Temporal and spatial control of gene expression inhorticultural crops

Biotechnology provides plant breeders an additional tool to improve various traits desired by growers and consumers of horticultural crops.It also provides genetic solutions to major problems affecting horticultural crops and can be a means for rapid improvement of a cultivar.With the availability of a number of horticultural genome sequences,it has become relatively easier to utilize these resources to identify DNA sequences for both basic and applied research.Promoters play a key role in plant gene expression and the regulation of gene expression.In recent years,rapid progress has been made on the isolation and evaluation of plant-derived promoters and their use in horticultural crops,as more and more species become amenable to genetic transformation.Our understanding of the tools and techniques of horticultural plant biotechnology has now evolved from a discovery phase to an implementation phase.The availability of a large number of promoters derived from horticultural plants opens up the field for utilization of native sequences and improving crops using precision breeding.In this review,we look at the temporal and spatial control of gene expression in horticultural crops and the usage of a variety of promoters either isolated from horticultural crops or used in horticultural crop improvement.

An optimized procedure for plant recovery from somatic embryos significantly facilitates the genetic improvement of Vitis

Plant regeneration from grapevine(Vitis spp.)via somatic embryogenesis typically is poor.Recovery of plants from Vitis rotundifolia Michx.(muscadine grape)is particularly problematic due to extremely low efficiency,including extended culture durations required for embryo–plant conversion.Poor plant recovery is an obstacle to the selection of improved genetically modified lines.Somatic embryos(SEs)of V.rotundifolia cultivar Delicious(Del-HS)and Vitis vinifera L cultivar Thompson Seedless(TS)were used to identify culture media and conditions that promoted embryo differentiation and plant conversion;this resulted in a two-step culture system.In comparative culture experiments,C2D medium containing 6%sucrose was the most effective,among four distinct formulae tested,for inducing precocious SE germination and cell differentiation.This medium,further supplemented with 4 mM 6-benzylaminopurine(C2D4B),was subsequently determined to enhance post-germinative growth of SE.MS medium supplemented with 0.5 mM 1-naphthaleneacetic acid(MSN)was then utilized to stimulate root and shoot growth of germinated SE.An average of 35%and 80%‘Del-HS’and‘TS’SE,respectively,developed into plants.All plants developed robust root and shoot systems and exhibited excellent survival following transfer to soil.Over 150 plants of‘Del-HS’were regenerated and established within 2.5 months,which is a dramatic reduction from the 6-to 12-month time period previously required.Similarly,88‘TS’plant lines were obtained within the same time period.Subsequently,seven out of eight Vitis cultivars exhibited significantly increased plant conversion percentages,demonstrating broad application of the two-step culture system to produce the large numbers of independent plant lines needed for selection of desired traits.

Production of Transgenic <i>Camelina sativa</i>Plants via <i>Agrobacterium</i>-Mediated Transformation of Shoot Apical Meristems

A method to produce transgenic Camelina sativa plants in cvs. PI650159 and PI650161 was developed. Micropropagated shoot meristem cultures were established from in vitro germinated seedlings and used as target tissues for Agrobacterium-mediated transformation. A plasmid harboring enhanced green fluorescent protein, β glucuronidase and neomycin phosphotransferase II genes were used to optimize parameters for transgenic plant production. Kanamycin at 40 mg&middot;l-1 was effective in suppression of non-transformed cells while permitting growth of transgenic tissues. Shoot apical meristems co-cultivated with Agrobacterium exhibited stable enhanced green fluorescence protein (EGFP) and β glucuronidase (GUS) expression after culture on plant regeneration medium. We observed transformation efficiencies of 53.33% in cv. PI650159 and 98.33% in cv. PI650161. The presence of transgenes in both cultivars was confirmed by PCR, while quantitative real-time PCR detected single copy integration in Pl650161 and two copy integration in Pl650159. Transgenic plants exhibited EGFP and GUS expression in all tissues including shoots, leaves, buds, floral organs, seeds, and pods. Our results demonstrate a simple and efficient technique using apical shoot meristems for production of transgenic C. sativa plants that can be used for transfer of desirable traits.

Clonal Fidelity of Micropropagated Psidium guajava L. Plants Using Microsatellite Markers

Micropropagation of Psidium guajava L. (guava) is a viable alternative to currently adopted techniques for large-scale plant propagation of commercial cultivars. Assessment of clonal fidelity in micropropagated plants is the first step towards ensuring genetic uniformity in mass production of planting material. In the present study, 31 plants of guava cultivar “Lucknow 49” regenerated by micropropagation were tested for genetic fidelity by comparing them to the mother plant from which explant material was obtained. Efficient rooting of in vitro proliferated shoots was obtained by culture on 1/2 strength MS medium supplemented with either 9.8 μM indole butyric acid (IBA) or 11.4 μM indole acetic acid (IAA). Leaf samples of 31 regenerated plants were compared to the mother plant using 17 simple sequence repeat (SSR) markers. While 16 SSRs detected the same allele, locus mPgCIR07 detected slight differences, where six micropropagated plants were 1 bp smaller (152 bp) than the parental genotype (153 bp). Differences in leaf tissues for anthocyanin pigmentation were also noted among micropropagated plants. Results of the study indicated efficient rooting of “Lucknow-49” cultivar for rapid propagation of planting material, and revealed that micropropagated plants were identical for 16 of the 17 loci examined. Although most mutations induced by tissue culture may not have an effect on phenotype, the possibility that novel phenotypes can be generated in a commercial setting exists.