Buffer 2

Manufactured by Thermo Fisher Scientific

Sourced in United States

Buffer II is a laboratory reagent designed to maintain the pH and ionic strength of solutions used in various biochemical applications. It is a commonly used buffer solution that helps to create a stable environment for biomolecules, such as proteins and nucleic acids, during experimental procedures.

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Lab products found in correlation

Close spelling variants of this product

AccuPrime PCR Buffer II (15 citations) PCR Buffer II (12 citations) Novex™ Tris-Glycine SDS Sample Buffer (2×) (9 citations) GeneAmp 10X PCR Buffer II (5 citations) 2-point standard buffers (4 citations) Quantigene 2.0 homogenizing buffer (4 citations) 4‐(2‐hydroxyethyl)‐1‐piperazineethanesulfonic acid buffer (4 citations) Environmental Master Mix (version 2.0) PCR buffer (3 citations) SPE buffer-2 (3 citations) NuPAGE 2× LDS sample buffer (3 citations)

5 protocols using buffer 2

DNA Extraction and 16S rRNA Sequencing

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The purified DNA was adjusted to 10~200 ng/μl, and PCR was carried out using with Buffer II (Thermo Fisher Scientific, USA). The PCR primer was 5’Balspec 16S (5′-CGCATGTATGAAAGAAGACCA-3′) and Balspec 16Sr 610 (5′-CCCCTTTTTAACTCTAGTCATATAGT-3′), with an expected product of 230 bp (Itoh et al. 2015 (link)). PCR conditions were 35 cycles of thermal denaturation at 95 °C for 30 s, annealing at 50 °C for 30 s, and elongation at 72 °C for 45 s. After PCR, bands were confirmed on a 1.2% agarose gel by electrophoresis, and then excised for sequencing by TA cloning using a Mighty TA-cloning kit (Takara Bio, Japan). The gene sequence was decoded using a model 3130 genetic analyzer (Applied Biosystems, USA) and analyzed using MEGA Ver. 7 software.
The 5′Balspec16S (5′-CGCATGTATGAAGAAGACCA-3′) and 3′Balspec 16S (5′-TTACCTATATAATTGTCGATACCA-3′) primer were also used for AHB strain sequencing, with an expected product of 1075 bp (Tavares et al. 2006 (link)).

Yamanouchi K., Arima H., Sakamoto Y., Kanto K., Kasai K., Ito K, & Inaba T. (2018). First report of the isolation of Balamuthia mandrillaris in the northern region of Japan. Parasitology Research, 117(9), 2895-2900.

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TCR Repertoire Analysis Protocol

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Genomic DNA was directly isolated from frozen sorted and unsorted cells as described above, except the phenol extraction was omitted. Vβ-Jβ rearrangements were amplified using TCRB tube A (Invivoscribe), Amplitaq GOLD (Thermo Fisher Scientific), and Buffer II (Thermo Fisher Scientific) according to the manufacturer's protocol. The TCD rearrangements were detected using a customized primer mix (Invivoscribe) containing 10 pmol of HEX Dδ3-3′ and Vδ1, Vδ2 Dδ2-5′ primers, Amplitaq GOLD (Thermo Fisher Scientific), 2 mM MgCl₂, dNTP (200 μM each), and Amplitaq GOLD Buffer II (Thermo Fisher Scientific). DNA input was 100 ng and PCR conditions were as follows: 7 min at 95°C and 40 cycles 45 s at 95°C, 45 s at 60°C, 35 s at 72°C. Fragments were separated on a 3730 DNA analyzer (Applied Biosystems), analyzed using the R package Fragman (version 1.0.9), and plotted using R version 3.4.4.

Themeli M., Chhatta A., Boersma H., Prins H.J., Cordes M., de Wilt E., Farahani A.S., Vandekerckhove B., van der Burg M., Hoeben R.C., Staal F.J, & Mikkers H.M. (2020). iPSC-Based Modeling of RAG2 Severe Combined Immunodeficiency Reveals Multiple T Cell Developmental Arrests. Stem Cell Reports, 14(2), 300-311.

