Determination of transcriptional level expressions by real-time pcr of some genes thought to be effective on cold stress tolerance in the plant Mentha x piperita L.

Mentha x piperita L., one of the most important medicinal aromatic plants belonging to the Lamiacea family, is a hybrid species formed by crossing watermint and spearmint. In addition to being the active ingredient of many drugs with the secondary metabolites it contains, it is used as a spice in a wide range from the food sector to the cosmetics industry thanks to its valuable essential oils. However, anthropogenic activities and abiotic factors such as sudden changes in temperatures that have occurred in climatic conditions from the past to the present affect many plant species negatively. In order to cope with these conditions that negatively affect the reproduction, growth and development of plants, it is necessary to discover plant stress tolerance mechanisms and molecules involved in signaling pathways. M. piperita, which is known to be very important medically and economically, cannot be produced by seed and is easily affected by suddenly changing environmental conditions, and the lack of any gene expression studies on cold stress in the plant reveals the need for studies in this field. It has been determined as a result of studies that M. piperita plant successfully regenerates after the cryopreservation application, which provides long-term protection by testing different parameters, and it is not known whether cold acclimation is effective in cryopreservation for M. piperita plant. For this purpose, the expression levels of six genes thought to be associated with cold stress in M. piperita were determined by Real-Time PCR. For this, first of all, cold stress-related genes were determined in the model organism Arabidopsis thaliana, since there is no sequence data on cold stress genes in M. piperita, and then BLAST between the two species in the NCBI database was performed to determine the sequences that matched the target genes the most. Primers specific to these sequences were again designed using the NCBI database. In order to determine the correlation of related genes with cold stress in M. piperita plant at the transcriptional level, experimental materials grown in vitro were exposed to cold stress for 24, 72 and 168 hours. Total RNA was isolated from leaf and stem samples of three different experimental groups and the control group. To be used in real-time PCR, cDNA was synthesized from total RNAs with reverse transcriptase enzyme. Real-Time PCR was performed to quantitatively detect the mRNA expression levels of target genes. CT values from the results were generated by 18S rRNA normalization using the 2-ΔCT method and mRNA expression plots of six genes compared to the control group. TTEST was performed for the significance of the results, and those with p ≤ 0.05 were considered significant.