Abstract
The gene editing using the CRISPR/Cas9 system has become an important biotechnological tool for studying gene function and improving crops. In this study, we have used CRISPR/Cas9 system for editing the phytoene desaturase gene (PDS) in popular Indian potato cultivar Kufri Chipsona-I. A construct (pHSE401) carrying two target gRNAs with glycine tRNA processing system under the control of Arabidopsis U6 promoter and the Cas9 protein was constructed and transformed in potato plants using Agrobacterium-mediated genetic transformations. The regeneration efficiency of 45% was observed in regenerated plants, out of which 81% of the putative transformants shoot lines exhibited mutant or bleached phenotype (albinism). The deletion mutations were detected within the StPDS gene in the genotyped plants and a mutation efficiency of 72% for gRNA1 and gRNA2 has been detected using Sanger sequencing. Hence, we set up a CRISPR/Cas9-mediated genome editing protocol which is efficient and generates mutations (deletions) within StPDS gene in potato. The bleached phenotype is easily detectable after only few weeks after Agrobacterium-mediated transformation. This is the first report as a proof of concept for CRISPR/Cas9-based editing of PDS gene in Indian potato cv. Kufri Chipsona-I. This study demonstrates that CRISPR/Cas9 can be used to edit genes at high frequency within the genome of the potato for various traits. Therefore, this study will aid in creating important mutants for modifying molecular mechanisms controlling traits of agronomic importance.
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Authors are very thankful to Indian Council of Agricultural Research-National Agricultural Science Fund (Grant Number: NASF/GT-7024/2018-19) for providing the funds for current research. We are also very thankful to, ICAR-Central Potato Research Institute, Shimla, India for providing the facilities to conduct this research work.
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Concept and design: SS, SaS, VB, RM, KS. Experimental work: SS, NSh, NSa, KT, SP, BS, HS. Supervision: SS, VB, AKT. Writing—original draft: NSh, BS. Writing—review and editing: SS, VB, VM, RT. Resource: VB, VK. All authors have read and approved the manuscript.
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Siddappa, S., Sharma, N., Salaria, N. et al. CRISPR/Cas9-mediated editing of phytoene desaturase (PDS) gene in an important staple crop, potato. 3 Biotech 13, 129 (2023). https://doi.org/10.1007/s13205-023-03543-w
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DOI: https://doi.org/10.1007/s13205-023-03543-w