Normalization of qRT-PCR data: the necessity of adopting a systematic, experimental conditions-specific, validation of references

Normalization of qRT-PCR data : the necessity of adopting a systematic, experimental conditions-specific, validation of references

Guenin, Stephanie (author)

Mauriat, Melanie (author): Swedish University of Agricultural Sciences,Sveriges lantbruksuniversitet,Institutionen för skoglig genetik och växtfysiologi,Department of Forest Genetics and Plant Physiology

Pelloux, Jerome (author)

Van Wuytswinkel, Olivier (author)

Bellini, Catherine (author): Swedish University of Agricultural Sciences,Sveriges lantbruksuniversitet,Institutionen för skoglig genetik och växtfysiologi,Department of Forest Genetics and Plant Physiology,National Institute of Agricultural Research (INRA)

Gutierrez, Laurent (author)

(creator_code:org_t)

2009-01-06
2009
English.
In: Journal of Experimental Botany. - : Oxford University Press (OUP). - 0022-0957 .- 1460-2431. ; 60:2, s. 487-493

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Abstract Subject headings

Quantitative RT-PCR (reverse transcription polymerase chain reaction, also known as qRT-PCR or real-time RT-PCR) has been used in large proportions of transcriptome analyses published to date. The accuracy of the results obtained by this method strongly depends on accurate transcript normalization using stably expressed genes, known as references. Statistical algorithms have been developed recently to help validate reference genes but, surprisingly, this robust approach is under-utilized in plants. Instead, putative ’housekeeping’ genes tend to be used as references without any proper validation. The concept of normalization in transcript quantification is introduced here and the factors affecting its reliability in qRT-PCR are discussed in an attempt to convince molecular biologists, and non-specialists, that systematic validation of reference genes is essential for producing accurate, reliable data in qRT-PCR analyses, and thus should be an integral component of them.

By the author/editor: Guenin, Stephani ...; Mauriat, Melanie; Pelloux, Jerome; Van Wuytswinkel, ...; Bellini, Catheri ...; Gutierrez, Laure ...

About the subject

AGRICULTURAL SCIENCES: AGRICULTURAL SCI ...; and Agriculture Fore ...; and Forest Science

By the university: Umeå University; Swedish University of Agricultural Sciences

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