Development of EST-SSR molecular markers in rice (Oryza sativa L.) under salinity stress and identification of key genes

Document Type : Research Paper


1 Department of Biotechnology, Faculty of Agriculture, Azarbaijan Shahid Madani University, Tabriz, Iran

2 Azarbaijan Shahid Madani University

3 a staff member of the Agriculture


Expressed sequence tags of simple sequence repeats (EST-SSRs) are used to investigate genetic diversity and develop molecular markers for plants under biotic and abiotic stresses. However, there are a limited number of molecular markers based on the ESTs in rice to cope with the abiotic stresses including salinity, for use in breeding programs. Among 8299 ESTs, available in the NCBI database for salinity stress, 525 contigs, and 1139 singleton sequences were obtained. Twenty EST-SSR markers could be introduced for the selection of tolerant varieties to salt stress in rice by analysis of contigs and singletons. In contigs and singletons, significant common gene ontology terms are mainly related to the single-organism cellular process and response to abiotic stresses. The highest percentage of transcription factors for contigs and singletons was related to ERF, Dof, MYB, C2H2, BBR-BPC, bZIP, and WRKY. Moreover, the HSP81-2 (heat shock protein 81-2) and regulators of complement activation were identified as proteins of hub genes that were related to the salt stress tolerance mechanisms. Three uncharacterized hub genes (OS02T0161900-01, OsJ_19443, and OsJ_04035) including EST-SSRs for function identification were investigated by the 3D protein structure homology-modeling. OS02T0161900-01 as tetra ubiquitin, OSj-19443 as a serine/threonine-protein kinase/endoribonuclease, and OSj-04035 as a triosephosphate isomerase were identified. Most of the hub genes were related to environmental stresses and our findings provided candidate genes and transcription factors involved in salinity stress. The development of functional markers associated with abiotic stress tolerance will be helpful to facilitate rice breeding programs. However, before using these markers, laboratory confirmation is necessary.


Main Subjects

Article Title [Persian]

توسعه مارکرهای مولکولی EST-SSR در برنج (Oryza sativa L.) تحت تنش شوری و شناسایی ژن های کلیدی

Abstract [Persian]

در گیاهان EST-SSRs یا توالی­ های ساده تکراری بیان شونده برای بررسی تنوع ژنتیکی و توسعه مارکرهای مولکولی مرتبط با تنش ­های زیستی و غیرزیستی استفاده می­ شوند. با وجود این، در برنج مارکر­های مولکولی محدودی در رابطه با EST برای سازش به تنش­ های غیرزیستی از جمله شوری برای استفاده در برنامه­ های اصلاح نباتات وجود دارد. از میان 8299 عدد EST  قابل دسترس در پایگاه داده NCBI برای تنش شوری در برنج، 525 contig  و 1139 singleton  به دست آمد. برای گزینش ارقام متحمل به تنش شوری در برنج 20 مارکر مولکولی EST-SSR از طریق آنالیز این contigها  و singletonها تعیین شد. آنتولوژی ژن مشترک و معن ی­دار برای contigها  و singletonها عمدتا مرتبط با فرایند سلولی و پاسخ به تنش ­های غیرزیستی بود. بیشترین درصد فاکتورهای رونویسی برای contigها  و singletonها با ERF، Dof،MYB ،C2H2 ، BBR-BPC،bZIP  وWRKY  مرتبط بود. علاوه بر این، پروتئین شوک حرارتی HSP81-2 و تنظیم‌کننده‌های فعال‌سازی مکمل (RCA) به‌عنوان محصول­ های ژن‌های هاب شناسایی شدند که با مکانیسم‌های تحمل به تنش شوری مرتبط بودند. سه ژن hub ناشناخته (OS02T0161900-01، OsJ_19443 و OsJ_0403) حاوی مارکر­های EST-SSR برای شناسایی عملکردشان از طریق مدل سازی همسان سازی ساختار پروتئین سه بعدی مورد بررسی قرار گرفتند و محصول OS02T0161900-01 به عنوان تترا یوبیکوئیتین،  OSj-19443  به عنوان سرین/ترئونین پروتئین کیناز/اندوریبونوکلئاز و OSj-04035 به عنوان ایزومراز تریوزفسفات شناسایی شد. اکثر ژن­ های hub به دست آمده در تحقیق حاضر مرتبط با تحمل به تنش­­ های غیرزیستی بودند که می ­توان آن ­ها را به عنوان ژن­ های کاندید و فاکتورهای رونویسی درگیر در تحمل به تنش شوری منظور کرد. توسعه مارکرهای مولکولی عملکردی می­ تواند در تسهیل برنامه ­های اصلاح برنج موثر واقع شود. باوجوداین، قبل از استفاده از این نشانگرها تائید آزمایشگاهی آن­ ها لازم است.

Keywords [Persian]

  • برنج
  • تنش غیرزیستی
  • شناسایی ژن
  • شوری
  • عملکرد
  • singleton
  • contig
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