Hieff qpcr sybr green master mix low rox plus

Manufactured by Yeasen

Sourced in China, United States

Hieff® qPCR SYBR Green Master Mix (Low Rox Plus) is a ready-to-use solution for quantitative real-time PCR (qPCR) experiments. It contains SYBR Green I dye, DNA polymerase, dNTPs, and necessary buffers and additives for efficient and sensitive qPCR amplification.

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Lab products found in correlation

S1000, by Bio-Rad (7 mentions) Trizol reagent, by Thermo Fisher Scientific (6 mentions) Hifair 3 1st strand cdna synthesis kit, by Yeasen (3 mentions) Hifair 3 1st strand cdna synthesis supermix for qpcr, by Yeasen (3 mentions) Trnzol universal, by Tiangen Biotech (3 mentions) Rnaprep pure plant kit, by Tiangen Biotech (3 mentions) Hifair 1st strand cdna synthesis supermix, by Yeasen (3 mentions) Trizol, by Thermo Fisher Scientific (3 mentions) Applied biosystems quantstudio 6 flex real time pcr system, by Thermo Fisher Scientific (3 mentions) Hifair 2 1st strand cdna synthesis kit, by Yeasen (2 mentions) Trizol, by Takara Bio (2 mentions) Hifair 2 1st strand cdna synthesis supermix for qpcr gdna digester plus, by Yeasen (2 mentions) Hifair 3 1st strand cdna synthesis supermix kit, by Yeasen (2 mentions) Hifair 3 1st strand cdna synthesis supermix for qpcr gdna digester plus, by Yeasen (2 mentions) Abi prism 7500 real time pcr system, by Thermo Fisher Scientific (2 mentions) Antibodies against β actin and hrp goat anti rabbit igg h l, by Beyotime (2 mentions) Hifair 2 1st strand cdna synthesis supermix for qpcr, by Yeasen (2 mentions) Nanodrop 2000, by Thermo Fisher Scientific (2 mentions) Hiscript 2 q select rt supermix for qpcr gdna wiper, by Vazyme (1 mentions) Prism 8, by GraphPad (1 mentions)

52 protocols using hieff qpcr sybr green master mix low rox plus

Quantitative Analysis of Gene Expression using GAPDH as Control

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The glyceraldehyde-3-phosphate dehydrogenase (GAPDH) gene was used as a control gene to analyze the relative expression of specific genes. Standard cDNA synthesis techniques were followed, and RT-qPCR was performed on a Bio-Rad S1000 with Hieff qPCR SYBR^® Green Master Mix (Low Rox Plus; YEASEN, China). Primer data are available in Supplementary File 1. The concentration of each transcript was then normalized to GAPDH mRNA level using the 2^–ΔΔCT method (17 (link)).
A list of the primers that were used for the detection of pre-mRNA splicing, and subsequently for the quantitative evaluation of the two distinct splicing isoforms of a given ASE using the quantitative polymerase chain reaction (qPCR) technique, is given in Supplementary File 1. To precisely amplify each of these two isoforms, primers complementary to the splice junction of the constitutive exon and alternative exon were designed. Standard cDNA synthesis protocols were followed, and RT-qPCR was conducted using a Bio-Rad S1000 with Hieff qPCR SYBR^® Green Master Mix (Low Rox Plus; YEASEN, China). The 2^–ΔΔCT technique was used to quantify PCR amplifications.

Zhao M., Zhou J., Tang Y., Liu M., Dai Y., Xie H., Wang Z., Chen L, & Wu Y. (2023). Genome-wide analysis of RNA-binding proteins co-expression with alternative splicing events in mitral valve prolapse. Frontiers in Immunology, 14, 1078266.

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Quantifying Gene Expression in Colon and Neurons

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Total RNA from human colon tissues and enteric neurons was isolated by TRIzol reagent (Invitrogen, CA, USA) following the manufacturer’s protocol, respectively. cDNA was synthesized using the HiScript II Q Select RT SuperMix for qPCR (+gDNA wiper) (Vazyme, R233-01, China) according to the manufacturer’s instructions. The RT-qPCR was performed using the Hieff qPCR SYBR Green Master Mix (Low Rox Plus) (YEASEN, 11202ES08, China) with the Applied Biosystems, Quant Studio5 (Applied Biosystems, CA, USA). The primers used for qPCR were listed in Table S1. The qPCR data were calculated by the 2^−ΔΔCt method, normalizing to 18S rRNA gene expression.

Xu Y., Wang F., Mi K., Wang X., Wang D., Zhao Q., Wang J., Liu Z., Zhang Q., Liu Y., Zhang X, & Liu X. (2023). Biglycan regulated colorectal cancer progress by modulating enteric neuron-derived IL-10 and abundance of Bacteroides thetaiotaomicron. iScience, 26(9), 107515.

