Total RNA was extracted by TRIzol reagent (Invitrogen), and HiScript III RT SuperMix for RT-qPCR (Vazyme) was used for reverse transcription according to the manufacturer’s instructions. The primers used for RT-qPCR are shown as follows: p27Kip1 (forward): 5’-GGCTAACTCTGAGGACACGCA-3’; p27Kip1 (reverse): 5’-TGGGGAACCGTCTGAAACAT-3’; GAPDH (forward): 5’-GGAGCGAGATCCCTCCAAAAT-3’; and GAPDH (reverse): 5’-GGCTGTTGTCATACTTCTCATGG-3’.
Hiscript 3 rt supermix for rt qpcr
Manufactured by Vazyme
Sourced in China
HiScript III RT SuperMix for RT-qPCR is a reverse transcription reagent kit designed for cDNA synthesis and real-time quantitative PCR (RT-qPCR) applications. It contains a reverse transcriptase enzyme, RNase inhibitor, and other necessary components for efficient and sensitive RNA-to-cDNA conversion.
Automatically generated - may contain errors
Lab products found in correlation
Trizol reagent, by Thermo Fisher Scientific (1 mentions)
Cfx manager 2, by Bio-Rad (1 mentions)
Total rna extraction reagent, by Vazyme (1 mentions)
Rnaiso plus, by Takara Bio (1 mentions)
Phusion high fidelity dna polymerase, by New England Biolabs (1 mentions)
Gel extraction kit, by CWBIO (1 mentions)
Rnaiso mate for plant tissue, by Takara Bio (1 mentions)
Cfx96 machine, by Bio-Rad (1 mentions)
Trizol, by Merck Group (1 mentions)
Sybr green mix, by Vazyme (1 mentions)
4 protocols using hiscript 3 rt supermix for rt qpcr
1
Quantitative RT-PCR Analysis of p27Kip1
2
RNA Extraction and RT-qPCR Analysis
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Total RNA was extracted using the RNA isolator Total RNA extraction reagent (TRIzol, Vazyme, Nanjing, China). cDNA was extracted using HiScript®III RT SuperMix for RT-qPCR (Vazyme) following the manufacturer's protocol. ChamQ Universal SYBR RT-qPCR Master Mix RT-qPCR (Vazyme) was utilized, with β-actin serving as the internal reference. The analysis of the results was performed using Bio-Rad CFX Manager 2.1. Additional file 1 provides information about the primers used.
3
Isolation and Cloning of Plant Transcripts
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Total RNA was isolated from leaves of the Shuchazao cultivar using RNAiso‐mate for Plant Tissue (Takara, Dalian, China) and RNAiso Plus (Takara) according to the manufacturer’s instructions. cDNA was synthesized from total RNA by reverse transcription using HiScript III RT SuperMix for RT-qPCR (Vazyme Biotech Co. Ltd, Nanjing, China). Open-reading frame sequences were amplified using Phusion High-Fidelity DNA Polymerase (New England Biolabs, Ipswich, MA). The PCR products were purified with a Gel Extraction Kit (CWBIO, Jiangsu, China), ligated into the pGEX-4T1 vector, and subsequently transformed into Trans1T1 competent cells.
4
RNA Extraction and RT-qPCR Analysis
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Total RNA was isolated by the TRIzol (Sigma) following the manufacturer's instructions. Then, the RNA was reverse transcribed into cDNA by the HiScript® III RT SuperMix for RT‐qPCR (Vazyme). The RT‐qPCR was performed on a CFX96 machine (Bio‐Rad) with the SYBR Green Mix (Vazyme). PCR cycle conditions were at 95°C for 10 min for denaturation, 40 cycles of 95°C for 10 s, 60°C for 10 s and 72°C for 20s. All experiments were done in triplicate. Primers used in this study were listed in Supporting Information Table S1 .
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