School Of Basic And Applied Sciences
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Item Silencing of a mannitol transport gene in Phelipanche aegyptiaca by the tobacco rattle virus system reduces the parasite germination on the host root(Taylor and Francis Ltd., 2022-11-24T00:00:00) Bari, Vinay Kumar; Singh, Dharmendra; Nassar, Jackline Abu; Aly, RadiRoot parasitic weed Phelipanche aegyptiaca is an obligate plant parasite that causes severe damage to host crops. Agriculture crops mainly belong to the Brassicaceae, Leguminosae, Cruciferae, and Solanaceae plant families affected by this parasitic weed, leading to the devastating loss of crop yield and economic growth. This root-specific parasitic plant is not able to complete its life cycle without a suitable host and is dependent on the host plant for nutrient uptake and germination. Therefore, selected parasitic genes of P. aegyptiaca which were known to be upregulated upon interaction with the host were chosen. These genes are essential for parasitism, and reduced activity of these genes could affect host-parasitic interaction and provide resistance to the host against these parasitic weeds. To check and examine the role of these parasitic genes which can affect the development of host resistance, we silenced selected genes in the P. aegyptiaca using the tobacco rattle virus (TRV) based virus-induced gene silencing (VIGS) method. Our results demonstrated that the total number of P. aegyptiaca parasite tubercles attached to the root of the host plant Nicotiana benthamiana was substantially decreased in all the silenced plants. However, silencing of the P. aegyptiaca MNT1 gene which encodes the mannitol transporter showed a significantly reduced number of germinated shoots and tubercles. Thus, our study indicates that the mannitol transport gene of P. aegyptiaca plays a crucial role in parasitic germination, and silencing of the PaMNT1 gene abolishes the germination of parasites on the host roots. � 2022 The Author(s). Published with license by Taylor & Francis Group, LLC.Item CRISPR/Cas9 mediated mutagenesis of MORE AXILLARY GROWTH 1 in tomato confers�resistance to root�parasitic weed Phelipanche aegyptiaca(Nature Research, 2021-02-17T00:00:00) Bari, Vinay Kumar; Nassar, Jackline Abu; Aly, RadiRoot parasitic weeds infect numerous economically important crops, affecting total yield quantity and quality. A lack of an efficient control method limits our ability to manage newly developing and more virulent races of root parasitic weeds. To control the parasite induced damage in most host crops, an innovative biotechnological approach is urgently required. Strigolactones (SLs) are plant hormones derived from carotenoids via a pathway involving the Carotenoid Cleavage Dioxygenase (CCD) 7, CCD8 and More Axillary Growth 1 (MAX1) genes. SLs act as branching inhibitory hormones and strictly required for the germination of root parasitic weeds. Here, we demonstrate that CRISPR/Cas9-mediated�targted editing of SL biosynthetic gene MAX1, in�tomato confers�resistance against root parasitic weed Phelipanche aegyptiaca. We designed sgRNA to target the third exon of MAX1 in tomato plants using the CRISPR/Cas9 system. The T0 plants were edited very efficiently at the MAX1 target site without any non-specific off-target effects. Genotype analysis of T1 plants revealed that the introduced mutations were stably passed on to the next generation. Notably, MAX1-Cas9 heterozygous and homozygous T1 plants had similar morphological changes that include excessive growth of axillary bud, reduced plant height and adventitious root formation relative to wild type. Our results demonstrated that, MAX1-Cas9 mutant lines exhibit resistance against root parasitic weed P. aegyptiaca due to reduced SL (orobanchol) level. Moreover, the expression of carotenoid biosynthetic pathway gene PDS1 and total carotenoid level was altered, as compared to wild type plants. Taking into consideration, the impact of root parasitic weeds on the agricultural economy and the obstacle to prevent and eradicate them, the current study provides new aspects into the development of an efficient control method that could be used to avoid germination of root parasitic weeds. � 2021, The Author(s).Item Targeted mutagenesis of two homologous ATP-binding cassette subfamily G (ABCG) genes in tomato confers resistance to parasitic weed Phelipanche aegyptiaca(Springer Japan, 2021-03-11T00:00:00) Bari, Vinay Kumar; Nassar, Jackline Abu; Meir, Ayala; Aly, RadiPhelipanche aegyptiaca and Orobanche spp. are obligate plant root-parasitic weeds that cause extensive damage in agricultural crop plants. Their germination requires exposure to strigolactones (SLs) exuded by the host plant roots. Here we studied genes in the host plant tomato involved in SL exudation and their impact on parasitic weeds. We provide evidence that CRISPR/Cas9-mediated targeted mutagenesis of two homologous ATP-binding cassette subfamily G (ABCG) genes, ABCG44 (Solyc08g067610) and ABCG45 (Solyc08g067620), in tomato significantly reduces SLs in the root exudate and abolishes germination of the root-parasitic weed P. aegyptiaca. Based on genome sequence similarity between ABCG44 and ABCG45, a 20-bp target sequence in their exon region was selected to design single guide RNA targeting both genes using CRISPR/Cas9. The plant binary vector constructs harboring the specific Cas9 and single guide RNA were transformed into tomato. Selected T0 mutated tomato plants showed different types of deletions at both gene loci. Genotype analysis of T1 plants suggested stable inheritance of the introduced mutations without any potential off-target effects. The phenotype of Cas9-mutated plants included increased shoot branching and growth of axillary buds, and reduced length of primary stems. Interestingly, reduced germination of P. aegyptiaca resulted from a decrease in the SL orobanchol in the root exudate of Cas9-mutated plants; however, orobanchol content in the root extract was unchanged compared to control plants. Moreover, in single and double ABCG mutants, expression of the SL-biosynthesis genes CCD8 and MAX1 decreased. The current study offers insights into CRISPR-mediated mutagenesis of ABCG genes, which could serve as an efficient control method to prevent root-parasitic weed germination. � 2021, The Botanical Society of Japan.Item Using biotechnological approaches to develop crop resistance to root parasitic weeds(Springer Science and Business Media Deutschland GmbH, 2021-04-12T00:00:00) Aly, Radi; Matzrafi, Maor; Bari, Vinay KumarMain conclusion: New transgenic and biotechnological approaches may serve as a key component in achieving crop resistance to root parasitic weeds. Abstract: Root parasitic weeds inflict severe damage to numerous crops, reducing yield quantity and quality. A lack of new sources of resistance limits our ability to manage newly developing, more virulent races. Having no effective means to control the parasites in most crops, innovative biotechnological solutions are needed. Several novel biotechnological strategies using regulatory RNA molecules, the CRISPR/Cas9 system, and T-DNA insertions have been acknowledged for engineering resistance against parasitic weeds. Significant breakthroughs have been made over the years in deciphering the plant genome and its functions, including the genomes of parasitic weeds. However, the basis of biotechnological strategies to generate host resistance to root parasitic weeds needs to be further developed. Gene-silencing and editing tools should be used to target key processes of host�parasite interactions, such as strigolactone biosynthesis and signaling, haustorium development, and degradation and penetration of the host cell wall. In this review, we summarize and discuss the main areas of research leading to the discovery and functional analysis of genes involved in host-induced gene silencing that target key parasite genes, transgenic host modification, and host gene editing to generate sustainable resistance to root parasitic weeds. � 2021, The Author(s), under exclusive licence to Springer-Verlag GmbH Germany, part of Springer Nature.