Quantitative RT-PCR was performed as described (Schulz et al. 2015 (link)). In brief, total RNA was extracted from leaf material using Trizol reagent (Invitrogen, Nidderau, Germany) and DNase treated (Ambion, Austin, TX, USA). First-strand cDNA was synthesized from 2.5 µg quality-proved total RNA using Superscript III reverse-transcriptase (Invitrogen). qRT-PCR was performed with an ABI PRISM 7900 HT 384-well plate Sequence Detection System (Applied Biosystems, Darmstadt, Germany). Reactions containing 2.5 µl 2 × SYBR Green Master Mix (Fast Power SYBR Green; Applied Biosystems), 0.5 µl cDNA (diluted 6-fold) and 2 µl of 0.5 µM primers were pipetted with an Evolution P3 pipetting robot (PerkinElmer, Zaventem, Belgium). The sequences of all primers, including those for quality checks, are given in Supplementary Table S8 . Ct values for the genes of interest were normalized by subtracting the mean of four reference genes and averages of three biological replicates were calculated per accession and condition (Supplementary Table S2 ).
Evolution p3 pipetting robot
Manufactured by PerkinElmer
Sourced in Belgium
The Evolution P3 is a pipetting robot designed for laboratory automation. It features high-precision liquid handling and can perform a range of pipetting tasks with speed and accuracy. The core function of the Evolution P3 is to automate repetitive pipetting workflows, improving efficiency and reproducibility in laboratory settings.
Automatically generated - may contain errors
Lab products found in correlation
Abi prism 7900 ht 384 well plate sequence detection system, by Thermo Fisher Scientific (2 mentions)
Sybr green master mix fast power sybr green, by Thermo Fisher Scientific (2 mentions)
Trizol reagent, by Thermo Fisher Scientific (2 mentions)
Dnase treated, by Thermo Fisher Scientific (2 mentions)
Superscript 3 reverse transcriptase, by Thermo Fisher Scientific (1 mentions)
2 protocols using evolution p3 pipetting robot
1
Quantitative RT-PCR Analysis of Gene Expression
2
Quantitative PCR Analysis of Arabidopsis Transcripts
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Total RNA was extracted from rosette material pooled from five plants in each of three replicate experiments using either Trizol reagent (Invitrogen) or a lysis buffer containing 100 mM Tris-HCl (pH 8.5–9.0), 25 mM EDTA, 25 mM EGTA, 2% SDS, 100 mM ß-mercaptoethanol supplemented with phenol, chloroform and isoamylalcohol. RNA samples were DNase treated (Ambion, Austin, TX, USA, or ThermoFisher Scientific). RNA quality controls, first strand cDNA synthesis and cDNA quality controls were performed as described recently23 (link). Quantitative PCR was performed with an ABI PRISM 7900 HT 384-well plate Sequence Detection System (Applied Biosystems, Darmstadt, Germany). Reactions containing 2.5 μl 2 × SYBR Green Master Mix (Fast Power SYBR Green; Applied Biosystems), 0.5 μl cDNA (diluted 5-fold) and 2 μl of 0.5 μM primers were pipetted using an Evolution P3 pipetting robot (PerkinElmer, Zaventem, Belgium). The sequences of all primers have been published23 (link). Ct values for the genes of interest were normalized by subtracting the mean Ct of four reference genes23 (link). All normalized expression values can be found in Suppl. Table 2 .
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