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Amv reverse transcriptase

Manufactured by Takara Bio
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Sourced in Japan, China, United States

AMV reverse transcriptase is an RNA-dependent DNA polymerase enzyme that can be used to synthesize complementary DNA (cDNA) from RNA templates. It is isolated from the Avian Myeloblastosis Virus (AMV) and is commonly used in reverse transcription reactions for gene expression analysis and other molecular biology applications.

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Lab products found in correlation

Trizol reagent, by Thermo Fisher Scientific (75 mentions) Trizol, by Thermo Fisher Scientific (16 mentions) Taqman pcr kit, by Thermo Fisher Scientific (13 mentions) Rnase free dnase, by Takara Bio (12 mentions) Taqman mirna probes, by Thermo Fisher Scientific (11 mentions) Random primer, by Takara Bio (11 mentions) Stem loop rt primer, by Thermo Fisher Scientific (10 mentions) Applied biosystems 7300 sequence detection system, by Thermo Fisher Scientific (10 mentions) Taqman probe, by Thermo Fisher Scientific (10 mentions) Sybr premix ex taq 2 kit, by Takara Bio (9 mentions) Trizol, by Takara Bio (9 mentions) Rneasy mini kit, by Qiagen (8 mentions) Sybr premix ex taq, by Takara Bio (7 mentions) Cfx96 real time pcr detection system, by Bio-Rad (7 mentions) Rnase inhibitor, by Takara Bio (7 mentions) Dnase 1, by Takara Bio (6 mentions) Trizol reagent, by Takara Bio (5 mentions) Sybr green pcr kit, by Roche (5 mentions) Real time pcr system, by Bio-Rad (5 mentions) Cfx96 real time pcr system, by Bio-Rad (5 mentions)

Spelling variants of this product

Reverse transcriptase (314 citations) AMV Reverse Transcriptase XL (36 citations) AMV transcriptase (8 citations) AMV reverse transcriptase kit (6 citations) AMV reverse transcriptase and random primer (5 citations) AMV reverse transcriptase reagent box (2 citations) AMV Reverse Transcriptase XL kit (2 citations) AMV reverse transcriptase system (2 citations)

189 protocols using amv reverse transcriptase

1

Quantitative RT-PCR Analysis of Gene Expression

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Total RNA was extracted from the cells using Trizol Reagent (Invitrogen, Carlsbad, CA) without detaching the cells. The complementary DNA (cDNA) was synthesised from 2 μg total RNA per sample with AMV reverse transcriptase (Takara, Japan) in a 20‐μL reaction solution containing 4 μL 5× buffer, 2 μL dNTP, 1 μL oligo‐(dT), 0.5 μL RNase inhibitor and 0.5 μL AMV reverse transcriptase and ddH2O. The mixture was then incubated at 30°C for 10 minutes, 42°C for 60 minutes, 95°C for 5 minutes and 5°C for 5 minutes.
Then, cDNA was amplified using a Power SYBR Green PCR master mix (×2) (Applied Biosystems, Foster City, CA) in a real‐time thermal cycler (Stratagene Mx3000PTM QPCR System, La Jolla, CA), and each measurement was repeated in triplicate. The optimised primers for qPCR analysis were listed in Table S3. The human housekeeping gene Hypoxanthine phosphoribosyl transferase 1 (HPRT1) was used as an internal control. Experiments were repeated in three cell samples.
Li Q., Tu T., Wu X., Wang W., Gao Z, & Liu W. (2022). Tissue chondrification and ossification in keloids with primary report of five cases. International Wound Journal, 19(7), 1860-1869.
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2

