Genome Engineering Tools for Functional Genomics and Crop Improvement in Legumes

2018 ◽  
pp. 219-234
Author(s):  
Rashi Khandelwal ◽  
Mukesh Jain
Keyword(s):  
Functional Genomics ◽  
Genome Engineering ◽  
Crop Improvement

scholarly journals Recent advances in CRISPR/Cas9 and applications for wheat functional genomics and breeding

aBIOTECH ◽  
2021 ◽  
Author(s):  
Jun Li ◽  
Yan Li ◽  
Ligeng Ma
Keyword(s):  
Functional Genomics ◽  
Genome Editing ◽  
Functional Annotation ◽  
Genome Engineering ◽  
Agronomic Traits ◽  
Wheat Breeding ◽  
Breeding Programs ◽  
Base Editing ◽  
The World ◽  
World Food Security

AbstractCommon wheat (Triticum aestivum L.) is one of the three major food crops in the world; thus, wheat breeding programs are important for world food security. Characterizing the genes that control important agronomic traits and finding new ways to alter them are necessary to improve wheat breeding. Functional genomics and breeding in polyploid wheat has been greatly accelerated by the advent of several powerful tools, especially CRISPR/Cas9 genome editing technology, which allows multiplex genome engineering. Here, we describe the development of CRISPR/Cas9, which has revolutionized the field of genome editing. In addition, we emphasize technological breakthroughs (e.g., base editing and prime editing) based on CRISPR/Cas9. We also summarize recent applications and advances in the functional annotation and breeding of wheat, and we introduce the production of CRISPR-edited DNA-free wheat. Combined with other achievements, CRISPR and CRISPR-based genome editing will speed progress in wheat biology and promote sustainable agriculture.

Delineating Calcium Signaling Machinery in Plants: Tapping the Potential through Functional Genomics

2021 ◽  
Vol 22 ◽  
Author(s):  
Soma Ghosh ◽  
Malathi Bheri ◽  
Girdhar K. Pandey
Keyword(s):  
Functional Genomics ◽  
Protein Kinases ◽  
Regulatory Networks ◽  
Crop Improvement ◽  
Functional Characterization ◽  
Genomic Analysis ◽  
Major Elements ◽  
Model Systems ◽  
Plant Responses ◽  
Dependent Protein

: Plant systems have developed calcium (Ca2+) signaling as an important mechanism of regulation of stress perception, developmental cues, and responsive gene expression. The post-genomic era has witnessed the successful unravelling of the functional characterization of genes and the creation of large datasets of molecular information. The major elements of Ca2+ signaling machinery involve Ca2+ sensors and responders such as Calmodulin (CaM), Calmodulin-like proteins (CMLs), Ca2+/CaM-dependent protein kinases (CCaMK), Ca2+-dependent protein kinases (CDPKs), Calcineurin B-like proteins (CBLs) as well as transporters, such as Cyclic nucleotide-gated channels (CNGCs), Glutamate-like receptors (GLRs), Ca2+-ATPases, Ca2+/H+ exchangers (CAXs) and mechanosensitive channels. These elements play an important role in the regulation of physiological processes and plant responses to various stresses. Detailed genomic analysis can help us in the identification of potential molecular targets that can be exploited towards the development of stress-tolerant crops. The information sourced from model systems through omics approaches helps in the prediction and simulation of regulatory networks involved in responses to different stimuli at the molecular and cellular levels. The molecular delineation of Ca2+ signaling pathways could be the stepping stone for engineering climate-resilient crop plants. Here, we review the recent developments in Ca2+ signaling in the context of transport, responses, and adaptations significant for crop improvement through functional genomics approaches.

Advances in potato functional genomics: implications for crop improvement

2022 ◽  
Author(s):  
Neha Sharma ◽  
Sundaresha Siddappa ◽  
Nikhil Malhotra ◽  
Kajal Thakur ◽  
Neha Salaria ◽  
...  
Keyword(s):  
Functional Genomics ◽  
Crop Improvement

scholarly journals Precision Genome Engineering Through Cytidine Base Editing in Rapeseed (Brassica napus. L)

2020 ◽  
Vol 2 ◽  
Author(s):  
Limin Hu ◽  
Olalekan Amoo ◽  
Qianqian Liu ◽  
Shengli Cai ◽  
Miaoshan Zhu ◽  
...  
Keyword(s):  
Genome Engineering ◽  
Molecular Breeding ◽  
Target Genes ◽  
Agronomic Traits ◽  
Crop Improvement ◽  
Brassica Napus L ◽  
Editing Efficiency ◽  
Base Editing ◽  
Expected Gain ◽  
Genome Wide