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RT-qPCR Validation of Dual RNA-seq Data

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RT-qPCR was used to validate the dual RNA-seq data. Total RNA of the coaggregated pellets and the monocultures was extracted and examined as previously described. RNA reverse transcription process was performed according to the manufacturer’s instructions (Hifair^® II 1st Strand cDNA Synthesis Kit, Yeasen Biotech, Shanghai, China). RT-qPCR was performed using Hieff^® qPCR SYBR Green Master Mix (Low Rox Plus) (Yeasen Biotech, Shanghai, China) following the manufacturer’s protocol. Relative expression was calculated by normalizing to the corresponding 16S rRNA gene transcripts. Three biological replicates were performed for all RT-qPCR reactions. PCR primers used in the present study are listed in Supplementary Materials (Table S1). Gene expression correlations between RT-qPCR results and RNA-seq results were evaluated by Pearson’s correlation coefficients using GraphPad Prism 8 (https://www.graphpad.com/scientific-software/prism/).

Liu T., Yang R., Zhou J., Lu X., Yuan Z., Wei X, & Guo L. (2022). Interactions Between Streptococcus gordonii and Fusobacterium nucleatum Altered Bacterial Transcriptional Profiling and Attenuated the Immune Responses of Macrophages. Frontiers in Cellular and Infection Microbiology, 11, 783323.

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Quantitative RNA Expression Analysis

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Total RNA was isolated using the RN001 RNA Quick Purification Kit (ESscience, China). Next, the mRNAs were reverse transcribed into cDNA using Hifair® III Reverse Transcriptase (Yeasen Biotech, China). Subsequently, Hieff® qPCR SYBR Green Master Mix (Low Rox Plus) (Yeasen Biotech, China) was used for qPCR on a QuantStudio 7 Flex System (Applied Biosystems, USA). The primer sequences used are shown in Table S1.

Hao Z., Ren L., Zhang Z., Yang Z., Wu S., Liu G., Cheng B., Wu J, & Xia J. (2022). A multifunctional neuromodulation platform utilizing Schwann cell-derived exosomes orchestrates bone microenvironment via immunomodulation, angiogenesis and osteogenesis. Bioactive Materials, 23, 206-222.

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Extracting and Quantifying Transcripts from GBM

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Total RNA was isolated from GBM tissues and cell lines using Trizol RNA isolation reagent (Invitrogen) and reversely transcribed to cDNA with a cDNA Synthesis kit (Yeasen Biotech Co., Shanghai, China). qRT-PCR was used to detect the gene expressions with Hieff^® qPCR SYBR Green Master Mix (Low Rox Plus) (Yeasen Biotech Co. China). The sequences of primers are shown in Supplementary Table S2.

Liu X., Huang Y., Qi Y., Wu S., Hu F., Wang J., Shu K., Zhang H., Bartsch J.W., Nimsky C., Dong F, & Lei T. (2022). The GBM Tumor Microenvironment as a Modulator of Therapy Response: ADAM8 Causes Tumor Infiltration of Tams through HB-EGF/EGFR-Mediated CCL2 Expression and Overcomes TMZ Chemosensitization in Glioblastoma. Cancers, 14(19), 4910.

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Quantitative Real-Time PCR Analysis of Gene Expression in Colorectal Cancer

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We obtained cancer and normal tissue samples from eight CRC patients who underwent curative resection at Tongji Hospital of Tongji Medical College (Wuhan, China). The MolPure^® Cell/Tissue Total RNA Kit (Yeasen) was used to extract total RNA, which was then reverse transcribed with the Hifair^® III 1st Strand cDNA Synthesis Kit (gDNA digester plus, Yeasen) in accordance with the manufacturer’s protocols. The target sequence was amplified with real-time PCR with the Hieff^® qPCR SYBR Green Master Mix(Low Rox Plus, Yeasen). The cycling parameters used were 95°C for 5 min, 95°C for 10 s, and 60°C for 30 s for 40 cycles. Melting curve analyses were performed, and Ct values were determined during the exponential amplification phase of real-time PCR. The 2^–ΔΔCt method was used to determine relative fold changes between tumor tissues and normal tissues as the following equation: 2 ^–ΔΔCt (ΔΔCt = ΔCt^tumor – ΔCt^normal). The primer sequences were listed in Supplementary Table 5.

Sun M., Ji X., Xie M., Chen X., Zhang B., Luo X., Feng Y., Liu D., Wang Y., Li Y., Liu B., Xia L, & Huang W. (2022). Identification of necroptosis-related subtypes, development of a novel signature, and characterization of immune infiltration in colorectal cancer. Frontiers in Immunology, 13, 999084.

