In vitro regeneration of sesame (Sesamum indicum L.) from seedling cotyledon and hypocotyl explants

Abstract

The goal of this study was to develop an efficient regeneration protocol to be used for genetic transformation of sesame. Published regeneration methods using benzyladenine (BA) and 1-naphthalene acetic acid (NAA) were unsuccessful for the cultivars used herein. Experiments were carried out using cotyledon and hypocotyl explants from the cultivar Mtwara-2. Later the optimised culture conditions were used to investigate the regeneration response of different genotypes. There was significant interaction between hormone treatments and macronutrients for shoot and root regeneration. Results also showed that shoot regeneration was significantly influenced by explant type. Shoots were only obtained from cotyledons whereas both cotyledons and hypocotyls could produce roots. Modified Murashige and Skoog (MS) medium with N6 macronutrients resulted in twice the shoot regeneration frequency obtained with ½MS macronutrients in the presence of thidiazuron (TDZ). The shoot regeneration frequency was significantly reduced when BA was used in place of TDZ. On shoot regeneration medium containing BA and NAA, only roots were formed. Replacing NAA with indole-3-acetic acid (IAA) greatly improved the regeneration of shoots. The optimum growth regulator combination for shoot regeneration was 20 lM TDZ together with 2.5 lM IAA, which gave a frequency of 63% and 4.4 shoots per regenerating explant for the best cultivar Ex-El. Genotypic differences were significant both for the number of explants regenerating shoots and the number of shoots produced per regenerating explant.

Agrobacterium tumefaciens-mediated genetic transformation of sesame (Sesamum indicum L.)

“…This is the first report of shoot regeneration from the cotyledonary node. In contrast to earlier reports, browning/necrosis and subsequent death of cotyledon explant was not noticed on MS basal medium (Baskaran and Jayabalan 2006) or in the presence of a cytokinin alone (Were et al 2006). This may be due to the different genotype used in these studies.…”

“…Moreover, in those reports the shoot regeneration frequency and the number of shoots per explant were comparatively low as compared with the present study, where regeneration frequency up to 88% and an average of 9.3 shoots/explants were achieved on a simple medium containing BA as sole growth regulator. The higher regeneration efficiency in the present study may be due to the use of the younger cotyledon explants excised from 2-day-old seedling, whereas in earlier reports cotyledon explants older than 3-4 days (Were et al 2006), 8-10 days (George et al 1987;Rao and Vaidyanath 1997b) and 1-2 weeks (Seo et al 2007) were used. Our results are in contrast to an earlier report where regeneration from cotyledons obtained from germinated seeds drastically declined (Seo et al 2007).…”

“…However, the main obstacle to genetic transformation is the recalcitrant nature of sesame to in vitro regeneration (Baskaran and Jayabalan 2006). There are very few reports on shoot regeneration, with low frequencies in a few genotypes from cotyledon and/or hypocotyl explants (Rao and Vaidyanath 1997a;Taskin and Turgut 1997;Younghee 2001;Were et al 2006;Seo et al 2007). Somatic embryos have also been induced from hypocotyl-derived calluses, but no plant regeneration was achieved (Mary and Jayabalan 1997).…”

Sesame

“…Some wild sesame species possess resistance genes, but post-fertilization barriers restrict their transfer to cultivated crops through conventional breeding. Establishment of in vitro plant regeneration is a prerequisite of any genetic transformation system that is already optimized (Were et al 2006 ) . Sesame has been reported as susceptible to Agrobactarium tumefaciens infection (Taskin et al 1999 ) .…”

“…Sesame has been reported as susceptible to Agrobactarium tumefaciens infection (Taskin et al 1999 ) . Protocol for genetic transformation and plant regeneration of sesame were optimized by Were et al ( 2006 ) . A signifi cant interaction between hormonal concentration and macronutrients for plant regeneration was recorded, and application of 20-m M TDZ along with 2.5-m M IAA was found the best for successful plant regeneration.…”