bioRxiv Science⌕ Search
Home/Archive/bioRxiv

bioRxiv · 10.1101/2022.09.20.508560

Development of an Agrobacterium-delivered codon-optimized CRISPR/Cas9 system for chickpea genome editing

GUPTA, S. K.·Vishwakarma, N. K.·Malakar, P.·Vanspati, P.·Sharma, N. K.·Chattopadhyay, D.
View original source ↗Export metadata ↓

Abstract

Chickpea is considered recalcitrant to in vitro tissue culture. The Clustered, Regularly Interspaced Short Palindromic Repeats/CRISPR-associated protein 9 (CRISPR/Cas9) based genome editing in chickpea can remove the bottleneck of limited genetic variation in this cash crop rich in nutrients and protein. However, the generation of stable mutant lines using CRISPR/Cas9 requires efficient and highly reproducible transformation approaches. We modified a binary vector pPZP200 by introducing a codon-optimized Cas9 gene for chickpea and the promoters of Medicago truncatula U6 snRNA for expressing guide RNA targeted to the Phytoene Desaturase (PDS) gene. The dissected single cotyledons with half embryo of chickpea were used as explants for genetic transformation. A single gRNA was found sufficient to achieve high efficiency (42%) editing with the generation of PDS mutants with albino phenotypes. A simple, rapid, highly reproducible, stable transformation and CRISPR/Cas9-based genome editing system for chickpea was established. For the first time, this study aimed to demonstrate this systems applicability by performing a gene knockout of the chickpea phytoene desaturase gene (CaPDS) in stable shoots using an improved chickpea transformation protocol.

Source connections

Explore related subjects

plant biologyPreprint

Keep this discovery

Explore connections, maps & timelines

BibTeXRIS

GUPTA, S. K., Vishwakarma, N. K., Malakar, P., Vanspati, P., Sharma, N. K., Chattopadhyay, D.. 2022-09-20. Development of an Agrobacterium-delivered codon-optimized CRISPR/Cas9 system for chickpea genome editing. https://doi.org/10.1101/2022.09.20.508560

Cite the original work for its findings. Save a collection to share your selection of sources.

KEEP EXPLORING

Related preprints

bioRxiv2026

Growth, yield, and quality of hot pepper (Capsicum annuum L.) as affected by the combined application of liquid bio-slurry and NPS fertilizer rates in Kucha, Gamo zone, south Ethiopia

Hot pepper (Capsicum annuum L.) is a key cash and nutritional crop in Ethiopia, yet its productivity remains constrained by inadequate and imbalanced soil nutrient management. Therefore, the field experiment was conducted in Kucha woreda, Gamo Zone, South Ethiopia, during the 2024/25 cropping season to investigate the effect of liquid bio-slurry and NPS fertilizer rates on growth, yield and quality of hot pepper. The experiment consisted of 16 treatments arranged in a factorial combination of four liquid bio-slurry (LBS) rates (0, 20, 40, and 60 m3 ha-1) and four NPS fertilizer rates (0, 75, 150, and 225 kg ha-1) using a randomized complete block design with three replications. All experimental data were collected following standard agronomic procedures and subjected to analysis of variance using GenStat statistical software (18th edition). The results revealed that both the main and interaction effects of liquid bio-slurry and NPS significantly (p < 0.01) influenced phenological, growth, yield, and quality parameters. The combined application of 150 kg ha-1 NPS and 60 m3 ha-1 LBS produced the highest marketable dry fruit yield (2.94 t ha-1) and total dry fruit yield (3.16 t ha-1), representing a 129.69% yield advantage over the control (1.28 t ha-1) in the case of marketable dry fruit yield. The findings demonstrate that integrated nutrient management using combined LBS and NPS fertilizers significantly enhanced the productivity and quality of hot pepper. Therefore, the application of 150 kg ha-1 NPS combined with 60 m3 ha-1 liquid bio-slurry is recommended for improved hot pepper production for the study area and similar agro-ecological conditions.

plant biology↗
bioRxiv2026

Higher stomatal density and smaller stomatal size associate with higher grain yield in winter wheat

In wheat (Triticum aestivum L.), flag leaf is crucial in determining grain-filling rate and final grain yield. Stomata, microscopic pores on plant surfaces, balance CO2 uptake and evaporative water loss; their numbers and aperture sizes depend on environmental factors. Previous studies have reported associations between stomatal traits and grain yield; mostly under controlled conditions. Our study aimed to investigate flag leaf stomatal anatomical and physiological traits, and their relationships with winter wheat grain yield under field conditions. We measured stomatal density and size, and stomatal conductance on adaxial and abaxial flag leaf surfaces in a three-year field study with twenty-five winter wheat varieties. Stomatal density ratio remained constant despite different stomatal densities occurring in different years, indicating coordinated adjustments of stomatal numbers between leaf surfaces in response to environmental conditions. Stomatal conductance ratios varied widely, indicating leaf-surface specific stomatal responses to environmental conditions. Grain yield was positively associated with stomatal density and negatively with stomatal size. Flag leaf removal reduced grain weight per spike and did not affect grain numbers per spike, unlike in spring wheat. Our field study provides valuable insights into the relationships between winter wheat stomatal traits and grain yield that can be used in breeding.

plant biology↗
bioRxiv2026

Robust planar morphogenesis in leaf-like organs requires callose-dependent coordination of division and growth

Planar organs, such as leaves, are key developmental innovations underlying the evolutionary success of plants. In vascular plants, building a planar organ requires coordinating cell division and growth both within the organ plane and across multiple, mechanically distinct cell layers. Phyllids, the leaf-like organs of bryophytes, typically consist of a single cell layer and offer a minimal system to decouple in-plane growth coordination from inter-layer mechanics. Here, we investigate how robust planar morphogenesis is achieved in phyllids, using the moss Physcomitrium patens as a bryophyte model. We show that impaired cytokinesis, caused by the loss of a callose synthase gene, disrupts growth gradients, ultimately yielding smaller, non-planar organs of variable shape. Combining quantitative live imaging with computational modeling, we show that the severity of shape defects in mutants depends on the spatial distribution and developmental timing of local growth arrests. These findings reveal how callose-dependent coordination between cell division and in-plane growth prevents mechanical conflicts and maintains planarity in a monolayered plant organ.

plant biology↗