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Total RNA Extraction and qRT-PCR Analysis

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Total RNA was extracted using AG RNAex Pro Reagent (Accurate Biotechnology, Hunan, China) according to the manufacturer’s instructions. Then 2 μg of total RNA was reversely transcribed into cDNA using HiScript III RT SuperMix for qPCR (+ gDNA wiper) (Vazyme Biotechnology, Nanjing, China). Quantitative real-time PCR (qRT-PCR) was performed using Hieff® qPCR SYBR Green Master Mix (Low Rox Plus) (Yeasen Biotechnology, Shanghai, China) and the Applied Biosystem™ QuantStudio™ 6 Flex Real-Time PCR System. The primers used for qRT-PCR are listed in Table S1 (Additional file 1). Three or more biological replicates (each replicate involved 20 or more individuals) were performed as previously described [6 (link), 33 (link)].

Jia Q., Yang L., Wen J., Liu S., Wen D., Luo W., Wang W., Palli S.R, & Sheng L. (2024). Cyp6g2 is the major P450 epoxidase responsible for juvenile hormone biosynthesis in Drosophila melanogaster. BMC Biology, 22, 111.

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Validating Transcriptomic Genes via qRT-PCR

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Candidate transcriptomic genes were validated with qRT-PCR analysis using three biological replicates. Total RNA was extracted as described above. cDNAs were synthesized using the HiScript II Reverse Transcriptase (Vazyme, Nanjing, China). qRT-PCR was performed on a Roche Light Cycler^® 480 Real-Time PCR System (Roche Diagnostics, Mannheim, Germany) using the Hieff qPCR SYBR Green Master Mix (Low Rox Plus) (Yeasen Biotechnology, Shanghai, China). The PCR procedure was as follows: denaturation for 5 min at 95 °C, followed by 40 cycles at 95 °C for 10 s and 60 °C for 30 s. The primers used in qRT-PCR were designed using Primer Premier v6.0 (Table S2). The reference gene Tubulin was used for normalization. The relative expression levels of the target genes were calculated using the 2^−∆∆Ct method.

Wang X., Ye Z.X., Wang Y.Z., Wang X.J., Chen J.P, & Huang H.J. (2023). Transcriptomic Analysis of Tobacco Plants in Response to Whitefly Infection. Genes, 14(8), 1640.

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qRT-PCR Analysis of Gene Expression

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Total RNA from the indicated cells that different treatments was extracted with TRIzol (TaKaRa, Otsu, Shiga, Japan, 9109) and reversed transcribed into cDNA by Hifair® Ⅱ 1st Strand cDNA Synthesis SuperMix for qPCR (Yeasen Biotech, Shanghai, China, 11123ES60). The relative qPCR was performed with Hieff® qPCR SYBR Green Master Mix (Low Rox Plus) (Yeasen Biotech, Shanghai, China, 11202ES08). Applied Biosystems 7500 was used for numerical determination, and the thermal cycling program consisted of 5 min at 95 °C followed by 45 cycles at 95 °C for 10 s, 57 °C for 20 s, and 72 °C for 34 s. All primers for qRT-PCR are listed in Supplementary Table S2.

Sun R., Guo Y., Zhang L., Zhang H., Yin B., Li X., Li C., Yang L., Zhang L., Li Z, & Huang J. (2024). PRRSV degrades MDA5 via dual autophagy receptors P62 and CCT2 to evade antiviral innate immunity. Virologica Sinica, 39(2), 264-276.

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Quantitative PCR Analysis of Insect Tissues

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Total RNA was extracted from the ovary, CA, epidermis, midgut, colleterial glands, fat body, or brain using TRIzol™ Reagent (Invitrogen, MA, USA). Total RNA (2 μg) was reverse transcribed into cDNA using a SMARTer® PCR cDNA Synthesis Kit (Takara, Dalian, China), following the manufacturer’s instructions. qPCR was performed using Hieff® qPCR SYBR Green Master Mix (Low Rox Plus) (Yeasen Biotech, Shanghai, China) and Applied Biosystems™ QuantStudio™ 6 Flex Real-Time PCR System (Thermo Fisher Scientific, MA, USA). The thermocycling conditions were as follows: 94 °C for 2 min, 40 cycles of 94°C for 10 s, and 56 °C for 30 s. Actin-5c was used as an internal reference [76 (link)]. The relative expression levels of the indicated genes were computed using the 2-ΔΔCt method [25 (link), 76 (link)]. The primer sequences used for qPCR are listed in Table S1 (Additional file 1).

Li Z., Zhou C., Chen Y., Ma W., Cheng Y., Chen J., Bai Y., Luo W., Li N., Du E, & Li S. (2022). Egfr signaling promotes juvenile hormone biosynthesis in the German cockroach. BMC Biology, 20, 278.

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