Sugar beet is considered as an essential crop to produce sugar, and its common name in plant breeding is Beta vulgaris L. Production of sugar beet is regularly limited by environmental circumstances which result in reduction of photosynthesis rates, sucrose accumulation and effects root development. Therefore research is needed to understand plant response to drought stress at the genomic level, to improve sugar beet crop drought tolerance. Various research efforts aimed to identify water deficit inducible genes by determining gene expression under a stresses abiotic. Our study focused on using qRT-PCR for studying gene expression in sugar beet under in vitro drought stress conditions. Treated plants were grown for 30 days on micro-propagation media with 0%, 3%, 5% and 7% PEG. Followed by total RNA extraction from leaves which reverse transcribed into cDNA, that is used as matrix in qPCR reaction using SYBR Green. The glutamine synthetase housekeeping gene, was employed as endogenous control, while alpha amylase and osmotin-like protein were used as target genes. The relative expression quantification values for our genes of interest were measured by the 2^−ΔΔCTmethod. Alpha amylase and osmotin-like protein genes under drought stress showed a significant up-regulation expression. Additionally, qRT-PCR protocol provides an accurate and efficient result in studying the potential of expression analysis for the candidate genes under water stress in sugar beet. Our study could help in identifying plant response to abiotic stress at the gene expression level.