Quantitative Gene Expression Analysis by qRT-PCR

Targeted gene expression analysis was performed with three or more biological replicates (n ≥ 3) for each genotype by qRT-PCR. Total RNA was isolated using the Trizol method (Sigma-Aldrich). DNase I (Sigma-Aldrich)-treated RNA was reverse transcribed using a high-capacity cDNA reverse transcription kit (Applied Biosystems). Gene-specific oligonucleotides were designed with Primer BLAST software (NCBI). The TIP41 gene was used as an endogenous control in gene expression analysis^{47 (link)}.

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Sonawane P.D., Gharat S.A., Jozwiak A., Barbole R., Heinicke S., Almekias-Siegl E., Meir S., Rogachev I., Connor S.E., Giri A.P, & Aharoni A. (2023). A BAHD-type acyltransferase concludes the biosynthetic pathway of non-bitter glycoalkaloids in ripe tomato fruit. Nature Communications, 14, 4540.

Publication 2023

Trizol method DNase I high-capacity cDNA reverse transcription kit

Biological Cdna Dnase i Gene Gene expression Gene expression analysis Genotype Oligonucleotides Primer Reverse transcription Trizol

Corresponding Organization : Max Planck Institute for Chemical Ecology

Other organizations : Weizmann Institute of Science

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Variable analysis

independent variables

Genotype

dependent variables

Targeted gene expression

control variables

Number of biological replicates (n ≥ 3)
RNA isolation method (Trizol)
DNase I treatment
Reverse transcription kit (Applied Biosystems)
Primer design (Primer BLAST, NCBI)
Endogenous control gene (TIP41)

controls

TIP41 gene used as endogenous control

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