Identification and evaluation of suitable reference genes for gene expression studies in the whitefly Bemisia tabaci (Asia I) by reverse transcription quantitative realtime PCR.
J Insect Sci
; 14: 63, 2014 May 02.
Article
en En
| MEDLINE
| ID: mdl-25373210
ABSTRACT
This study presents a reliable method for performing reverse transcription quantitative realtime PCR (RT-qPCR) to measure gene expression in the whitefly Bemisia tabaci (Asia I) (Gennadius) (Hemiptera Aleyrodidae), utilising suitable reference genes for data normalisation. We identified orthologs of commonly used reference genes (actin (ACT), cyclophilin 1 (CYP1), elongation factor 1α (EF1A), glyceraldehyde 3-phosphate dehydrogenase (GAPDH), ribosomal protein L13a (RPL13A), and α-tubulin (TUB1A)), measured the levels of their transcripts by RT-qPCR during development and in response to thermal stress, and evaluated their suitability as endogenous controls using geNorm, BestKeeper, and NormFinder programs. Overall, TUB1A, RPL13A, and CYP1 were the most stable reference genes during B. tabaci development, and TUB1A, GAPDH, and RPL13A were the most stable reference genes in the context of thermal stress. An analysis of the effects of reference gene choice on the transcript profile of a developmentally-regulated gene encoding vitellogenin demonstrated the importance of selecting the correct endogenous controls for RT-qPCR studies. We propose the use of TUB1A, RPL13A, and CYP1 as endogenous controls for transcript profiling studies of B. tabaci development, whereas the combination of TUB1A, GAPDH, and RPL13A should be employed for studies into thermal stress. The data pre- sented here will assist future transcript profiling studies in whiteflies.
Texto completo:
1
Colección:
01-internacional
Banco de datos:
MEDLINE
Asunto principal:
Regulación de la Expresión Génica
/
Proteínas de Insectos
/
Reacción en Cadena de la Polimerasa de Transcriptasa Inversa
/
Reacción en Cadena en Tiempo Real de la Polimerasa
/
Hemípteros
Tipo de estudio:
Diagnostic_studies
/
Prognostic_studies
Límite:
Animals
Idioma:
En
Revista:
J Insect Sci
Asunto de la revista:
BIOLOGIA
Año:
2014
Tipo del documento:
Article
País de afiliación:
Reino Unido