Evaluation of Candidate Reference Genes for Normalization of RT-PCR Data with HeLa Cells

Автор: N.A. Krainova, N.A. Khaustova, D.S. Makeeva, N.N. Fedotov, E.A. Gudim, E.A. Riabenko, M.Yu. Shkurnikov, V.V. Galatenko, D.A. Sakharov, and D.V. Maltseva

Страница: 42-50

Evaluation of Candidate Reference Genes for Normalization of RT-PCR Data with HeLa Cells

Biotekhnologiya, 2013, N 1, P. 42-50

UDC 57.04, 57.017.3, 577.1

Section:  “Producers, Biology, Selection, and Gene Engineering”

N.A. Krainova ¹,  N.A. Khaustova ¹,  D.S. Makeeva ^{1, 2},  N.N. Fedotov ³,  E.A. Gudim ⁴,  E.A. Riabenko ¹,  M.Yu. Shkurnikov ⁴,  V.V. Galatenko ³,  D.A. Sakharov ¹,  and  D.V. Maltseva ^{1, *}

¹  The Research-and-Technique Center BioClinicum, 115088, Moscow Russia

²  The Lomonosov Moscow State University, Faculty of Bioinformatics and Bioengineering,  119991, Moscow Russia

³  The Lomonosov Moscow State University, Faculty of Mechanics and Mathematics,  119991, Moscow Russia

⁴  The Research Institute for Pathology and Pathophysiology, Russ. Acad. Med. Sci.,  125315, Moscow Russia

e-mail:  d.maltseva@bioclinicum.com

Stability in expression under all studied conditions is a main criterion for the selection of reference gene for normalizing of RT-PCR results. In this work, we have used the basic methods for the assessment of the stability of candidate reference genes, ΔCt, geNorm, qBase and the Haller’s criterion of equivalence, to evaluate the applicability of reference genes (HPRT1, ACTB, GAPDH, RPS18, HSPC3, UBC, and SDHA) to RT-PCR to the most widely spread cell model, the HeLa line. The experiments were performed under standard conditions, heat shock and relaxation. Based on the summarized results obtained by all the methods, the RPS18 and HSPC3 genes that displayed the greatest expression stability within the frames of the used model were selected as reference.

Key words:  expression stability,  heat shock,  HeLa cell line,  HSPA1A,  real-time PCR,  reference genes.

The full English version of the article was published in “Applied Biochemistry and Microbiology”, 2013, Vol. 49, Issue 9, pp. 743-749 as N. A. Krainova, N. A. Khaustova, D. S. Makeeva, N. N. Fedotov, E. A. Gudim, E. A. Ryabenko, M. U. Shkurnikov, V. V. Galatenko, D. A. Sakharov, and D. V. Maltseva “Evaluation of potential reference genes for qRT-PCR data normalization in HeLa cells”.

It is contained at the SpringerLink website:  http://link.springer.com/article/10.1134/S0003683813090032  and  at the Russian Scientific Electron Library website:  http://elibrary.ru/item.asp?id=21888579

DOI: 10.1134/S0003683813090032

27.04.2015, 1602 просмотра.