Setting up a gene expression study for tissue cells by method of quantitative real-time PCR

Daliborka Dušanić (Author), Luka Bolha (Author), Mojca Narat (Author), Irena Oven (Author)

Abstract

Gene expression analysis by quantitative reverse transcription polymerase chain reaction (RT-qPCR) allows accurate and sensitive measurment of gene expression levels. However, a series of steps needs to be taken to ensure the relevance, accuracy, correct interpretation and repeatability of the RT-qPCR experiment. We describe here a simple experiment of determining relative gene expression for caspase 8 gene in chicken chondrocytes treated with an apoptosis inducing compound 5-fluorouracil. We use this example to point out some important guidelines in setting up a gene expression study in tissue cells, analyzing and interpreting the results and reporting on the findings.

Keywords

molecular genetics;RT-qPCR;gene expression;tissue cells;

Data

Language:	English
Year of publishing:	2012
Typology:	1.01 - Original Scientific Article
Organization:	UL BF - Biotechnical Faculty
Publisher:	Biotehniška fakulteta
UDC:	575
COBISS:	3081864
ISSN:	1581-9175
Parent publication:	Acta agriculturae Slovenica
Views:	1149
Downloads:	378
Average score:	0 (0 votes)
Metadata:

Other data

Secondary language:	Slovenian
Secondary abstract:	Analiza izražanja genov z metodo reverznega pomnoževanja in kvantitativne verižne reakcije s polimerazo v realnem času (RT-qPCR) omogoča natančno in občutljivo določanje nivoja izražanja tarčnih genov. Vendar je za zagotavljanje točnosti, natančne interpretacije in ponovljivosti eksperimenta, predvsem pa relevantnosti rezultatov, pomembno natančno poznavanje korakov postopka. V tem prispevku opisujemo enostaven poskus, v katerem smo v celični liniji kokošjih hondrocitov, ki smo jih tretirali s spojino 5-fluorouracil, ki sproži apoptozo, določili relativno izražanje gena, ki kodira encim kaspazo 8.Ta primer smo uporabili za izpostavitev nekaterih pomembnih smernic pri oblikovanju študije izražanja genov v tkivnih celicah, analizi in interpretaciji rezultatov ter oblikovanju zaključkov.
Secondary keywords:	molekularna genetika;kvantitativni PCR v realnem času;genetsko izražanje;tkivne celice;
URN:	URN:NBN:SI
Type (COBISS):	Not categorized
Pages:	str. 19-28
Volume:	ǂLetn. ǂ100
Issue:	ǂšt. ǂ1
Chronology:	2012
Keywords (UDC):	mathematics;natural sciences;naravoslovne vede;matematika;biological sciences in general;biologija;general genetics;general cytogenetics;splošna genetika;splošna citogenetika;
DOI:	10.2478/v10014-012-0009-0
ID:	1445737