Use of an adsorbent and antioxidants to reduce the effects of leached phenolics in in vitro plantlet regeneration of faba bean

Development of a reliable in vitro regeneration protocol is necessary to facilitate genetic transformation of faba bean. However, leaching of phenolics from the explants of most genotypes of faba bean to theculture medium causes browning, and eventually kills the explants, hindering in vitro regeneration. This study is aimed to minimize the effect of phenolics and to identify the most suitable types of explants forin vitro regeneration. We pre-treated faba bean seeds in polyvinylpyrrolidone (PVP), then cultured different types of explants on tissue culture media supplemented with an adsorbent (activated charcoal) and antioxidants (ascorbic acid, cysteine and silver nitrate). Our results showed that treating the over night soaked seed (after removing the seed coat) with PVP solution (1000 mg/l) for 1 h, followed by culturing in Murashige and Skoog medium (MS medium) with 3% (w/v) sucrose, 0.8% (w/v) agar, 2 mg/l 6-benzylaminopurine and 2 mg/l thidiazuron, supplemented with ascorbic acid (1 mg/l) or activatedcharcoal (10 g/l), greatly reduced lethal browning in explants and improved shoot regeneration. The shoots rooted on half-strength MS medium supplemented with 0.5 mg/l -naphthaleneacetic acid. Thecotyledonary node is the most suitable type of explant for regeneration. Regenerated plantlets were successfully established in pots and set seeds in the green house

Development of an efficient regeneration system for bombarded calli from immature embryos of Moroccan durum wheat varieties

ArticleOne of the biggest obstacles limiting genetic transformation of durum wheat is the lack of an efficient regeneration system for bombarded tissues. Our study aims to optimize culture conditions for regenerating bombarded calli from immature embryos of four durum wheat varieties ‘Amria’, ‘Chaoui’, ‘Isly’ and ‘Marouane’, through comparing the effects of phytohormones (IAA, zeatin and their interaction) and nitrogen amount and sources on callus induction and plant regeneration. Both tested induction media induced approximately the same rate of induced calli for all the tested varieties. However, the interaction of the induction and the regeneration media showed a highly significant effect on plantlet regeneration for all tested varieties. After bombardment, IM1/RM2 combination proved to be the favourable medium with up to 200% and 120% plantlets regenerated for ‘Chaoui’ and ‘Isly’ varieties respectively. Encouraging results obtained in this study will help to promote the research in genetic transformation and its improvement

Agrobacterium-mediated transformation of mature embryo tissues of bread wheat (Triticum aestivum L.) genotypes

Although significant progress has been made on Agrobacterium-mediated wheat transformation, current methodologies using immature embryos as recipient tissues are labor intensive, time consuming and expensive. The use of mature embryos as explants is increasingly being recognized as an optimal method for developing regenerable cell lines during wheat transformation. Therefore, we have developed an Agrobacterium-based transformation protocol using mature embryos while adjusting several factors that influence genetic transformation efficiency. In this study, we focussed on acetosyringone concentrations, genotypes and different types of mature embryos (intact or longitudinally halved-embryos or fragmented into four latitudinal pieces) used as a source of explants for the genetic transformation. A. tumefaciens strain EHA101 harboring the plasmid vector pTF101.1 carrying the barley HVA1 gene and bar-selectable marker gene were used. Mature intact-embryos and longitudinally halved-embryos yielded the highest number of putative transgenic plantlets on the selection medium. However, no plantlets were obtained from latitudinal fragmented mature embryos. ‘Amal’ and ‘Rajae’ genotypes regenerated the highest number of putative transgenic plants and 200 μM acetosyringone was found to be the optimal concentration for their transformation. A total of 47 transgenic plants were selected with 11 plantlets showing resistance to leaf painting. Molecular analysis revealed that 1% and 0.66% of T0 regenerated plantlets were successfully transformed and carried the HVA1 gene for the ‘Amal’ and ‘Rajae’ genotypes, respectively. Additional analysis shows the transgene is stably inherited in the T1 generation. Based on the results, we conclude that among the influencing factors tested, genotypes, mature embryo explant types and acetosyringone concentration contribute significantly to the success of bread wheat transformation

First draft genome assembly of the Argane tree (Argania spinosa) [version 1; peer review: 1 approved, 1 approved with reservations]

Background: The Argane tree (Argania spinosa L. Skeels) is an endemic tree of southwestern Morocco that plays an important socioeconomic and ecologic role for a dense human population in an arid zone. Several studies confirmed the importance of this species as a food and feed source and as a resource for both pharmaceutical and cosmetic compounds. Unfortunately, the argane tree ecosystem is facing significant threats from environmental changes (global warming, over-population) and over-exploitation. Limited research has been conducted, however, on argane tree genetics and genomics, which hinders its conservation and genetic improvement. Methods: Here, we present a draft genome assembly of A. spinosa. A reliable reference genome of A. spinosa was created using a hybrid de novo assembly approach combining short and long sequencing reads. Results: In total, 144 Gb Illumina HiSeq reads and 7.2 Gb PacBio reads were produced and assembled. The final draft genome comprises 75 327 scaffolds totaling 671 Mb with an N50 of 49 916 kb. The draft assembly is close to the genome size estimated by k-mers distribution and covers 89% of complete and 4.3% of partial Arabidopsis orthologous groups in BUSCO. Conclusion: The A. spinosa genome will be useful for assessing biodiversity leading to efficient conservation of this endangered endemic tree. Furthermore, the genome may enable genome-assisted cultivar breeding, and provide a better understanding of important metabolic pathways and their underlying genes for both cosmetic and pharmacological purposes.This work was supported by the Iridian Genome Foundation (MD, USA). H.G. is supported by a Grant from the NIH (MD, USA) for H3ABioNet/H3Africa (grant numbers U41HG006941 and U24 HG006941). O.B. and B.C. are Fulbright JSD (USA) grant recipients. This work also benefited from support of Midterm Research Program of INRA-Morocco through the use of its bioinformatics platform