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BRAF Exon 15 Sanger Sequencing Protocol

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Sanger sequencing was performed on tumor samples using 50 ng of genomic DNA with 0.83 µM of each primer, 0.6 U Ampli-Taq GOLD (Thermo Fisher Scientific, Waltham, Massachusetts, USA), 10 × Buffer II (Thermo Fisher Scientific) an 0.2 mM each dNTP. Cycling conditions were denaturation an 95°C for 10 min, followed by 40 cycles of 30 s 94°C, 30 s of 60°C and 30 s of 72°C, with final elongation at 72°C for 10 min.
Sanger sequencing was performed using the BigDye Terminator v1.1 Cycle Sequencing Kit (Thermo Fisher Scientific) and analyzed using Sequencing Analysis Software (Applied Biosystems). Primers used to amplify BRAF exon 15 were 5’-CATAATGCTTGCTCTGATAGGAAAATG-3’ (forward) and 5’-CATCCACAAAATGGATCCAGACA-3’ (reverse).

Reitsam N.G., Märkl B., Dintner S., Waidhauser J., Vlasenko D, & Grosser B. (2022). Concurrent loss of MLH1, PMS2 and MSH6 immunoexpression in digestive system cancers indicating a widespread dysregulation in DNA repair processes. Frontiers in Oncology, 12, 1019798.

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Rotavirus VP7 Genotyping Protocol

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The first-round PCR mix was prepared with 10X buffer II (Invitrogen) 4.5 μl, 50 mM MgCl2 2.0 μl, dNTPs (10 mM) 1.0 μl, Taq polymerase (5 U/μl) (Invitrogen) 0.2 μl, Primer VP7-F (20 pmol/μl) 1.0 μl, Primer VP7-R (20 pmol/μl) 1.0 μl, RNase-free H2O 35.3 μl, to make total volume 45.0 μl. 45 μl of PCR mix was added to each PCR tube, and 5 μl of cDNA (from the RT reaction) was added, and tubes transferred to thermal cycler and cycle conditions as provided. The samples were then separated in 2% agarose gel electrophoresis to see positive samples.
The second-round PCR mix was then prepared with 10X buffer II 4.8 μl, 50 mM MgCl2 2.5 ul, dNTPs (10 mM) 1.0 ul, Taq polymerase (5 U/μl) 0.2 ul, Primer VP7-R (20 pmol/μl) 1.0 μl, and 1.0 μl (20 pmol/μl) of each strain specific primers G1 to G12, RNase-free H₂O 30.5 µl to make total volume of 48.0 μl. 48 μl of second-round mix was then added to each PCR tube and then 2 μl of first round product was added, and PCR was run under the following temperature cycles conditions.

Mchaile D.N., Philemon R.N., Kabika S., Albogast E., Morijo K.J., Kifaro E, & Mmbaga B.T. (2017). Prevalence and genotypes of Rotavirus among children under 5 years presenting with diarrhoea in Moshi, Tanzania: a hospital based cross sectional study. BMC Research Notes, 10, 542.

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cDNA Synthesis from Extracted RNA

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About 40 μl of extracted nucleic acid were transferred to a PCR tube. The dsRNA was denatured at 97 °C for 5 min, and then chilled on ice for 2 min. The 10X buffer II (Invitrogen) 7.0 μl, 50 mM MgCl₂ 7.0 μl, Random primers 1.0 μl, dNTPs (10 mM) 2.0 μl, M-MLV (200 U/μl) Invitrogen 2.0 μl, RNase-free H2O 11.0 μl, to make total volume of 30.0 μl. Then 30 μl of RT mix was added to each tube containing the extracted RNA. Incubated at 42 °C for 50 min, and then was incubate again at 95 °C for 5 min followed by chilling on ice for 2 min. The total volume was 70 μl of cDNA was obtained and stored at − 20 °C ready for PCR.

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