Cdna synthesis kit

Manufactured by Quanta Biosciences

Sourced in United States

The cDNA Synthesis Kit is a laboratory tool designed to convert RNA into complementary DNA (cDNA). It provides the necessary reagents and enzymes required for the reverse transcription process, which is a fundamental step in various molecular biology applications, such as gene expression analysis, RNA sequencing, and cDNA library construction.

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Lab products found in correlation

Rneasy mini kit, by Qiagen (4 mentions) Trizol reagent, by Thermo Fisher Scientific (3 mentions) Lightcycler 480, by Roche (3 mentions) Trizol, by Thermo Fisher Scientific (2 mentions) Perfecta sybr green fastmix rox, by Quanta Biosciences (2 mentions) Steponeplus real time pcr system, by Thermo Fisher Scientific (2 mentions) Sybr green supermix kit, by Bio-Rad (1 mentions) Cfx connect real time system, by Bio-Rad (1 mentions) T100 thermal cycler, by Bio-Rad (1 mentions) Sybr green 1 pcr master mix, by Roche (1 mentions) Hiseq 2000 sequencer, by Illumina (1 mentions) Sybr green master mix, by Bio-Rad (1 mentions) Dnasei kit, by Thermo Fisher Scientific (1 mentions) Cfx384 c1000 touch thermal cycler, by Bio-Rad (1 mentions) Qscript microrna cdna synthesis kit, by Quanta Biosciences (1 mentions) Lightcycler 480 2, by Roche (1 mentions) Rnaeasy mini kit, by Qiagen (1 mentions) Rneasy isolation kit, by Qiagen (1 mentions) Taqman hpsc scorecard panel, by Thermo Fisher Scientific (1 mentions) Quanta sybr green supermix, by Quanta Biosciences (1 mentions)

Close spelling variants of this product

QScript microDNA cDNA Synthesis Kit First-strand cDNA synthesis kit QScript™ cDNA SuperMix synthesis kit QScript complementary DNA (cDNA) synthesis kit Quanta cDNA synthesis kit QScript cDNA synthesis kit QScript microRNA cDNA Synthesis Kit IScripts cDNA Synthesis kit QScript miRNA cDNA Synthesis Kit QScript Flex cDNA Synthesis Kit

22 protocols using cdna synthesis kit

Analyzing STAT3 Activation and Target Gene Expression

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Post transfection, cells were treated with 10 ng/ml EGF for 1.5 hr to activate STAT3. Trizol (Invitrogen, Carlsbad, CA) was then used to extract total RNA. One microgram of total RNA was reverse-transcribed using a cDNA synthesis kit (Quanta Biosciences) using a T100 Thermal Cycler (BioRad). Real-time qPCR was performed using a SYBR Green Super Mix kit on a CFX connect Real-Time System (BioRad). Gene-specific primers for STAT3 target genes were used to assess mRNA levels normalized to GAPDH mRNA levels as internal control, and the ratio of normalized mRNA to the control conditions was determined using the comparative ∆CT method for analysis. The primers used for real-time qPCR are as follows: Bcl-xL, (F) 5ʹ-ATGCAGGTATTGGTGAGTCG-3ʹ; (R) 5ʹ-CTGCTGCATTGTTCCCATAG-3ʹ; GAPDH, (F) 5ʹ-GGA GCG AGA TCC CTC CAA AAT-3ʹ; (R) 5ʹ-GGC TGT TGT CAT ACT TCT CAT GG-3ʹ; IL-6, (F) 5ʹ-ACT CAC CTC TTC AGA ACG AAT TG-3ʹ; (R) 5ʹ-CCA TCT TTG GAA GGT TCA GGT TG-3ʹ; c-MYC, (F) 5ʹ CCGCATCCACGAAACTTTG-3ʹ; (R) 5ʹ- GGGTGTTGTAAGTTCCAGTGCAA-3ʹ; TIF1 (F) 5′-TTCGGAGAAAGGCATTAGA-3′; (R) 5′ –TCCAGGGCTTCATTCATAT-3′; IRF7 (F) 5′-ATGGGCAAGTGCAAGGTGTA-3’; (R) 5-ACCAGCTCTTGGAAGAAGACTC-3’; IL-8 (F) 5-’TCTGTTAAATCTGGCAACCC-3’; (R) 5-’TAAAGGAGAAACCAAGGCAC- 3; IL-1β (F) 5′- TTCTTCGACACATTGGATAACG-3′ and (R) 5′-TGGAGAACACCACTTGTTGCT-3′.