RNA Isolation and qPCR Analysis

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According to the manufacturer’s protocol, total RNA was isolated from cells by the Trizol reagent. For mRNA detection, 500 ng of total RNA were used for complementary DNA synthesis (One Step SYBR PrimeScript™ RT-PCR Kit, Takara, Shiga, Japan), according to the manufacturer’ instructions. For miR-21 detection, 2 µg of total RNA was used for first-strand DNA synthesis using AMV reverse transcriptase (Takara) and a stem-loop RT-primer (Life Technology). Real-time q-PCR was performed by 7300 real time PCR System (Applied Bio-systems, Foster City, CA) according to the manufacturer’s protocol. mRNA level was normalized to β-actin, while miR-21 expression was normalized to small nuclear RNA U6.
Sun Q., Xiao L., Cui Z., Yang Y., Ma J., Huang Z., Zhang J, & Chen J. (2022). 3,3′-Diindolylmethane improves antitumor immune responses of PD-1 blockade via inhibiting myeloid-derived suppressor cells. Chinese Medicine, 17, 81.
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3

Relative Transcript Levels Analysis

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Total RNA was extracted from the soybean leaves under the stress-treatment and unstressed using an RNA plant extraction kit (OMEGA, China) according to the manufacture’s instruction, and approximately 1 μg of purified total RNA was reverse transcribed to cDNA in a 20 μL reaction volume using AMV reverse transcriptase (Takara) according to the supplier’s instructions. To analyze the relative transcript levels of selected genes, real-time PCR was performed with specific primers according to the Bio-Rad Real-time PCR system (Foster city, CA, USA) and the SYBR Premix Ex II system (TakaRa Perfect Real Time). The qRT-PCR program was: 95 °C for 30 s;40 cycles of 95 °C for 5 s and 60 °C for 34 s; 95 °C for 15 s. The primers used for RT-PCR are listed in Additional file 4: Table S9.In each case, three technical replicates were performed for each of at least three independent biological replicates [31 (link)]. Quantities of standard RNA were prepared by diluting cDNA (1, 10− 1, 10− 2, 10− 3, 10− 4, and 10− 5) and only Ct values less than 40 were used to calculate correlation coefficients (R2 values) and amplification efficiencies (E) from the slope generated in Microsoft Excel 2013, based on the eq. E = [10−(1/slope) − 1] × 100%. All PCR assays showed efficiency values between 95% and 110% (Additional file 4: Table S9) [32 (link)].
Zhang M., Liu Y., Shi H., Guo M., Chai M., He Q., Yan M., Cao D., Zhao L., Cai H, & Qin Y. (2018). Evolutionary and expression analyses of soybean basic Leucine zipper transcription factor family. BMC Genomics, 19, 159.
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4

Subcellular RNA Fractionation and RT-qPCR

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To determine the ratio of nuclear to cytosolic mRNA for specific genes, subcellular RNA fractionation was carried out using an RNA Subcellular Isolation Kit (Active Motif, USA) according to the manufacturer's instructions. The reverse transcription PCR and the real-time quantitative PCR (RT-qPCR) were performed according to a modified version of the method described previously (28 (link)). Briefly, cDNA was synthesized using AMV Reverse Transcriptase (Takara Bio, Japan), and RT-qPCR was performed with a FastStart Essential DNA Green Master Kit (Roche Diagnostics, USA) using a Real-time PCR LightCycler 96 (Roche Diagnostics, Switzerland). The MIQE (29 (link)) checklist is provided in Supplementary Table S2.
Bong S.M., Bae S.H., Song B., Gwak H., Yang S.W., Kim S., Nam S., Rajalingam K., Oh S.J., Kim T.W., Park S., Jang H, & Lee B.I. (2020). Regulation of mRNA export through API5 and nuclear FGF2 interaction. Nucleic Acids Research, 48(11), 6340-6352.
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5

