Jaindra Tripathi, PhD - International Institute of Tropical Agriculture - Dr.

Jaindra Tripathi

PhD

International Institute of Tropical Agriculture

Dr.

Nairobi | Kenya

ORCID logohttps://orcid.org/0000-0002-6366-917X

Jaindra Tripathi, PhD - International Institute of Tropical Agriculture - Dr.

Jaindra Tripathi

PhD

Introduction

Primary Affiliation: International Institute of Tropical Agriculture - Nairobi , Kenya

Experience

Apr 2001
International Institute of Tropical Agriculture
Consultant
Biosciences

Publications

6Publications

-Reads

41Profile Views

35PubMed Central Citations

Robust CRISPR/Cas9 mediated genome editing tool for banana and plantain (Musa spp.)

https://doi.org/10.1016/j.cpb.2019.100128

Current Plant Biology

Highlights

•CRISPR)/Cas9 system enables targeted and precise editing of banana and plantain genome.
•Efficient CRISPR/Cas9 based genome editing protocol developed for banana and plantain using multiple gRNAs targeting) PDS gene.
•Edited plants showed albino and variegated phenotypes indicating mutations disrupting function of PDS.
•Robust gene editing tool pave the way for genetic improvement of banana and plantain, which are major staple food crops cultivated in Africa.
Abstract
The clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPR-associated protein 9 (Cas9) system enables precision editing of the genome of many plant species. Developing robust gene editing tools in banana and plantain would pave the way for the improvement of these crops. Here, we developed efficient CRISPR/Cas9 genome editing protocol for banana and plantain using multiple gRNAs targeting phytoene desaturase (PDS) gene. CRISPR/Cas9 construct containing two gRNAs was delivered into embryogenic cell suspension cultures of banana cultivar ‘Sukali Ndiizi’ and plantain cultivar ‘Gonja Manjaya’ by Agrobacterium-mediated transformation. The regenerated genome-edited events of ‘Sukali Ndiizi’ and ‘Gonja Manjaya’ showed albino and variegated phenotypes indicating mutations at the targeted sites disrupting the function of PDS gene. The majority of events (52/77 for ‘Sukali Ndiizi’ and 16/17 for ‘Gonja Manjaya’) were albino. Sequencing of the target sites confirmed presence of indels in all the 18 events sequenced demonstrating mutation efficiency of 100 % in both cultivars. The majority of events (6/8) of ‘Gonja Manjaya’ showed indels at both the target sites of PDS gene, however only 2/10 events of ‘Sukali Ndiizi’ showed indels at both sites, with one event (S24) having a knockout of large fragment (723 bp) indicating that both gRNAs were effective. Several of the albino events of both ‘Sukali Ndiizi’ and ‘Gonja Manjaya’ showed homozygous mutations. Further sequencing of four potential off-target sites in five events showed no mutations indicating CRISPR/Cas9 based editing in banana and plantain is targeted and precise with a very low probability of off-target sites. This study could provide a methodological framework for single or multiple knockouts in banana and plantain.



View Article
November 2019

Impact Factor 1.800

3 Reads

A simple and rapid protocol for the genetic transformation of Ensete ventricosum

Matheka, J., Tripathi, J.N., Merga, I. et al. A simple and rapid protocol for the genetic transformation of Ensete ventricosum. Plant Methods 15, 130 (2019) doi:10.1186/s13007-019-0512-y

Plant Methods

Enset (Ensete ventricosum), also known as Ethiopian banana, is a food security crop for more than 20 million people in Ethiopia. As conventional breeding of enset is very challenging, genetic engineering is an alternative option to introduce important traits such as enhanced disease resistance and nutritional value. Genetic transformation and subsequent regeneration of transgenic enset has never been reported mainly due to challenges in developing transformation protocols for this tropical species. Agrobacterium-mediated transformation could be a practical tool for the genetic improvement of enset. However, the efficiency of the transformation system depends on several parameters such as plant regeneration, genotype, explant, selection agent and Agrobacterium strains. As a first step towards the development of transgenic enset, a simple and rapid plant regeneration system was developed using multiple buds as explants. Induction and proliferation of multiple buds from shoot tip explants was achieved on Murashige and Skoog (MS) medium supplemented with 5 and 10 mg/l of 6-benzylaminopurine (BAP), respectively. Shoots were regenerated from multiple buds on MS media containing 2 mg/l BAP and 0.2% activated charcoal. Based on the optimized regeneration protocol, an Agrobacterium-mediated transformation method was developed using multiple buds as explants and the binary plasmid pCAMBIA2300-GFP containing the green florescent protein (gfp) reporter gene and neomycin phosphotransferase II (nptII) selection marker gene. Transgenic plantlets were obtained within 4 months at a frequency of about 1.25%. The transgenic lines were validated by PCR analysis using primers specific to the nptII gene. To obtain uniformly transformed plantlets, chimerism was diluted by subculturing and regenerating the transgenic shoots on a selective medium containing kanamycin (150 mg/l) for fivecycles. The uniformity of the transgenic plants was confirmed by Southern blot hybridization and RT-PCR analyses on different tissues such as leaf, pseudostem and root of same transgenic plant. In the present study, we report a simple Agrobacterium-mediated transformation system for generating transgenic events of enset. To the best of our knowledge, this is the first report on the stable transformation and regeneration of transgenic events of enset. The transformation system established in this study can be used for the generation of transgenic enset with important traits such as disease resistance.