Njatcha C., Farooqui M., Kornberg A., Johnson D.E., Grandis J.R, & Siegfried J.M. (2018). STAT3 Cyclic Decoy Demonstrates Robust Antitumor Effects in Non-Small Cell Lung Cancer. Molecular cancer therapeutics, 17(9), 1917-1926.

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Transcriptome Analysis of Gene Expression

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RNA was extracted with the RNeasy mini kit (Qiagen) following the manufacturer’s instruction. cDNA was synthesized by reverse transcription using a cDNA synthesis kit (Quanta-Bioscience). Real-time PCR reactions were performed using SYBR Green I PCR Master Mix (Roche) and the Roche LightCycler 480. Expression was normalized to the housekeeping gene Rpo. Expression was normalized to the housekeeping gene Rpo. All the primers used are listed in Supplementary Information as Supplementary Table 2.
Paired-end RNA-sequencing was performed using 1 µg RNA and two samples per lane to achieve maximum sequencing depth. The genomics unit performed the quality control and library preparation. The libraries were sequenced using Illumina HiSeq2000 sequencer. Genes with a fold-change of 1.5 were considered to be differentially expressed.

Santanach A., Blanco E., Jiang H., Molloy K.R., Sansó M., LaCava J., Morey L, & Di Croce L. (2017). The Polycomb group protein CBX6 is an essential regulator of embryonic stem cell identity. Nature Communications, 8, 1235.

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Tomato Root RNA Extraction and qRT-PCR

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Total RNA from tomato roots was extracted using spectrum plant total RNA kit (Sigma- STRN50-1KT) according to the manufacturer’s protocol. 500 ng of total RNA was used to obtained cDNA according to the protocol of Quanta Bioscience cDNA Synthesis Kit. Quantitative real-time PCR (qRT-PCR) was performed in a volume of 10 μl using PerfeCTa SYBR Green FastMix ROX (Quanta biosciences) with 5 times diluted cDNA as template. Tomato elongation factor 1-α was used as an internal control gene. Specificity of the primers was confirmed by melting curve analysis. The generated Ct values of target genes were normalized to the Ct value of internal reference EF1-α gene. Relative expression was calculated using 2^−ΔΔCt method and expressed as fold increase with respect to control^{67 (link)}.

Bari V.K., Nassar J.A, & Aly R. (2021). CRISPR/Cas9 mediated mutagenesis of MORE AXILLARY GROWTH 1 in tomato confers resistance to root parasitic weed Phelipanche aegyptiaca. Scientific Reports, 11, 3905.

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Quantitative Real-Time PCR Analysis

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RNA was isolated from Trizol-lysed cells and reverse-transcribed using Quanta
Biosciences cDNA Synthesis kit. cDNA, primers, and SYBRGreen master mix (BioRad)
were mixed and run on a Stratagene MX3000 instrument. Ct values were normalized
using two housekeeping genes.

Ding B., Parmigiani A., Divakaruni A.S., Archer K., Murphy A.N, & Budanov A.V. (2016). Sestrin2 is induced by glucose starvation via the unfolded protein response and protects cells from non-canonical necroptotic cell death. Scientific Reports, 6, 22538.

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RNA Isolation and Quantitative PCR

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Total RNA from cell lines and mice tissues was isolated using Trizol reagent (Invitrogen, Carlsbad, CA USA). RNA was treated with DNaseI using the DNaseI Kit (Ambion, Ambion Inc., Austin, TX, USA). cDNA was synthesized from 0.5–1 μg total RNA using the Quanta Biosciences qScript ™ (Quanta Biosciences Inc., Gaithersburg, MD, USA) cDNA Synthesis Kit (95047-100) for mRNA analysis and the qScript™ microRNA cDNA Synthesis Kit (95107-100) for miRNA analysis. Quantiative PCR (qPCR) of miRNAs and mRNAs in RNA isolated from cultured cell or mice livers was performed using the CFX384, C1000 touch thermal cycler (Bio-Rad, Hercoles, CA, USA) and a SYBR Green PCR Kit: Quanta Cat. #84018 and #84071, respectively. All experiments were performed in triplicate. qRT-PCR of RNA isolated from human samples was performed using the following specific TaqMan (TAqMan, Invitrogen, CA, USA) predesigned probes: Hs00399294_g1 (H19),Hs04187499_m1 (FADD), and hsa-miR-675-002005 (miR-675).