Hypothalamus Total RNA Extraction

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Total RNA was extracted from the hypothalamus of the sacrificed rodents using TRIzol reagent (D9108A, TaKaRa, Osaka, Japan). The concentration and the purity of total RNA were examined by the A260/A280 ratio using an ultraviolet spectrophotometer (Eppendorf, Hamburg, Germany). The integrity of total RNA was also checked using agarose gel electrophoresis. According to the instructions of TaKaRa RNA PCR Kit (AMV) 3.0, all RNA samples were reverse-transcribed using the AMV reverse transcriptase (2621, TaKaRa, Osaka, Japan) and an oligodeoxythymine (oligo(dT)18) (3806, TaKaRa, Osaka, Japan). All cDNAs obtained were stored at −80 °C.
Qi Y., Xue H., Xu J., Wu M., Chen L, & Xu L. (2023). Effect of PACAP/PAC1R on Follicle Development of Djungarian Hamster (Phodopus sungorus) with the Variation of Ambient Temperatures. Biology, 12(2), 315.
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6

Extracting and Analyzing Murine Integrin Transcripts

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Genomic RNA was extracted from the tongue or lung tissues of suckling mice using an RNeasy Mini kit, as previously described [29 ]. The use of all animals in this study was approved by the Review Board of Lanzhou Veterinary Research Institute, Chinese Academy of Agricultural Sciences. cDNA was synthesized from the extracted RNA with AMV reverse transcriptase (Takara Bio, Japan) using random primers (20 pmol/mL) and used as the template for amplification of the αv and β3 transcripts with PCR. The PCR primer pairs, αvF/αvR and β3R/β3F, respectively, are shown in Table 1. The PCR reaction was performed in a volume of 40 µL. The cycling parameters were as follows: 40 cycles of denaturation at 95℃ for 3 min, annealing at 58℃ for 30 sec, and elongation at 72℃ for 4 min, followed by a final elongation step at 72℃ for 10 min before the reaction was cooled to 4℃ for further processing. The amplicons were purified with a QIAquick Gel Extraction Kit (Takara Bio) and cloned separately into the pGEM-T Easy vector (Promega, USA). The PCR products were verified by electrophoresis and sequenced in both directions by GeneWiz (China).
Zhang W., Lian K., Yang F., Yang Y., Zhu Z., Zhu Z., Cao W., Mao R., Jin Y., He J., Guo J., Liu X, & Zheng H. (2015). Establishment and evaluation of a murine αvβ3-integrin-expressing cell line with increased susceptibility to Foot-and-mouth disease virus. Journal of Veterinary Science, 16(3), 265-272.
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7

Quantification of CSFV RNA Levels

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The viral RNA in the supernatants was extracted with a viral RNA minikit (catalog no. W7091; Watson Biotechnologies). Synthesis of cDNA was performed in a 20-μl volume containing 200 ng of total RNA, 20 U of avian myeloblastosis virus (AMV) reverse transcriptase (catalog no. D2620; TaKaRa), 200 μM deoxynucleoside triphosphates (dNTPs) (catalog no. D4030A; TaKaRa), 0.4 μM random primers (catalog no. D6045; TaKaRa), 0.5 μl of RNase inhibitor (catalog no. D2313A; TaKaRa), and 4 μl of 5× AMV reverse transcriptase buffer. Quantification of CSFV genome copy numbers was performed with Premix Ex Taq (Probe qPCR) (catalog no. RR390A; TaKaRa) using a previously described real-time reverse transcription-PCR (RT-qPCR) assay (34 (link)).
Wang J., Chen S., Liao Y., Zhang E., Feng S., Yu S., Li L.F., He W.R., Li Y., Luo Y., Sun Y., Zhou M., Wang X., Munir M., Li S, & Qiu H.J. (2016). Mitogen-Activated Protein Kinase Kinase 2, a Novel E2-Interacting Protein, Promotes the Growth of Classical Swine Fever Virus via Attenuation of the JAK-STAT Signaling Pathway. Journal of Virology, 90(22), 10271-10283.
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8

