Improving fruit quality in tomato (Lycopersicum esculentum Mill) under heat stress by silencing the vis 1 gene using small interfering RNA technology

Abstract

Constantly elevated temperatures cause an array of physio-biochemical changes in tomato (Lycopersicum esculentum Mill.) which make the fruit ripen quickly and up to 50 % yield loss. The development of tomato cultivars, using genetic engineering approaches which delay ripening, offers a new way to keep tomatoes healthy under heat stress. Over-expression of small heat shock protein gene, viscosity 1 (vis 1) plays a role in increasing juice viscosity, early ripening and tissue soften which emphasizes the importance of this gene in premature ripening. The aim of this work was therefore to develop a useful system for silencing the vis1 gene using small interfering RNA strategy. Agrobacterium strain GV3101 harbouring the binary vector pICBV19 containing the gus and bar genes was used to adapt the transformation process in this study. The primers were designed to amplified the first exon of the vis 1 gene and the amplified fragment was used for cloning into the pFGC5941 at XhoI/ NcoI site at sense orientation then additional fragment was subsequently cloned at BamHI/XbaI to form sense/antisense cloned fragment interrupted by the CHSA-intron Agrobacterium strain LBA4404 with the binary vector pFGC5941 which harbors vis1 gene under the control of the 35S promoter containing bar gene under the control of a mannopine synthase 2 0 (Mas2 0 ) as selectable marker, was used to reduce the expression of vis1 gene in fruit. Polymerase chain reaction (PCR), RT-PCR and northern blotting analysis were applied to detect putative transgenic plants. Significantly, silencing of vis 1 gene was potently occur and new transgenic tomato cultivars were produced with enhanced ripening qualities for recommendation for growing under heat stress.

Impact of application of zinc oxide nanoparticles on callus induction, plant regeneration, element content and antioxidant enzyme activity in tomato (Solanum lycopersicum Mill.) under salt stress

“…Subsequently, they were maintained in a 16-h photoperiod under 60 µmol m-2 s -1 illumination supplied by cool white fluorescent light for three weeks. Cotyledon explants were excised from 27-day-old seedlings cut into halves with a sterile scalpel blade and placed abaxial side facing down on callus induction medium as previously described [23]. Callus induction medium (M2) was prepared as previously described [24].…”

Tomato tolerance to abiotic stress: a review of most often engineered target sequences

“…In plants, RNAi can be induced in two ways: by a transgene (TIGS, transgene induced gene silencing) or a virus (VIGS, virus-induced gene silencing). The first approach was used for instance to silence gene vis1 (viscosity) in tomato fruit to obtain transgenic lines with delayed ripening under heat stress (Metwali et al 2015). Since VIGS represents a useful tool for the identification of gene function, in the other approach different types of viruses (e.g.…”

AGROBACTERIUM-MEDIATED TRANSFORMATION OF TOMATO (LYCOPERSICON ESCULENTUM Mill.) USING A SYNTHETIC Cry1Ab GENE FOR ENHANCED RESISTANCE AGAINST TUTA ABSOLUTA (Meyrick)

“…Agrobacterium strain GV3101 harbouring the binary vector pICBV19 containing the gus and bar genes (Figure 1) was used to adapt the transformation process in tomato (Ehab et al, 2015). Cry1Ab gene was subcloned into the BamHI site of pBI121.…”