Influence of polyethylene glycol (PEG) mediated osmotic stress on reactive oxygen species (ROS) scavenging machinery of Chinese potato (Solenostemon rotundifolius (Poir.) J. K. Morton) was investigated. Five genotypes of Chinese potato were raised in Murashige and Skoog (MS) basal medium containing 6-benzylaminopurine (BAP, 1 mg L-1) along with various concentrations of PEG-6000 mediated stress conditions (0,-0.2 and-0.5 MPa) and evaluated for osmotic stress tolerance in vitro. The medium containing PEG-6000 had a detrimental effect on plantlet growth and development while compared with the control. Accumulation of H 2 o 2 was lower in Sreedhara and Subala and higher in Nidhi under PEG stress, which was evident by in situ detection in leaves. Lipid peroxidation product such as malondialdehyde (MDA) content was increased due to PEG stress which was more in susceptible genotype than that in tolerant ones. An enhanced ROS-scavenging antioxidant enzyme was observed under stress with respect to the control. The enzymes of ascorbate-glutathione cycle showed an important role in scavenging ROS. The imposition of PEG stress also increased the non-enzymatic antioxidants viz., the ascorbate and reduced glutathione content which was prominent in tolerant genotypes in comparison to susceptible. The present study indicated that, Sreedhara and Subala showed more tolerance to osmotic stress with better ROS scavenging machineries which would be the lines of interest for augmenting future breeding strategies in this climate resilient minor tuber crop. Chinese potato (Solenostemon rotundifolius (Poir.) J. K. Morton), belongs to the family Lamiaceae, is one of the important minor tuber crops grown in the tropics of the world. The tubers of this crop are rich in carbohydrates and minerals 1 which provide essential dietary and energy supplements during the lean periods. It could be the better alternative to potato (Solanum tuberosum L.) for the tropics of the world under changing climatic conditions; where potato fails to grow due to global warming. Chinese potato can grow in a wide range of environmental conditions. Despite its wide adaptability, the productivity (18-20 t/ha) was severely affected due to osmotic stress. Under natural conditions, plant growth and development often challenged by various stresses 2 , of which, osmotic stress is one of the most limiting factors which can cause 20-98% of yield reductions 3. To elucidate the plant response to osmotic stress, in vitro cultures are preferred as it minimizes nutrient and environmental variations under control conditions 4. In this investigation, we have used polyethylene glycol (PEG-6000) to induce osmotic stress conditions in vitro. In plant osmotic stress studies, PEG is widely used as a potential osmoticum in the nutrient medium to induce water deficit 5. Induced osmotic stress resulted in overproduction of reactive oxygen species (ROS) which was considered as a hallmark of plant stress response. To scavenge the toxic consequences of ROS, plant deploys antioxidative mecha...
A protocol for high-frequency direct organogenesis from root explants of Kachai lemon (Citrus jambhiri Lush.) was developed. Full-length roots (~3 cm) were isolated from the in vitro grown seedlings and cultured on Murashige and Skoog basal medium supplemented with Nitsch vitamin (MSN) with different concentrations of cytokinin [6-benzylaminopurine, (BAP)] and gibberellic acid (GA3). The frequency of multiple shoot proliferation was very high, with an average of 34.3 shoots per root explant when inoculated on the MSN medium supplemented with BAP (1.0 mg L–1) and GA3 (1.0 mg L–1). Optimal rooting was induced in the plantlets under half strength MSN medium supplemented with indole-3-acetic acid (IAA, 0.5–1.0 mg L–1). IAA induced better root structure than 1-naphthaleneacetic acid (NAA), which was evident from the scanning electron microscopy (SEM). The expressions of growth regulating factor genes (GRF1 and GRF5) and GA3 signaling genes (GA2OX1 and KO1) were elevated in the regenerants obtained from MSN+BAP (1.0 mg L-1)+GA3 (1.0 mg L-1). The expressions of auxin regulating genes were high in roots obtained in ½ MSN+IAA 1.0 mg L-1. Furthermore, indexing of the regenerants confirmed that there was no amplicons detected for Huanglongbing bacterium and Citrus tristeza virus. Random amplified polymorphic DNA (RAPD) and inter simple sequence repeat (ISSR) markers detected no polymorphic bands amongst the regenerated plants. This is the first report that describes direct organogenesis from the root explant of Citrus jambhiri Lush. The high-frequency direct regeneration protocol in the present study provides an enormous significance in Citrus organogenesis, its commercial cultivation and genetic conservation.
A protocol for high-frequency direct organogenesis from root explants of Citrus jambhiri Lush. was developed. Full-length roots (~3cm) were isolated from the in vitro grown seedlings and cultured on Murashige and Skoog basal medium supplemented with Nitsch vitamin (MSN) with different concentrations of cytokinin [6-benzylaminopurine, (BAP)] and gibberellic acid (GA3). The frequency of multiple shoot proliferation was very high, with an average of 34.3 shoots per root explant when inoculated on the MSN medium supplemented with BAP (1.0 mg L–1) and GA3 (1.0 mg L–1). Optimal rooting was induced in the plantlets under half strength MSN medium supplemented with indole-3-acetic acid (IAA, 0.5-1.0 mg L–1). IAA induced better root structure than 1-naphthaleneacetic acid (NAA), which was evident from the result of scanning electron microscopy (SEM). The expressions of growth-regulating factor genes (GRF1 and GRF5) and GA3 signaling genes (GA2OX1 and KO1) were high in the regenerants obtained using MSN+BAP (1.0 mg/L)+GA3 (1.0 mg/L). The expressions of auxin regulating genes were high in roots obtained in ½ MSN+IAA1.0 mg L-1. Furthermore, the virus indexing of the regenerants confirmed that there were no virus amplicons detected for Huanglongbing and Citrus tristezavirus. Random amplified polymorphic DNA (RAPD) and inter simple sequence repeat (ISSR) markers detected no polymorphic bands amongst the regenerated plants. The high-frequency direct regeneration protocol in the present study provides an enormous significance in Citrus organogenesis, it’s commercial cultivation and genetic conservation.
An efficient shoot and root organogenesis protocol has been developed from synseeds derived from various explants of Citrus jambhiri Lush. Optimum synseeds were developed using sodium alginate (0.5-0.75%) in 1.0% CaCl2 solution. Shoot organogenesis was examined under various basal vitamin mediums (MS Nitsch and MSB5) supplemented with various concentrations of adenine sulfate (ADS) and 6-benzyl amino purine (BAP) from different explants such as cotyledonary junction, shoot tip, and nodal explants. The synseed regeneration response ranged between 60-100% among the vitamins, cytokinins and explants used. The number of shoots per synseeds was higher (13.4) in MSB5-BAP (1.5 mg L-1), followed by 12.8 in MSN-ADS (2.0 mg L-1), 11.2 in MSN-ADS (1.0 mg L-1), and 10.8 in MSB5-ADS (1.0 mg L-1) from the synseeds developed using 0.75% sodium alginate. The mean number of roots per explant was higher (4.2) in ½MSN+IAA (1.0 mg L-1). Similarly, the mean root length was higher (5.2 cm) in ½MSN+IAA (0.5 mg L-1) followed by 4.2 cm in ½MSN+IAA (1.0 mg L-1). Regenerants derived from synseeds have shown no somaclonal variations, confirming that the plantlets are true-to-type to their parental progenies. The encapsulated plantlets showed >90% survivability while transferred at Kachai village, Manipur, India. The results of the present study encourage the use of various vitamin mediums and explants for large-scale propagation of C. jambhiri through synseeds.
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