In vitro regeneration and optimization of conditions for Agrobacterium mediated transformation in jute, Corchorus capsularis

Abstract

Jute is a crop of commercial importance that is widely cultivated for its bast fiber production but susceptible to many diseases that results in major economic loss. New genes can be introduced into this plant through Agrobacterium mediated genetic transformation for its genetic improvement, which is dependent on the availability of suitable in vitro techniques. An efficient regeneration system has been developed for in vitro culture of jute (Corchorus capsularis) from the distal cut ends of cotyledonary petioles. High frequency shoot regeneration was obtained on Murashige and Skoog (MS) nutrient agar medium supplemented with 0.5 mg/l NAA, 0.5 mg/l BAP and 36 g/l sucrose. On transfer to soil, the regenerated plantlets survived and appeared to be morphologically similar to the normal seed-grown plants. They developed pods and set fertile seeds. Histological analysis revealed de novo origin of shoot buds in the in vitro cultured cotyledonary petioles. Parameters affecting transformation were optimized by assaying GUS activity in these regenerable tissues after cocultivation with Agrobacteria. These tissues appear to be susceptible for infection and transformation by Agrobacterium carrying uid (GUS INT) and nptII genes, as well as shoot multiplication. The cells at the cut end of the petioles were found competent to take up the DNA, which was monitored by transient GUS gene expression. EHA105 at 0.3 O.D and LBA4404 at 0.5 O.D were found to be compatible in giving optimal levels of transient GUS expression.

Establishment of Efficient Genetic Transformation Systems and Application of CRISPR/Cas9 Genome Editing Technology in Lilium pumilum DC. Fisch. and Lilium longiflorum White Heaven

“…However, the receptor materials might become very fragile if the infection duration is too long. Similarly, a low concentration of bacterial solution may not provide sufficient Agrobacterium to ensure effective infection, while a high concentration might lead to a rapid increase in bacterial growth, causing excessive damage to the receptor materials [54,55]. In this study, the transformation efficiencies with different bacterial concentrations and infection durations were significantly different.…”

Studies on Explant Regeneration and Protoplast Culture from Hypocotyl Segments of Jute, Corchorus capsularis L.

“…Similar observations were made by Sihachkr and Ducreux (1987) in sweet potato. Micro-calli were obtained after 60 days of repeated dilutions with medium but regeneration was not obtained even on the most responsive medium standardized for C. capsularis (Pushyami et al, 2011) for petiole explants. Though 7.5 mgl -1 6-BAP along with 50 mgl -1 AdSO 4 was proved beneficial for hypocotyl regeneration in JRC 698, no shoot regeneration could be observed even after 30 days of callus establishment in protoplast cultures.…”

“…C. capsularis (white jute) is one of the two cultivated species in the genus. In vitro plant regeneration protocols using cotyledon and petiole attached cotyledon explants for different cultivars from both the species are well documented (Abbas et al, 1997;Islam et al, 1982;Seraj et al, 1992;Saha et al, 1999;Huda et al, 2007;Sarker et al, 2007;Pushyami et al, 2011). Jute considered recalcitrant to regeneration and the process being genotype dependent (Naher et al, 2003), different types of explants need to be tried for establishing a repeatable regeneration protocol.…”

Multiple Shoot Regeneration from Excised Embryonic Axis Explants of Jute,Corchorus capsulariscv. JRC321

“…But, due to the highly recalcitrant nature and genotype dependent of the two cultivated species of Corchorus in tissue culture, major improvement of this crop by means of genetic transformation could not be achieved yet 17 . However, standardization of preliminary transformation and regeneration using cotyledonary petiole explants of jute (JRC321) was documented 18 . Also, limited plantlets (2-3 shoots) regeneration using shoot meristem-tip and epicotyl explants of seedling, and immature shoot bud explant after Agrobacterium transformation 19 .…”