Abstract
BackgroundIn crop plants, to cope up with the demand of food for rising population, revolutionary crop improvement programmes are being implemented for higher and higher yields. Abiotic stress, especially at flowering stage, causes drastic effect on yield in plants. Deforestation and urbanization made the water table very low and changed the climate which led to untimely and unforeseen rains which affect the yield of a crop through stress, both by lack of water as well as water logging (abiotic stress). Development of tolerant plants through breeding is a time-consuming programme and does not perform well in normal conditions. Development of stress-tolerant plants through transgenic technology is the better solution. Maize is a major crop used as food and fodder and has the commercial value in ethanol production. Hence, the genes viz., nced (9-cis-epoxycarotenoid dioxygenase) and rpk (receptor-like protein kinase), which play the key roles in the abscisic acid pathway and upstream component in ABA signaling have been transferred into maize plants through Agrobacterium-mediated transformation by optimizing several parameters to obtain maximum frequency of transformation. ResultsCultures raised from immature embryos of 2-mm size isolated from maize cobs, 12–15 days after pollination, were used for transformation. rpk and nced genes under the control of leaP and salT promoters respectively, cloned using gateway technology, have been introduced into elite maize inbred lines. Maximum frequency of transformation was observed with the callus infected after 20 days of inoculation by using 100 μM acetosyringone, 10 min infection time, and 2 days incubation period after co-cultivation resulted in maximum frequency of transformation (6%) in the NM5884 inbred line. Integration of the genes has been confirmed with molecular characterization by performing PCRs with marker as well as gene-specific primers and through southern hybridization. Physiological and biochemical characterization was done in vitro (artificial stress) and in vivo (pot experiments). ConclusionsChanges in the parameters which affect the transformation frequency yielded maximum frequency of transformation with 20-day-old callus in the NM5884 inbred line. Introducing two or more genes using gateway technology is useful for developing stable transgenic plants with desired characters, abiotic stress tolerance in this study.
Highlights
In crop plants, to cope up with the demand of food for rising population, revolutionary crop improvement programmes are being implemented for higher and higher yields
These two inbred lines which were found to be more regenerative and yielded larger quantities of E.callus have been used in the present study by following in vitro practices adopted in our previous studies viz., Linsmaier and Skoog’s (LS) medium [28] supplemented with 1.5 mg/L 2, 4-D for callus induction and LS with BAP and Kn (0.5 mg/L each) for regeneration [19]
Maximum survival and regeneration frequencies of 46 and 25 were found in NM5884 inbred line where infection was done after 20 days, and lowest survival (1%) and regeneration (1%) frequencies were observed in NM74C inbred line
Summary
To cope up with the demand of food for rising population, revolutionary crop improvement programmes are being implemented for higher and higher yields. Development of tolerant plants through breeding is a time-consuming programme and does not perform well in normal conditions. Plants are sessile and exposed to different biotic and abiotic stresses, which refers to the conditions imposed externally and cause drastic effect in their growth and development which influence the productivity of the plant. Most of the plants and or crops are sensitive to abiotic stresses viz., drought, salinity, temperatures, and other environmental extremes. For compensating such adverse conditions, plants develop unique mechanisms to react with the tough environmental conditions by changing themselves, flexible to adapt the conditions. Molecular genetics and biotechnology play an important role to get the resistant plants either by enhancement of native genes or by introducing the resistance genes from the other plant species under the control of stress-induced promoter
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