Neutral phenol chloroform isoamyl alcohol

Manufactured by Thermo Fisher Scientific

Neutral phenol–chloroform-isoamyl alcohol is a solution used in the extraction and purification of nucleic acids, such as DNA and RNA, from biological samples. It is a mixture of phenol, chloroform, and isoamyl alcohol, and is commonly used in the standard phenol-chloroform extraction method.

Automatically generated - may contain errors

Lab products found in correlation

Nanodrop 2000, by Thermo Fisher Scientific (2 mentions) Acid phenol chloroform isoamyl alcohol, by Thermo Fisher Scientific (2 mentions) 0.1 mm glass beads, by Biospec (1 mentions) Miseq platform, by Illumina (1 mentions) Qiaquick pcr purification column, by Qiagen (1 mentions) Sds 20, by Thermo Fisher Scientific (1 mentions) Platinum hot start pcr master mix, by Thermo Fisher Scientific (1 mentions) Hiseq 2000 sequencer, by Illumina (1 mentions) Truseq stranded total rna library kit, by Illumina (1 mentions) Truseq small rna library kit, by Illumina (1 mentions) Turbo dnase, by Thermo Fisher Scientific (1 mentions)

Close spelling variants of this product

Chloroform (747 citations) Phenol/chloroform/isoamyl alcohol (121 citations) Phenol (68 citations) Phenol/chloroform (56 citations) Isoamyl alcohol (6 citations) Phenol:chloroform:isoamyl (3 citations)

4 protocols using neutral phenol chloroform isoamyl alcohol

Total RNA Extraction from Bacterial Cultures

Check if the same lab product or an alternative is used in the 5 most similar protocols

Total RNA was extracted from exponential phase cultures growing either in LB or in MOPS EZ rich defined medium using hot phenol lysis method described previously (Massé et al. 2003 (link)). Briefly 700 µL of samples were removed from growing cultures and added to a mixture containing 800 µL of acid phenol–chloroform-isoamyl alcohol (pH of 4.3; Fisher Scientific) and 100 µL of lysis buffer (320 mM sodium acetate [pH 4.6], 8% SDS, 16 mM EDTA) equilibrated to 65°C. Samples were mixed at 65°C for 5 min and centrifuged for 30 min at 4°C to separate phases. The upper aqueous phase was extracted a second time with equal volume of neutral phenol–chloroform-isoamyl alcohol (pH of 6.7; Fisher Scientific). RNA was alcohol-precipitated and resuspended in DEPC-treated water. RNA concentration was measured using Nano Drop 2000 (Thermo Fisher Scientific).

Sinha D., Matz L.M., Cameron T.A, & De Lay N.R. (2018). Poly(A) polymerase is required for RyhB sRNA stability and function in Escherichia coli. RNA, 24(11), 1496-1511.

+ Open protocol

+ Expand

RNA Stability Determination Protocol

Cited 1 time

Check if the same lab product or an alternative is used in the 5 most similar protocols

To determine RNA stabilities, cultures were grown in BHI to exponential phase (OD₆₂₀ ≈ 0.15) as described above, and a culture sample (0 h after the end of log-phase growth [T₀]) was collected. Rifampin was added to inhibit transcription, and additional samples were collected 5, 10, 20, and 30 min after rifampin addition. All samples were subjected to hot phenol lysis as described previously (80 (link)). Briefly, 700 μl of sample was added to a mixture containing 800 μl of acid phenol–chloroform-isoamyl alcohol, pH 4.3 (Fisher Scientific), and 100 μl of lysis buffer (320 mM sodium acetate [pH 4.6], 8% [wt/vol] SDS, and 16 mM EDTA) equilibrated to 65°C. Samples were mixed at 65°C for 5 min and centrifuged for 30 min at 4°C to separate phases. The upper aqueous phase was extracted a second time with an equal volume of neutral phenol–chloroform-isoamyl alcohol, pH 6.7 (Fisher Scientific). RNA was ethanol precipitated and resuspended in DEPC-treated water. RNA concentration was measured using a NanoDrop 2000 (Thermo Fisher Scientific).

Sinha D., Frick J.P., Clemons K., Winkler M.E, & De Lay N.R. (2021). Pivotal Roles for Ribonucleases in Streptococcus pneumoniae Pathogenesis. mBio, 12(5), e02385-21.

