In vitro minimal growth storage of Saccharum spp. hybrid (genotype 88H0019) at two stages of direct somatic embryogenic regeneration

Abstract

In vitro culture via somatic embryogenesis of Saccharum spp. germplasm is used routinely in our laboratories for micropropagation and production of transgenic lines. At times, due to greenhouse and field constraints, it is necessary to hold back material in culture. For this purpose, methods were established to slow down growth and development at two stages of a direct somatic embryogenesis protocol, viz. before embryo maturation and before plantlet acclimatization. Storage of globular somatic embryos of a single genotype, 88H0019, was achieved by transferring three-week old cultures to half-strength Murashige and Skoog (MS) basal salts and vitamin medium, 5 g l -1 sucrose, 0.6 mg l -1 2,4-dichlorophenoxyacetic acid (2,4-D), 0.5 g l -1 casein hydrolysate, 8 g l -1 agar, pH 5.8, at both 18 and 24 ± 2°C for 12 weeks, after which they were transferred to standard regeneration medium. For storage of whole plantlets, the highest survival rates and shoot regrowth after 8 months was observed at 18°C on halfstrength MS and 10 g l -1 sucrose, with or without 30 g l -1 sorbitol. Genetic integrity of the plants was established by amplified fragment length polymorphism (AFLP) analysis and a low polymorphic rate of 0.005% was observed. Phenotypic stability was investigated by conducting standard agronomic and yield measurements after a further 8 months of field growth. Although stalk diameter was significantly smaller from all the minimal growth-derived plants, sucrose yield was not compromised when compared to a conventionally propagated field control.

EfficientIn VitroRegeneration of Sugarcane (Saccharum OfficinarumL.) from Bud Explants

“…Despite the interest in S. officinarum, there are only a few earlier reports on its regeneration through shoot tip cultures (11,17,18,34), apical meristem (10,33), leaf sheath (14), shoot tops (35), young leaf segments (19), nodal buds (8), young meristematic leaf sheath (31), immature leaf explants (39), leaf spindle explants, germinated seeds (9), shoot apex (22), tops with growing apices (30) The aim of the present study was to develop an efficient, reliable and promising protocol for regeneration of S. officinarum from bud explants.…”

Cryopreservation of in vitro-grown shoot-tips of Rabdosia rubescens by encapsulation-dehydration and evaluation of their genetic stability

“…In vitro tissue culture conservation is susceptible to contamination and somaclonal variation (Watt et al 2009;Gonçalves et al 2010). Cryopreservation is the most promising choice for conservation of vegetatively propagated plants owing to its safety, repeatability and long-term storage possibility (Pacheco et al 2009;Hazubska-Przbyl et al 2010).…”

A two-stage pretreatment of seedlings improves adventitious shoot regeneration in sugar beet (Beta vulgaris L.)

“…Sugar beet is known to be a recalcitrant species with respect to in vitro culture. Also, a high degree of genotypic variation, primarily because of its highly heterozygous nature due to outcrossing, is a serious problem for the optimization of both regeneration (Gurel 1997;Gurel et al 2001;Zhang et al 2008) and transformation protocols in sugar beet (Krens et al 1996;Zakharchenko et al 2000;Hisano et al 2004;Ninkovic et al 2010) as well as in many other species (Arellano et al 2009;Watt et al 2009;Liu et al 2010;Kwapata et al 2010). The effects of pretreating leaf explants or preconditioning the source of explants (seedlings) on the subsequent regeneration efficiency were previously reported for sugar beet (Jacq et al 1993;Zhong et al 1993a, b;Zhang et al 2001;Gurel et al 2003a) and several other species (D'Onofrio and Morini 2006;Thomas 2007).…”