Molecular Mechanism of Apoptosis Induction

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4-(2-Hydroxyethyl)-1-piperazineethanesulfonic acid and trypsin were purchased from AMRESCO. Bovine serum albumin (BSA) was purchased from Sijiqing Hangzhou Bioengineering Company. Dulbecco’s Modified Eagle’s Medium (DMEM) was purchased from GIBCO BD. MTT, diethylpyrocarbonate, and ethidium bromide were purchased from Sigma. Dimethyl sulfoxide was purchased from Shanghai Ling Feng Chemical Co., Ltd. RNAiso Reagent, AMV reverse transcriptase, deoxyribonucleoside triphosphate (dNTP), Oligo(dT)18, Taq DNA polymerase, 100 bp DNA Marker, RNasin, and RNase free DNase I were purchased from Takara. From Shanghai Shenneng Gaming Biotechnology Co. Ltd. (Whitehouse Station, NJ, USA), 20×TBE, agarose, caspase-3 primer, caspase-8 primer, caspase-9 primer, and β-actin primer were purchased. C225 solution for infusion was purchased from Merck & Co, Germany. All other chemicals were commercially available and of analytical grade.
Lu Q., Dai X., Zhang P., Tan X., Zhong Y., Yao C., Song M., Song G., Zhang Z., Peng G., Guo Z., Ge Y., Zhang K, & Li Y. (2018). Fe3O4@Au composite magnetic nanoparticles modified with cetuximab for targeted magneto-photothermal therapy of glioma cells. International Journal of Nanomedicine, 13, 2491-2505.
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9

Quantifying Cytokine Expressions in RAW 264.7 Cells

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After RAW 264.7 cells were stimulated, the expressions of TNF-α, MCP-1, and IL-1β were measured by qRT-PCR as reported previously (35 (link)). Briefly, the total RNA of cells was extracted using the TRIzol® reagent (Invitrogen, Paisley, UK). Then, complementary DNA was synthesized from 4 µg of the total RNA by using AMV reverse transcriptase (Takara, Japan), as previously described (36 (link)). qRT-PCR was performed using ViiA™ 7 Software (Applied Biosystems) with SYBR green PCR Kit (Roche). All of the primers used in qRT-PCR were listed in Table 2. The relative amounts of target gene expression were normalized with GAPDH housekeeping gene, using 2−ΔΔCt method (37 (link)).
Zhang Q., Huang J., Yu J., Xu Z., Liu L., Song Y., Sun X., Zhang A, & Jin M. (2017). HP1330 Contributes to Streptococcus suis Virulence by Inducing Toll-Like Receptor 2- and ERK1/2-Dependent Pro-inflammatory Responses and Influencing In Vivo S. suis Loads. Frontiers in Immunology, 8, 869.
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10

Circadian Clock Gene Expression Analysis

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Blood samples were taken from the forearm vein at 09:00 [7 (link)]. Leucocytes were isolated immediately using lymphocyte separation media (Dakewe Biotech, Shenzhen, China). The isolation of total RNA was performed using the RNAiso Plus reagent (TaKaRa, Otsu, Japan) according to the manufacturer's instructions. 1 μg of total RNA was treated with RNase-free DNase I for 10 minutes at 37°C to remove any contaminating DNA and reverse-transcribed with AMV reverse transcriptase (TaKaRa, Otsu, Japan). Gene expression was analyzed by real-time quantitative PCR using the ABI PRISM 7000 Sequence Detection System (Applied Biosystems). The GenBank accession numbers, assay ID, and the target exons were as follows: NM_001178.4 and Hs00154147_m1, 9-10 for BMAL1; NM_002616.1 and Hs00242988_m1, 22-23 for PER1; NM_022817.1 and Hs00256143_m1, 8-9 for PER2; and NM_016831.1 and Hs00213466_m1, 15-16 for PER3. Relative mRNA levels of each transcript were determined by the comparative Ct method with the reference gene (GAPDH).
Huang Y., Wang H., Li Y., Tao X, & Sun J. (2017). Poor Sleep Quality Is Associated with Dawn Phenomenon and Impaired Circadian Clock Gene Expression in Subjects with Type 2 Diabetes Mellitus. International Journal of Endocrinology, 2017, 4578973.
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