Optimization of regeneration and transformation parameters in tomato and improvement of its salinity and drought tolerance

As part of our efforts to improve tomato tolerance to abiotic stress, we have undertaken this study to introduce two candidate genes encoding: a sodium antiporter and a vacuolar pyrophosphatase, previously shown to enhance drought and salt tolerance in transgenic Arabidopsis plants. First, we evaluated the potential of primary leaves from three to four week-old in vitro-grown tomato seedlings as alternative explants to cotyledons for tomato transformation. Our results demonstrated that primaryleaves are three times more efficient then cotyledons in terms of regeneration percentage, productivity, and transformation frequencies independently of the medium and genetic construct used. Second,primary leaves were used to introduce the genes of interest using Agrobacterium-mediated transformation. Many transgenic tomato plants were easily recovered. The presence of the transgenes and their expression were confirmed by PCR and RT-PCR analysis. The transformation frequencies for primary leaf explants ranged from 4 to 10% depending on the genetic construct used. The time requiredfrom inoculation of primary leaves with Agrobacterium cells to transfer of transgenic tomato plants to soil was only 2 months compared to 3 to 4 months using standard tomato transformation protocols. The transgenic tomato plants obtained in the current study were more tolerant to salinity and drought stress than their wild-type counterparts

Heterologous expression of the yeast Tpo1p or Pdr5p membrane transporters in Arabidopsis confers plant xenobiotic tolerance

This deposit is composed by the main article plus the supplementary materials of the publication.Soil contamination is a major hindrance for plant growth and development. The lack of effective strategies to remove chemicals released into the environment has raised the need to increase plant resilience to soil pollutants. Here, we investigated the ability of two Saccharomyces cerevisiae plasma-membrane transporters, the Major Facilitator Superfamily (MFS) member Tpo1p and the ATP-Binding Cassette (ABC) protein Pdr5p, to confer Multiple Drug Resistance (MDR) in Arabidopsis thaliana. Transgenic plants expressing either of the yeast transporters were undistinguishable from the wild type under control conditions, but displayed tolerance when challenged with the herbicides 2,4-D and barban. Plants expressing ScTPO1 were also more resistant to the herbicides alachlor and metolachlor as well as to the fungicide mancozeb and the Co(2+), Cu(2+), Ni(2+), Al(3+) and Cd(2+) cations, while ScPDR5-expressing plants exhibited tolerance to cycloheximide. Yeast mutants lacking Tpo1p or Pdr5p showed increased sensitivity to most of the agents tested in plants. Our results demonstrate that the S. cerevisiae Tpo1p and Pdr5p transporters are able to mediate resistance to a broad range of compounds of agricultural interest in yeast as well as in Arabidopsis, underscoring their potential in future biotechnological applications.Fundação para a Ciência e a Tecnologia grants: (EXPL/AGR-PRO/1013/2013, PTDC/BIA-PLA/1084/2014, SFRH/BPD/44640/2008, SFRH/BPD/81221/2011, PD/BD/105735/2014, PD/00133/2012, SFRH/BD/92552/2013, UID/BIO/04565/2013, UID/Multi/04551/2013). Programa Operacional Regional de Lisboa 2020 grant: (Project N. 007317).info:eu-repo/semantics/publishedVersio

A serum substitute for fed-batch culturing of hybridoma cells

We developed a substitute for serum to produce fed-batch cultures of hybridoma cells in serum-free medium and confirmed that the cells could be successfully cultivated this way. Our substitute consisted of 12 components. The specific production rates of lactate and ammonia, which are harmful byproducts from the cells, were significantly reduced compared with a conventional serum-containing batch culture. This reduction led to a higher cell concentration and a longer production lifetime. As a result, the final concentration of monoclonal antibody was 400 mg/L, or five times greater than that in the conventional serum-containing batch culture. The developed substitute is expected to enable fed-batch cultivation in a serum-free condition