+ Open protocol

+ Expand

Gut Microbiome Profiling from Fecal and Intestinal Samples

Cited 1 time

Check if the same lab product or an alternative is used in the 5 most similar protocols

For DNA from human fecal samples, ~200 mg (average wet weight) fecal sample was suspended in 600μl PBS. For mouse intestinal samples, tissues were dissected, homogenized in 5mL PBS, and 500μl of the homogenate used for DNA extraction. ~500μl 0.1mm glass beads (BioSpec), 210μl SDS %20, and 500μl neutral phenol:chloroform:isoamyl-alcohol (24:24:1, Fisher Scientific) were added to each sample, and samples were lysed by bead-beating followed by ethanol precipitation (Hsiao et al., 2014 (link)).
The V4 variable region of bacterial 16S ribosomal RNA genes was amplified in 25μl total volume reactions comprising 1μl of extracted DNA as template, 10μl Platinum Hot Start PCR Master Mix (ThermoFisher), 13μl PCR-grade water and 0.5μl of forward and reverse primers (10μM). Cycling conditions were 94°C for 3 min, followed by 33 cycles (94°C for 45 sec, 50°C for 60 sec, 72°C for 90 sec), and 72°C for 10 min. An equal amount of each amplicon (~240ng) was pooled into libraries, which were then purified using QIAquick PCR purification columns (Qiagen) and subjected to sequencing using the Illumina MiSeq platform. Paired-end 150nt reads were assembled, de-multiplexed, rarefied to >900 reads per sample, and analyzed using the QIIME 1.9.1 software package (Caporaso et al., 2010 (link)). Sequencing run results are summarized in Table S2A and S3.

Alavi S., Mitchell J.D., Cho J.Y., Liu R., Macbeth J.C, & Hsiao A. (2020). Interpersonal Gut Microbiome Variation Drives Susceptibility and Resistance to Cholera Infection. Cell, 181(7), 1533-1546.e13.

+ Open protocol

+ Expand

Genome-wide Transcriptome Profiling of Bacteria

Check if the same lab product or an alternative is used in the 5 most similar protocols

Total RNA samples collected from cultures at an OD₆₀₀ of 1.0 were subjected to DNase treatment (DNase Turbo; Ambion) following the manufacturer's protocol. Sample mixtures (total reaction volume of 100 μl) were incubated for 1 h at 37°C and the reaction was stopped by addition of 100 μl of DEPC-treated water and 200 μl of neutral phenol–chloroform–isoamyl alcohol (Fisher Scientific). DNase-treated RNA samples were phenol extracted, alcohol precipitated, and RNA concentration was measured. 5 μg of DNase-treated RNA was subjected to ribosomal RNA removal (RiboZero™ rRNA Removal for Gram-negative Bacteria, Illumina). For mRNA sequencing, libraries were constructed by Macrogen (Rockville, MD) from the rRNA-depleted samples using the Illumina TruSeq Stranded Total RNA library kit. For short RNA sequencing of total RNA and RNA immunoprecipitated with Hfq, rRNA-depleted samples were subjected to RNA fragmentation using the Ambion RNA fragmentation kit (AM8740) followed by RNA 5′ polyphosphatase treatment (Epicenter), which was performed to facilitate 5′ adapter ligation. Libraries were generated by Macrogen (Rockville, MD, USA) using the TruSeq Small RNA library kit (Illumina). All high throughput RNA sequencing was performed using 100 bp paired end read sequencing with an Illumina HiSeq2000 sequencer.

Cameron T.A., Matz L.M., Sinha D, & De Lay N.R. (2019). Polynucleotide phosphorylase promotes the stability and function of Hfq-binding sRNAs by degrading target mRNA-derived fragments. Nucleic Acids Research, 47(16), 8821-8837.

+ Open protocol

+ Expand

About PubCompare

Our mission is to provide scientists with the largest repository of trustworthy protocols and intelligent analytical tools, thereby offering them extensive information to design robust protocols aimed at minimizing the risk of failures.

We believe that the most crucial aspect is to grant scientists access to a wide range of reliable sources and new useful tools that surpass human capabilities.

However, we trust in allowing scientists to determine how to construct their own protocols based on this information, as they are the experts in their field.

Ready to get started?

Revolutionizing how scientists
search and build protocols!