 μM [14C] L-Thr, 5–40 mM NaHCO3, 0.4–3.2 μM mt-tRNA, 1 μM YRDC (17–279a.a.), and 2 μM OSGEPL1 (35–414a.a.). To stop the reaction, the mixture was spotted on a Whatman 3MM filter paper, soaked in 5% TCA, washed three times with 5% TCA, and rinsed with ice-cold ethanol. Radioactivity of [14C] L-Thr on the dried paper was measured by liquid scintillation counting on a Tri-Carb 2810TR (PerkinElmer).
Genotyping mRFP1 and BAT Hmgcr KO Mice
Genomic DNA Extraction from Fish Fins
DNA Extraction from Mucosal Swabs
Chromatin Immunoprecipitation in Yeast
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