Total RNA was extracted from various plant tissues using NucleoZOL Reagent Kit according to the manufacturer’s protocol (MACHEREY-NAGEL GmbH & Co. KG). The genomic DNA removal and the first stand cDNA synthesis were carried out using the ReverTra Ace qPCR RT Master Mix Kit (Toyobo, Japan). Fast Star Essential DNA Green Master Kit (Roche, Switzerland) was chosen for qRT-PCR analysis and performed on a Thermal Cycle Dice® Real Time System (TaKaRa, Japan). An ubiquitin gene (LOC_Os03g13170) was used as an internal control. Primers used for qRT-PCR are listed in Additional file 5 : Table S2. Three replicates were used for each biological sample and the means were used for analysis.
Nucleozol reagent kit
Manufactured by Macherey-Nagel
Sourced in Germany
NucleoZOL Reagent Kit is a complete solution for the isolation of high-quality RNA from a variety of sample types, including cells, tissues, and microorganisms. The kit uses a specialized reagent that facilitates the lysis of samples and the subsequent separation of RNA from other cellular components.
Automatically generated - may contain errors
Lab products found in correlation
Faststar essential dna green master kit, by Roche (4 mentions)
Thermal cycle dice real time system, by Takara Bio (3 mentions)
Revertra ace qpcr rt master mix, by Toyobo (2 mentions)
Revertra ace qpcr rt master mix kit, by Toyobo (1 mentions)
Thermal cycler dice real time system 2, by Takara Bio (1 mentions)
4 protocols using nucleozol reagent kit
1
Quantitative RNA Expression Analysis
2
RNA Extraction and qRT-PCR Analysis
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Total RNA was extracted using a NucleoZOL Reagent Kit (MACHEREY-NAGEL, Düren, Germany) according to the manufacturer’s instructions. RNA was reverse-transcribed using the ReverTra Ace qPCR RT Master Mix with genomic DNA (gDNA) Remover Kit (Toyobo, Osaka, Japan). Real-time fluorescent quantitative PCR (qRT-PCR) was carried out using the FastStar Essential DNA Green Master Kit (Roche, Basel, Switzerland) and performed on a Thermal Cycle Dice Real Time System (Takara, Kusatsu, Japan). Rice ubiquitin (LOC_Os03g13170) was used as an internal control. The primers used for qRT-PCR are listed in Table S2 . The means from three replicates were used for analysis.
3
Total RNA Extraction and qRT-PCR Analysis
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Total RNA was extracted from leaves of rcd2 and WT using NucleoZOL Reagent Kit according to the manufacturer’s instructions (Macherey-Nagel, Düren, Germany). RNA was reverse-transcribed using the ReverTra Ace qPCR RT Master Mix with genomic DNA (gDNA) Remover Kit (Toyobo, Osaka, Japan). The quantitative real-time PCR (qRT-PCR) was carried out using the FastStar Essential DNA Green Master Kit (Roche, Basel, Switzerland) and performed on a Thermal Cycler Dice Real Time System II (Takara, Kusatsu, Japan), following the method described previously.22 (link) The rice Ubiquitin was used as the internal control to normalize expression levels. The means from three replicates were used for analysis by Student’s t-test with the Excel 2016 software. All primers used in qRT-PCR are listed in Supplemental Table 1.
4
Quantifying Gene Expression in Rice Mutants
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The total RNA was isolated from flag leaves of WT and the mutants at the heading stage using the NucleoZOL Reagent Kit according to the manufacturer’s instructions (MACHEREY-NAGEL, Düren, Germany). The first strand of copy DNA (cDNA) was synthesized using the ReverTra Ace qPCR RT Master Mix with genomic DNA (gDNA) Remover Kit (Toyobo, Osaka, Japan). Real-time fluorescent quantitative PCR was carried out using the FastStar Essential DNA Green Master Kit (Roche, Basel, Switzerland) and performed on a Thermal Cycle Dice® Real Time System (Takara, Kusatsu, Japan). All target genes were normalized to the rice internal control gene Ubiquitin (LOC_Os03g13170) to detect the relative expression levels. Three biological repeats were conducted to obtain the final results. The primers for the qRT-PCR are listed in Table S1 .
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