Regeneration and genetic transformation of Populus deltoides Bartr ex Marsh for cellulose enhancement

Abstract

Wood is the primary raw material for the production of pulp and paper. The Indian pulp and paper sector has made considerable progress during the last decades and characterized one of the key industries in India with a turnover of about 6.5 billion euros. India is considered the fastest-growing paper market globally. Populus spp. is gaining attention because of its faster growth and multipurpose wood. Species of Populus and their hybrids are grown for various uses such as plywood, timber, paper, fuelwood, and bioenergy production. Because of its higher commercial value, there is a need to adopt strategies for developing productive clones of Populus. Populus deltoides, a native of North America, is a widely planted species in India. For establishment of micropropagation protocol Populus deltoides elite clones G48 and L34 were selected for the present study based on their higher biomass productivity and wood quality. Further based on the regeneration and transformation efficiencies P. deltoides clone G48 was used to enhance the cellulose content to increase its economic value. A successful micropropagation and regeneration protocol was established. Agrobacterium tumefaciens mediated transformation protocol was optimized for cloning of two essential genes, Korrigan and sucrose synthase, directly involved in cellulose biosynthesis. Southern blot analysis was used to confirm the stable integration of transgene and check the copy number of the gene, which was further confirmed by qRT-PCR. The genes were successfully cloned, and their expression pattern was studied. This study has established a highly efficient regeneration and transformation protocols. The direct shoot organogenesis was achieved from leaf explant of two commercially important clones of Populus deltoides on MS medium enriched with 15 mg/L adenine sulphate, 5 mg/L Ascorbic acid, 250 mg/L (NH4)2 SO4 (referred to as PD1 medium) supplemented with 2.5 µM each of 6-benzylaminopurine and indole-3-acetic acid.