View Article
November 2019

Impact Factor 4.500

2 Reads

Molecular Basis of Disease Resistance in Banana progenitor Musa balbisiana against Xanthomonas campestris pv. musacearum

Scientific Reports

Banana Xanthomonas wilt disease, caused by Xanthomonas campestris pv. musacearum (Xcm), is a major threat to banana production in east Africa. All cultivated varieties of banana are susceptible to Xcm and only the progenitor species Musa balbisiana was found to be resistant. the molecular basis of susceptibility and resistance of banana genotypes to Xcm is currently unknown. transcriptome analysis of disease resistant genotype Musa balbisiana and highly susceptible banana cultivar pisang Awak challenged with Xcm was performed to understand the disease response. The number of differentially expressed genes (DeGs) was higher in Musa balbisiana in comparison to pisang Awak. Genes associated with response to biotic stress were up-regulated in Musa balbisiana. the DeGs were further mapped to the biotic stress pathways. our results suggested activation of both pAMp-triggered basal defense and disease resistance (R) protein-mediated defense in Musa balbisiana as early response to Xcm infection. This study reports the first comparative transcriptome profile of the susceptible and resistant genotype of banana during early infection with Xcm and provide insights on the defense mechanism in Musa balbisiana, which can be used for genetic improvement of commonly cultivated banana varieties.
Banana Xanthomonas wilt (BXW), caused by the bacterium Xanthomonas campestris pv. musacearum (Xcm), is one of the most devastating disease endangering the livelihood of millions of farmers in east Africa, which is the largest banana-producing and -consuming region in Africa1. The impacts of BXW disease are both rapid and extreme, unlike those of other diseases, which cause gradually increasing losses over years. The disease has caused estimated economic losses of about $2-8 billion over the decade and significant reductions in production have resulted in major price increases1. The disease affects all banana cultivars grown in east Africa2. Only diploid Musa balbisiana, which is a wild type banana native to Asia and one of the progenitors of modern cultivated bananas, was found to be resistant2. Resistant cultivars could play an important role in controlling the BXW disease in east Africa, where the consumption of bananas is highest in the world at 220 to 460 kg per person annually3. There is an ongoing project for developing transgenic bananas resistant to BXW using sweet pepper genes4. However, knowledge of resistance mechanism in Musa balbisiana against Xcm can be utilized for developing resistant varieties through cis-genesis using Musa genes associated with defense, or editing of genes related to susceptibility and/or negative regulation of plant immunity. Currently, there is no understanding about the molecular mechanism for disease resistance or susceptibility in response to Xcm infection. Therefore, to obtain insight into the molecular basis of disease resistance, the transcriptome-wide differential gene expression was investigated between the BXW-resistant genotype Musa balbisiana and BXW-susceptible genotype Pisang Awak in response to Xcm. Further the differentially expressed transcripts were mapped to biotic stress pathways using Mapman to identify genes associated with defense mechanism.

View Article
May 2019
1 Read

Application of genetic modification and genome editing for developing climate- smart banana

Food and energy security

Banana is a major staple food crop feeding more than 500 million people in tropical and subtropical countries. Its production is largely constrained by diseases and pests in addition to other factors such as declining soil fertility, narrow genetic diversity in germplasm, and inadequate availability of clean planting material. The impact of climate change, particularly a rise in temperature and drought, is predicted to affect production adversely due to direct effect on plant agronomy and also influence on pathogens, pests, and their interactions with host plants. There is need to develop climate- smart varieties of banana with multiple and durable resistance to combat abiotic stresses such as extreme temperature and drought, and biotic stresses such as pathogens and pests. Modern breeding tools, including genetic modification and genome editing, can be applied for the improvement of banana bypassing the natural bottlenecks of traditional breeding. Intensive efforts using genetic modification have been made to develop improved banana varieties with resistance to biotic stresses; however, these need to be coupled with tolerance to abiotic stresses. Genome editing, an emerging powerful tool, can be applied for developing sustainable solutions to adapt to climate change by resisting biotic and abiotic stresses. CRISPR/Cas9- based genome editing has been lately established for banana, paving the way for functional genomics allowing identification of genes associated with stress- tolerant traits, which could be used for the improvement of banana for adaptation to a changing climate. This article presents an overview of recent advancements and prospective on the application of genetic modification and genome editing for developing climate- smart banana.


View Article
April 2019
11 Reads

Field resistance of transgenic plantain to nematodes has potential for future African food security

Tripathi, L. et al. Field resistance of transgenic plantain to nematodes has potential for future Af

Scientific Reports

Plant parasitic nematodes impose losses of up to 70% on plantains and cooking bananas in Africa. Application of nematicides is inappropriate and resistant cultivars are unavailable. Where grown, demand for plantain is more than for other staple crops. Confined field testing demonstrated that transgenic expression of a biosafe, anti-feedant cysteine proteinase inhibitor and an anti-root invasion, non-lethal synthetic peptide confers resistance to plantain against the key nematode pests Radopholus similis and Helicotylenchus multicinctus. The best peptide transgenic line showed improved agronomic performance relative to non-transgenic controls and provided about 99% nematode resistance at harvest of the mother crop. Its yield was about 186% in comparison with the nematode challenged control non-transgenic plants based on larger bunches and diminished plant toppling in storms, due to less root damage. This is strong evidence for utilizing this resistance to support the future food security of 70 million, mainly poor Africans that depend upon plantain as a staple food.

View Article
January 2015
7 Reads

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