Harari-Steinfeld R., Gefen M., Simerzin A., Zorde-Khvalevsky E., Rivkin M., Ella E., Friehmann T., Gerlic M., Zucman-Rossi J., Caruso S., Leveille M., Estall J.L., Goldenberg D.S., Giladi H., Galun E, & Bromberg Z. (2021). The lncRNA H19-Derived MicroRNA-675 Promotes Liver Necroptosis by Targeting FADD. Cancers, 13(3), 411.

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Quantifying Gene Expression Profiles

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The expressions of COX-2, MIP1β and TNFα transcripts were determined with real-time PCR performed on Light cycler 480 (Roche). Cells were cultured and treated as described above, and total RNA was isolated with an RNAeasy minikit (Qiagen) according to manufacturer’s instructions. RNA concentration was determined using a Take 3 module of Epoch micro plate reader (Biotek) at 260/280 nm. 1µg of total RNA was used for reverse transcription using cDNA synthesis kit from Quanta Biosciences, containing MgCl₂, dNTPs, recombinant RNAse inhibitor protein, qScript Reverse Transcriptase, random primers, oligo (dT) primers and stabilizers. Gene expression was assayed by quantitative real-time PCR on LightCycler® 480 II (Roche Applied Science) using LightCycler® 480 SYBR Green I Master mix, cDNA prepared as described above and COX-2, MIP1β, TNFα and GAPDH forward and reverse Primers. Following are the forward (F) and reverse (R) primers used
MIP1β–F- CCAGCCAGCTGTGGTATTR-CAGTTCAGTTCCAGGTCATACATNFα-F- CCAGGGACCTCTCTCTAATCAR-TCAGCTTGAGGGTTTGCTACCOX-2-F-CAACTCTATATTGCTGGAACATGGAR-TGGAAGCCTGTGATACTTTCTGTACTGAPDH-F-TGCACCACCAACTGCTTAGCR-GGCATGGACTGTGGTCATGAGThe ΔΔCt values for COX-2, MIP1β, and TNFα were calculated relative to the GAPDH levels and values were expressed as fold change over the control (Duah et al., 2013 (link)).

Al-Azzam N., Kondeti V., Duah E., Gombedza F., Thodeti C.K, & Paruchuri S. (2015). Modulation of mast cell proliferative and inflammatory responses by Leukotriene D4 and Stem Cell Factor signaling interactions. Journal of cellular physiology, 230(3), 595-602.

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Quantitative Real-Time PCR for Gene Expression

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Relative gene expression was determined using a two-step quantitative real-time PCR method. Total RNA was isolated with the RNeasy Isolation kit with on-column DNase I treatment (Qiagen) and reverse-transcribed using the cDNA Synthesis Kit (Quanta biosciences). Quantitative reverse transcription PCR (RT–PCR) was performed with the Quanta SYBR Green Supermix (Quanta biosciences) on the ABI Prism 7500 Real-Time PCR System (Applied Biosystems). Fold changes in gene expression were determined using the comparative C_T method (ΔΔCt) with normalization to the housekeeping genes GAPDH or B2M. The primer sequences used in the study are shown in Supplementary Table 2. The TaqMan hPSC Scorecard panel was used to assess pluripotency and trilineage differentiation potential according to manufacturer's instructions (Life Technologies). Data analysis was performed using the hPSC Scorecard software (Life Technologies).

Karakikes I., Stillitano F., Nonnenmacher M., Tzimas C., Sanoudou D., Termglinchan V., Kong C.W., Rushing S., Hansen J., Ceholski D., Kolokathis F., Kremastinos D., Katoulis A., Ren L., Cohen N., Gho J.M., Tsiapras D., Vink A., Wu J.C., Asselbergs F.W., Li R.A., Hulot J.S., Kranias E.G, & Hajjar R.J. (2015). Correction of human phospholamban R14del mutation associated with cardiomyopathy using targeted nucleases and combination therapy. Nature Communications, 6, 6955.

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Quantification of Inflammatory Mediator Expression

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Expressions of CysLT₁R, CysLT₂R, EP₁, EP₂, EP₃, EP₄, MIP-1β, TNF-α, IL-8, COX-1, COX-2, and PKG transcripts were determined with real-time PCR performed on the LightCycler 480 (Roche). Total RNA was isolated from LAD2 cells after respective treatment with an RNeasy Mini Kit (Qiagen, Hilden, Germany), and cDNA was synthesized with the cDNA synthesis kit from Quanta BioSciences (Gaithersburg, Md). Real-time PCR was performed by using primers mentioned in Table E1 in this article’s Online Repository at www.jacionline.org, the levels of respective genes relative to GAPDH were analyzed, and the ΔΔ cycle threshold values relative to control were expressed as the fold change.

