Naslov
Comparison of three RT-PCR-based methods for relative quantification of mRNA.

Autori
Breljak, Davorka ; Ambriović-Ristov, Andreja ; Kapitanović, Sanja ; Čačev, Tamara ; Gabrilovac, Jelka

Izvornik
Food technology and biotechnology (1330-9862) 43 (2005), 4; 379-388

Vrsta, podvrsta i kategorija rada
Radovi u časopisima, članak, znanstveni

Ključne riječi
RT-PCR; mRNA quantification; aminopeptidase N; APN; HL-60

Sažetak
Comparison of three RT-PCR based methods: semi-quantitative, competitive and real-time RT-PCR for relative quantification of mRNA is presented. Aminopeptidase N expressed on human promyeloid HL-60 cell line, at basal and activated state, served as a model for comparison. HL-60 cells were stimulated with IFN-gamma (6 ng/mL) for 72 h at 37 oC, total cellular RNA was isolated, reverse transcribed to cDNA and semi-quantitative, competitive and real-time RT-PCR were performed to obtain the relative levels of mRNA for aminopeptidase N. The data obtained showed that all three RT-PCR based methods give reliable and comparable results, i.e. approximately two-fold increase of aminopeptidase N mRNA on IFN-gamma stimulated HL-60 cells. Thus, in spite of rapid advances made in the area of real-time RT-PCR, end-point RT-PCR such as competitive and semi-quantitative RT-PCR, although laborious and time consuming, may still remain useful techniques for relative mRNA quantification when small number of samples are to be analyzed.

Izvorni jezik
Engleski

Znanstvena područja
Biologija, Temeljne medicinske znanosti

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