Internal standards for quantitative RT-PCR studies of gene expression under drought treatment in barley (Hordeum vulgare L.): the effects of developmental stage and leaf age

Marcin Rapacz , Agata Stępień , Katarzyna Skorupa


Drought is the most significant abiotic stress in agriculture; thus, this area of studies seems to be one of the most important challenges in plant biology. Data about gene expression under drought are crucial to study drought response mechanisms and to select the genes for a transgenic approach. Quantitative RT-PCR is a powerful method for gene expression analysis; however, obtaining proper data normalization requires internal reference genes with stable level of expression. In the present paper ten potential reference genes were examined in two developmental stages of barley for their expression stability during leaf growth and increasing drought level. The results indicated that leaf growth per se affects the expression of studied genes to the similar extent as the drought and showed that different genes were most stably expressed in the seedling and the heading stage. As a result, different sets of reference genes were selected for different applications. For instance, ADP-ribozylation factor 1 and ubiquitin encoding genes were most suitable to study drought-induced changes in gene expression at the seedling stage, whereas actin and GAPDH genes were useful during heading, and ADP-ribozylation factor 1 and HSP90 allowed for the comparison between these two stages. Our data proved the necessity for validation of commonly used reference genes. The results indicate that expression of ADP seems to be the least affected by all the factors studied in the present experiment. However, when the effect of only one factor among those investigated in this work will be studied, different genes should be considered to be used as the references due to the higher stability of their expression
Author Marcin Rapacz (FoAE / DoPP)
Marcin Rapacz,,
- Department of Plant Physiology
, Agata Stępień
Agata Stępień,,
, Katarzyna Skorupa
Katarzyna Skorupa,,
Journal seriesActa Physiologiae Plantarum, ISSN 0137-5881, [1861-1664], (A 25 pkt)
Issue year2012
Publication size in sheets0.5
Keywords in EnglishBarley Developmental stage Leaf age Drought Real-time PCR Reference gene Abb
ASJC Classification1110 Plant Science; 1102 Agronomy and Crop Science; 1314 Physiology
Languageen angielski
Score (nominal)25
Publication indicators WoS Citations = 34; Scopus SNIP (Source Normalised Impact per Paper): 2012 = 0.999; WoS Impact Factor: 2012 = 1.305 (2) - 2012=1.53 (5)
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