Kondeti V., Al-Azzam N., Duah E., Thodeti C.K., Boyce J.A, & Paruchuri S. (2015). Leukotriene D4 and prostaglandin E2 signals synergize and potentiate vascular inflammation in a mast cell–dependent manner through cysteinyl leukotriene receptor 1 and E-prostanoid receptor 3. The Journal of allergy and clinical immunology, 137(1), 289-298.

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Quantitative gene expression analysis of lung tissue

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We isolated mRNA from lung tissue samples lysed in buffer RLT (RNeasy Lysis Buffer for tissues, Qiagen) using RNeasy kits (QIAGEN). 500 ng of total RNA was used in a cDNA synthesis reaction, using a cDNA Synthesis Kit (Quanta BioSciences). The qPCR reactions utilized probe-based qPCR Master Mix (Integrated DNA Technologies [IDT]) and commercially pre-designed and validated primer/probe gene expression assays (available from IDT) for the following genes: cytokines (Interleukin [IL]-4, IL-5, IL-10, IL-13 and IL-33), chemokines (C-C Motif Chemokine Ligand [Ccl] 2, Ccl5, Cl11, and Ccl24), growth factors (Transforming growth factor beta 1 [Tgfb1], and Insulin-like growth factor 1 [Igf1]) and tissue remodeling markers (Caspase 1 [Casp1], NLR Family Pyrin Domain Containing 3 [Nlrp3], IL-1β, Fibroblast growth factor receptor [Fgfr] 1, Fgfr2, Aldolase A [Aldoa], SRY-box 2 [Sox2], Nestin [Nes], Tenascin C [Tnc], Vimentin [Vim], and Periostin [Postn]). The qPCR reactions were run using a StepOnePlus Real-Time PCR System (Applied Biosystems). Gene expression was calculated relative to the expression of GAPDH (housekeeping gene) and was reported as true copies of the gene of interest per 10⁴ copies of GAPDH as previously described [21 (link)].

Chiarella S.E., Cuervo-Pardo L., Coden M.E., Jeong B.M., Doan T.C., Connelly A.R., Rodriguez R.I., Queener A.M, & Berdnikovs S. (2023). Sex differences in a murine model of asthma are time and tissue compartment dependent. PLOS ONE, 18(10), e0271281.

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Quantification of IP3R and miR-204 in Mice

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Total RNA from cultured cells or tissues of mice was isolated by the TRIZOL (Invitrogen) method. Real-time PCR was performed using the Prism 7000 Sequence Detection System (Applied Biosystems, Foster City, CA) with the Super Script III Platinum SYBR Green One-Step qRT-PCR Kit (Invitrogen). The following primers purchased from Exiqon were used: Mouse IP₃R1 forward 5′- TGG GCA ACT GTG GGA CCT T-3′, reverse 5′- GGA ACT CCA CAT CCA GAA CCA-3′; mouse IP₃R2 forward 5′- AGG AAG GTG AGG ATG GCA TAG A-3′, reverse 5′-CAC GGT GAC AAT GCA CAT GA-3′; mouse IP₃R3 forward 5′-CAG AAC GAC CGC AGG TTT GT-3′, reverse 5′- CAG GAC ATT CTG CCC ATT GTT-3′; mouse GAPDH forward 5′- GGC AAA TTC AAC GGC ACA-3′, reverse 5′-CGC TCC TGG AAG ATG GTG AT-3′. MiR-204 forward and universal reverse primers were procured from Quanta Biosciences (cDNA synthesis kit, Quanta Biosciences, Beverly, MA, USA). Mouse GAPDH and RNU6 (Quanta Biosciences, Beverly, MA, USA) were used as internal controls for mRNA and miR quantification, respectively.

Gabani M., Liu J., Ait-Aissa K., Koval O., Kim Y.R., Castañeda D., Vikram A., Jacobs J.S., Grumbach I., Trebak M., Irani K, & Kassan M. (2019). MiR-204 regulates type 1 IP3R to control vascular smooth muscle cell contractility and blood pressure. Cell calcium, 80, 18-24.

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