A fully in vitro protocol towards large scale production of recombinant inbred lines in wheat (Triticum aestivum L.)
- 417 Downloads
A bottleneck for genetic research and breeding of crop plants is the time taken to producing large pure line segregating populations so called recombinant inbred lines (RILs). One way to overcome this problem is through use of the single-seed-decent (SSD) process under in vitro conditions. A number of factors that may affect in vitro SSD approach of wheat including temperature, light duration and intensity, salt strength and carbohydrate concentration were investigated in this study. Under the in vitro conditions, 45 days per generation was achieved for an early flowering wheat genotype Emu Rock, allowing eight generations per annum; 58 days per generation was achieved for mid flowering genotypes, allowing six generations per annum. The results showed that a variation of growth environment before and after three-leaf stage allowed in vitro seed-set with a relatively short generation time. Specifically, the plantlets were first grown under 22 °C with a light intensity of 145 μmol m−2 s −1 (16 h d−1) for 20 days (around three-leaf stage), and then moved to an environment of 28 °C and 500 μmol m−2 s−1 (20 h d−1) light. The culture medium was 1/2 strength Murashige and Skoog (MS) with modification of adding ten times of extra KH2PO4 and 4% sucrose. The fully in vitro protocol resulted in 100% flowering rate and average seed set rate of 91.5% in Emu Rock and Zippy. It can be further fine-tuned to suit different genotypes and it has a potential for factory scale mass-production of RILs for genetic studies and practical breeding programs.
KeywordsWheat Triticum aestivum L. Fast generation cycling Single-seed-decent Flowering and seed-set in vitro
Financial supports from the Australian Council of Grain Grower Organisations (COGGO) Research Fund 2014–2015, National Natural Science Foundation of China (41373118), and Guangzhou Education Bureau Innovation Team Project (No.13C02) are appreciated. Y. Yao would like to thank China Scholarship Council (CSC) for her visiting scholarship at The University of Western Australia.
YY, HL, ZL, GY conducted experimental design, data analysis and manuscript writing. PZ was involved in strategic experimental input. YY and HL conducted the in vitro experiments.
Compliance with ethical standards
Conflict of interest
The authors declare that they have no conflict of interest.
- Croser JS, Pazos-navarro M, Bennett RG, Tschirren S, Edwards K, Erskine W, Creasy R, Ribalta FM (2016) Time to flowering of temperate pulses in vivo and eneration turnover in vivo–in vitro of narrow-leaf lupin accelerated by low red to far-red ratio and high intensity in the far-red region. Plant Cell Tissue Organ Cult. doi: 10.1007/s11240-016-1092-4 Google Scholar
- El-Hennawy MA, Abdalla AF, Shafey SA, Al-Ashkar IM (2011) Production of doubled haploid wheat lines (Triticum aestivum L.) using anther culture technique. Ann Agric Sci 56:63–72Google Scholar
- Liu D, Liu W, Zhu D, Jin H, Guo L, Zuo Z, Liu L (2010) Effect of phosphate fertilizer supply on growth, yield and nutrient absorption of Chrysanthemum morifolium (Ramat.). S Chin J Agr Sci 123:1575–1580Google Scholar
- Nelson MN, Berger JD, Erskine W (2010) Flowering time control in annual legumes: prospects in a changing global climate. CAB Rev Perspect Agric Vet Sci Nutr Nat Res 5:49–62Google Scholar
- Ribalta FM, Pazosnavarro M, Nelson K, Edwards K, Ross JJ, Bennett RG, Munday CM, Erskine W, Ochatt SJ, Croser J (2016) Precocious floral initiation and identification of exact timing of embryo physiological maturity facilitate germination of immature seeds to truncate the lifecycle of pea. Plant Growth Regul. doi: 10.1007/s10725-016-0211-x Google Scholar
- Zhang Q, Tu D (1986) Flowered and fruited of the wheat in tube. J Hen Norm Univ 49:105–107Google Scholar
- Ziv M, Naor V (2006) Flowering of geophytes in vitro. Propag Ornam Plants 6:3–16Google Scholar