Biolistic co-transformation of rice using gold nanoparticles

Document Type : Full Article

Authors

1 Agricultural Biotechnology Research Institute of Iran (ABRII), Agricultural Research, Education and Extension Organization (AREEO), Karaj, I. R. Iran

2 Department of Plant Inbreeding, College of Agriculture, Zanjan University, Zanjan,, I. R. Iran

Abstract

ABSTRACT- In order to produce transgenic rice lines lacking selectable marker gene, biolistic co-transformation technique using gold nanoparticles was adopted. In the first step, the efficiency of different sizes of gold particles was evaluated. The results showed that the efficiency of the nanoparticles in the transformation was comparable to that of the micro particles. Subsequently, two separate plasmids including p UBC harboring cry IA (c) gene and pTRA132 carrying hph gene as selectable marker gene were mixed and coated with gold nanoparticles. Embryo genic calli originated from mature rice seeds were targeted with plasmid-coated nan projectiles. Putative transgenic cells were selected from selection media after three subcultures in MS medium were supplemented with 50 mg hygromycine B l-1 in three weeks' intervals. The selected calli were then grown into putative transgenic plantlets. Molecular analysis performed on the regenerated lines showed that at least one copy of both genes was inserted into the genome of some transgenic lines. RT-PCR analysis also confirmed successful transcription of the cry IA (c) gene. Accordingly, it could be concluded that the biolistic co-transformation method using gold nanoparicles could be a successful method for gene delivery into rice varieties. Nevertheless, expression of cry IA (c) gene and loci of both genes should still be investigated in future studies.

Keywords

Article Title [Persian]

تراریزش توام گیاه برنج به روش زیست‌پرتابی با استفاده از نانوذرات طلا

Authors [Persian]

  • سید الیاس مرتضوی 1
  • زهرا ظهرابی 2
1 پژوهشکده بیوتکنولوژی کشاورزی ایران، سازمان تحقیقات، آموزش و ترویج کشاورزی، کرج، ج. ا. ایران
2 گروه اصلاح نباتات کشاورزی، دانشکده کشاورزی، دانشگاه زنجان، زنجان، ج. ا. ایران
Abstract [Persian]

چکیده-به منظور تولید لاین‌های برنج تراریخته مقاوم به آفات که فاقد ژن نشانگر انتخابی باشد، تکنیک ریزپرتابی با استفاده از نانوذرات طلا مورد استفاده قرار گرفت. در نخستین گام، کارایی نانوذرات طلا با اندازه‌های مختلف مورد ارزیابی قرار گرفت. نتایج نشان داد که نانوذرات طلا می‌توانند به اندازه میکروذرات در انتقال ژن کارایی داشته باشند. در گام بعد، دو پلاسمید مجرا مورد استفاده قرار گرفتند. پلاسمید puBC حامل ژن cry1A(c) و فاقد ژن مارکر انتخابی به همراه پلاسمید  pTRA132 حاوی ژن مقاومت به هیگرومایسین به عنوان نشانگر انتخابی مخلوط شده و سپس با نانوذرات طلا پوشش داده شدند. این مخلوط پلاسمیدها به کالوس‌های جنین‌زایی که از بذور رسیده منشا گرفته و دارای توان تقسیم سلولی و باززایی بالایی بودند، به روش ریزپرتابه وارد شدند. سلول‌های تراریخته احتمالی از بافت‌های بمباران شده پس از 3 دوره گزینش در محیط کالوس‌زایی N6 حاوی 50 میلی‌گرم در لیتر هیگرومایسین B مورد گزینش قرار گرفتند. نهایتاً کالوس‌های مقاوم به هیگرومایسین در محیط باززایی MS  حاوی 50 میلی‌گرم در لیتر هیگرومایسین B باززا شدند. در نهایت گیاهان تراریخته احتمالی حاصل با آنالیزهای مولکولی مورد بررسی قرار گرفتند. این آنالیزها نشان داد که حداقل یک نسخه از ژن مورد نظر به گیاهان تراریخته وارد شده است. آنالیز RT-PCR نیز تأیید کرد که ژن cryIA(c) در گیاهان تراریخته قابلیت رونویسی و بیان ژن را دارد. لذا می‌توان استنباط کرد که استفاده از روش ریزپرتابه و نانوذرات طلا می‌تواند یه روش کارا و موثر در انتقال ژن و تولید گیاهان تراریخته به ویژه در گیاه برنج مورد استفاده قرار گیرد.

Keywords [Persian]

  • واژه‏های کلیدی:
  • ریزپرتابی
  • تراریزش توأم
  • cryIA(c)
  • نانوذرات طلا
  • برنج

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Iran Agricultural Research
Volume 37, Issue 1
June 2018
Pages 75-82

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