Callus Induction and Plant Regeneration of Chrysanthemum morifolium and C. coccineum via Direct and Indirect Organogenesis and Genetic Fidelity Analysis Using IRAP, ISSR and SCoT Molecular Markers
محورهای موضوعی : مجله گیاهان زینتی
Fardin Nasri
1
(
Department of Horticultural Sciences, Faculty of Agricultural Sciences, University of Guilan, Rasht, Iran
)
Hedayat Zakizadeh
2
(
Department of Horticultural Sciences, Faculty of Agricultural Sciences, University of Guilan, Rasht, Iran
)
Yavar Vafaee
3
(
Department of Horticultural Science, Agriculture Faculty, University of Kurdistan, Sanandaj, Iran
)
Ali Akbar Mozafari
4
(
Department of Horticultural Science, Agriculture Faculty, University of Kurdistan, Sanandaj, Iran
)
کلید واژه: Callus induction, Embryogenesis, Cultivar response, Direct regeneration, Genetic fidelity,
چکیده مقاله :
In vitro propagation of C. morifolium cv. ‘Homa’ and cv. ‘Delkash’ and wild C. coccineum via direct and indirect organogenesis and somatic embryogenesis were investigated. BAP at 0, 1, 2 and 3 mg l-1 or NAA at 0, 0.05, 0.1 and 0.2 mg l-1 concentrations were used to induce direct and indirect organogenesis of shoot tip explants. To study the callus induction and somatic embryogenesis, the young leaf explants were cultured on MS medium containing BAP (0, 1, 2 or 3 mg l-1) and 2,4-D (0, 1, 2 or 3 mg l-1). Direct shoot regeneration was achieved from shoot tip explants of ‘Homa’ and ‘Delkash’ as well as C. coccineum. The highest number of shoots through direct regeneration (13.78 and 8.89 shoots per explant for C. coccineum and C. morifoilum ‘Homa’, respectively) were observed in the treatment with 2 mg l-1 BAP and 0.05 mg l-1 NAA. In both species, the highest frequency of callus formation and embryogenesis were obtained on medium containing 2.0 mg l-1 2,4-D and 2 mg l-1 BAP. Genetic fidelity of 10 acclimatized plants derived from direct regeneration of each species was confirmed using six inter-retrotransposon amplified polymorphism (IRAP), inter-simple sequence repeat (ISSR) and start codon targeted (SCoT) primers. A total of 56, 56 and 39 fragments were amplified for IRAP, ISSR, and SCoT, respectively. In general, our results showed that finding a better response of explants to embryogenesis or organogenesis in a specific cultivar and with special PGRs combinations and concentrations play an important role in the in vitro propagation efficiency of chrysanthemum species.
در این تحقیق، تکثیر درون شیشهای داوودی گونه C. morifolium ارقام ’هما‘ و ’دلکش‘ و گونه بومی (C. coccineum) از طریق اندام زایی مستقیم، غیر مستقیم و جنین زایی غیرجنسی گزارش شده است. BAP در غلظتهای صفر، 1، 2 و 3 میلیگرم در لیتر و NAA در غلظتهای صفر، 05/0، 1/0 و 2/0 میلیگرم در لیتر جهت القای اندامزایی مستقیم و غیر مستقیم در ریزنمونههای نوک شاخههای انتهایی استفاده شد. جهت مطالعه القای کالوس و جنین زایی غیرجنسی، ریزنمونههای جوان برگ بر روی محیط کشت MS حاوی BAP (صفر، 1، 2 و 3 میلیگرم در لیتر) و 2و4-دی (صفر، 1، 2 و 3 میلیگرم در لیتر) کشت شدند. باززایی مستقیم شاخه از ریزنمونههای نوک شاخههای انتهایی ارقام ’هما‘ و ’دلکش‘ و همچنین در گونه بومی C. coccineum مشاهده شد. بالاترین تعداد شاخهها از طریق اندامزایی مستقیم (78/13 و 89/8 شاخه در هر ریزنمونه به ترتیب برای C. coccineumو ’هما‘) با 2 میلی گرم در لیتر BAP و 05/0 میلیگرم در لیتر NAA مشاهده شد. در هر دو گونه، بالاترین فراوانی تشکیل کالوس و جنینزایی روی محیط کشت حاوی 2 میلیگرم در لیتر 2و4-دی و 2 میلیگرم در لیتر BAP بدست آمد. پایداری ژنتیکی 10 گیاه سازگار یافته حاصل از اندامزایی مستقیم از هر گونه با استفاده از6 پرایمر از نشانگرهای مولکولی (IRAP, ISSR, SCoT) به اثبات رسید. در کل 56، 56 و 39 قطعه برای IRAP، ISSR و SCoT بهترتیب تکثیر شدند. در کل، نتایج ما نشان داد که یافتن عکسالعمل بهتر ریزنمونهها به جنینزایی یا اندامزایی در یک رقم ویژه و با یک ترکیب هورمونی و غلظتهای آنها نقش مهمی را در کارآیی تکثیر درون شیشهای گونههای داوودی، بازی میکند.
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