آنالیز Rt-Qpcr برخی از اعضای خانواده ژنی Devil تحت تنش شوری در گیاه آلوروپوس لیتورالیس (.Aeluropus Littoralis (Gouan) Parl)

Devil‌ها پپتیدهای کوچکی هستند که نقش مهمی در رشد و نمو گیاهان داشته و در کنترل پاسخ‌های گیاه نسبت به تغییرات محیطی حاصل از تنش‌ها موثر می‌باشند. در این تحقیق به بررسی تغییرات بیان چهار ژن aldvl1، aldvl2، aldvl3 و aldvl6 به همراه ژن‌های مرجع اختصاصی بافت برگ شامل algtf و alu2snrnp و ژن‌های مرجع اختصاصی برای بافت ریشه شامل alprs3 و alef1a در گیاه شورزی آلوروپوس لیتورالیس پرداخته شد. گیاهان تحت تنش شوری 600 میلی‌مولار کلریدسدیم و در پنج دوره زمانی صفر (کنترل)، 3، 6، 48 ساعت و یک هفته قرار گرفتند. نتایج تحقیق نشان داد که الگوی بیانی ژن‌های aldvl در اندام ریشه و برگ به شکل معنی‌داری تحت‌تاثیر تنش شوری قرار گرفته که این افزایش بیان در تمامی ژن‌ها به جز ژن aldvl6، در اندام ریشه بیشتر از برگ گیاه بود. در بین ژن‌های مورد مطالعه، بیشترین افزایش بیان برای ژن aldvl2 در بافت برگ در زمان 48 ساعت (12.12) و در اندام ریشه در زمان یک هفته (14.83) مشاهده شد. همچنین، الگوی رفتاری ژن‌ها در بافت‌های ریشه و برگ در حالت افزایش و یا کاهش بیان نسبت به یکدیگر تفاوت داشته و وابسته به بافت بود. بررسی همبستگی بیان ژن‌ها بیانگر تغییرات الگوی رفتاری بیان ژن‌‌ها در بافت‌های مختلف بود به‌طوری‌که همبستگی بیان ژن aldvl6 و aldvl3 در بافت برگ و ریشه به‌صورت منفی و معنی‌دار (0.6-) بود که نشان‌دهنده ارتباط معکوس بیان ژن در این دو بافت بوده است. با توجه به ماهیت گیاهان شورزی در تحمل شوری محیط و نیز تغییر بیان ژن‌های مورد مطالعه پس از اعمال تنش شوری، می‌توان به نقش احتمالی این ژن‌ها در افزایش قدرت تحمل گیاه اشاره نمود.

RT-qPCR analysis of some members of DEVIL gene family in Aeluropus littoralis (Gouan) Parl. under salinity stress

IntroductionAbiotic stress has always affected the plants growth and development through morphological and physiological, biochemical and anatomical alterations, leading to reduce yield and limit the crop area. Development is a completed process that depends on a cell to cell communication to regulate growth and differentiation. In the past, phytohormones were considered the important players of cell signaling. Highthroughput techniques have led to finding many new plant peptides over the last years. DEVILS are small peptides that play an important role in plant growth and development and are effective in controlling plant responses to environmental changes caused by stress. These small peptides act in the growth process of plants similar to phytohormones and are called peptide hormones that play a role in plants as molecular messengers. Aeluropus littoralis, belongs to Poaceae that can tolerate NaCl up to 600 mM and grows in marshes, salty and drought lands. Because of C4 photosynthesis system and proper physiological properties, A. littoralis as a halophyte plant could be known as a great source of genes related to salt tolerance. Materials and methods In this research, the changes of expression of four genes namaly AlDVL1, AlDVL2, AlDVL3, and AlDVL6 were investigated under salinity stress (600 mM) in Aeluropus littoralis as a halophyte plant at five timepoints of 0, 3, 6, 48 and 168 hours post stress (hps). Leaves and roots were sampled and grounded in liquid nitrogen, and total RNA was extracted using Threezol reagent (Riragene). First strand cDNA synthesis and RTqPCR were performed according to kit instructions. PCR amplification was performed in C1000™ Thermal Cycler (BioRad, USA) according to the company’s suggestions. The 2ΔΔct method was applied for the relative expression analysis. Results and discussionThe results showed that the expression pattern of AlDVL genes was affected by salinity stress and had a significant change in root and leaf tissues. In all of genes except AlDVL6, expression changes in the leaf were higher than the root. The highest value of expression in the leaf and root tissue was observed at 48 hps (12.12) and 168 hps (14.83) for AlDVL2, respectively. Expression pattern of AlDVL genes was tissuespecific, and different pattern was observed in root and leaf tissues, while in each tissue, all genes had similar pattern. For example, in all of the studied genes in leaf tissue, the amount of expression of genes was first reduced in the initial timepoint, and then the expression increased to 48 hours after the stress and eventually decreased over 168 hps. This trend of change was observed among different genes with a slight difference in leaf tissue. In the root tissue, the trend of gene expression was similar except for the AlDVL3 gene. From the first hours to 48 hps, gene expression value was declined, but was peaked at 168 hps. According to the results, it can be stated that the genes act specifically in each tissue. The results of correlation analysis in different tissues showed that AlDVL6 and AlDVL3 expression correlation in both leaf and root tissues had a negative and significant relation (0.6) with inverse effects. ConclusionsHalophyte plants by regulating their gene expression can respond to different various abiotic stresses. Expression pattern of phytohormones related genes in leaf and root tissue can be taken as an indication of their functional relevance at different timepoints of salt stress.