Rapeseed is one of the world's most important sources of oilseed crops. Single nucleotide substitution is the basis of most genetic variation underpinning important agronomic traits. Therefore, genome-wide and target-specific base editing will greatly facilitate precision plant molecular breeding. In this study, four CBE systems (BnPBE, BnA3A-PBE, BnA3A1-PBE, and BnPBGE14) were modified to achieve cytidine base editing at five target genes in rapeseed. The results indicated that genome editing is achievable in three CBEs systems, among which BnA3A1-PBE had the highest base-editing efficiency (average 29.8% and up to 50.5%) compared to all previous CBEs reported in rapeseed. The editing efficiency of BnA3A1-PBE is ~8.0% and fourfold higher, than those of BnA3A-PBE (averaging 27.6%) and BnPBE (averaging 6.5%), respectively. Moreover, BnA3A1-PBE and BnA3A-PBE could significantly increase the proportion of both the homozygous and biallelic genotypes, and also broaden the editing window compared to BnPBE. The cytidine substitution which occurred at the target sites of both BnaA06.RGA and BnaALS were stably inherited and conferred expected gain-of-function phenotype in the T1 generation (i.e., dwarf phenotype or herbicide resistance for weed control, respectively). Moreover, new alleles or epialleles with expected phenotype were also produced, which served as an important resource for crop improvement. Thus, the improved CBE system in the present study, BnA3A1-PBE, represents a powerful base editor for both gene function studies and molecular breeding in rapeseed.

scholarly journals A potential clue for the application of prime editing in tomato, a dicot plant

2021 ◽  
Author(s):  
Tien Van Vu ◽  
Jihae Kim ◽  
Swati Das ◽  
Jae-Yean Kim
Keyword(s):  
High Frequency ◽  
Mammalian Cells ◽  
Genome Engineering ◽  
Crop Improvement ◽  
Plant Genome ◽  
Guide Rna ◽  
Genome Modification ◽  
Potential Applications ◽  
Targeted Deep Sequencing ◽  
Synthetic Sequence

Precision genome editing is highly desired for crop improvement. The recently emerged CRISPR/Cas technology offers great potential applications in precision plant genome engineering. A prime editing (PE) approach combining a reverse transcriptase (RT) with a Cas9 nickase and a priming extended guide RNA has shown a high frequency for precise genome modification in mammalian cells and several plant species. However, the applications of the PE approach in dicot plants are still limited and inefficient. We designed and tested prime editors for precision editing of a synthetic sequence in a transient assay and for desirable alleles of 10 loci in tomato by stable transformation. However, our data obtained by targeted deep sequencing also revealed inefficient PE activity in both the tobacco and tomato systems. Further assessment of the activities of the PE components uncovered potential reasons for the inefficiency of the PE complexes. These data could also help explain the recent successes of some prime editors in plants using improved expression systems. Our work provides an important clue for the application of the PE approach in crop improvement.

RNAi: New Era of Functional Genomics for Crop Improvement

2012 ◽  
pp. 24-38 ◽  
Author(s):  
Pradeep Kumar ◽  
Madhu Kamle ◽  
Ashutosh Pandey
Keyword(s):  
Functional Genomics ◽  
Crop Improvement ◽  
New Era

scholarly journals Modern Trends in Plant Genome Editing: An Inclusive Review of the CRISPR/Cas9 Toolbox

2019 ◽  
Vol 20 (16) ◽  
pp. 4045 ◽  
Author(s):  
Ali Razzaq ◽  
Fozia Saleem ◽  
Mehak Kanwal ◽  
Ghulam Mustafa ◽  
Sumaira Yousaf ◽  
...  
Keyword(s):  
Genome Editing ◽  
Genome Engineering ◽  
Crop Improvement ◽  
Crop Plants ◽  
Targeted Mutagenesis ◽  
Plant Genome ◽  
Zinc Finger Nucleases ◽  
Base Substitution ◽  
Transcription Activator ◽  
Abiotic Stressors

Increasing agricultural productivity via modern breeding strategies is of prime interest to attain global food security. An array of biotic and abiotic stressors affect productivity as well as the quality of crop plants, and it is a primary need to develop crops with improved adaptability, high productivity, and resilience against these biotic/abiotic stressors. Conventional approaches to genetic engineering involve tedious procedures. State-of-the-art OMICS approaches reinforced with next-generation sequencing and the latest developments in genome editing tools have paved the way for targeted mutagenesis, opening new horizons for precise genome engineering. Various genome editing tools such as transcription activator-like effector nucleases (TALENs), zinc-finger nucleases (ZFNs), and meganucleases (MNs) have enabled plant scientists to manipulate desired genes in crop plants. However, these approaches are expensive and laborious involving complex procedures for successful editing. Conversely, CRISPR/Cas9 is an entrancing, easy-to-design, cost-effective, and versatile tool for precise and efficient plant genome editing. In recent years, the CRISPR/Cas9 system has emerged as a powerful tool for targeted mutagenesis, including single base substitution, multiplex gene editing, gene knockouts, and regulation of gene transcription in plants. Thus, CRISPR/Cas9-based genome editing has demonstrated great potential for crop improvement but regulation of genome-edited crops is still in its infancy. Here, we extensively reviewed the availability of CRISPR/Cas9 genome editing tools for plant biotechnologists to target desired genes and its vast applications in crop breeding research.

scholarly journals Transgene-Free Genome Editing in Tomato and Potato Plants Using Agrobacterium-Mediated Delivery of a CRISPR/Cas9 Cytidine Base Editor

2019 ◽  
Vol 20 (2) ◽  
pp. 402 ◽  
Author(s):  
Florian Veillet ◽  
Laura Perrot ◽  
Laura Chauvin ◽  
Marie-Paule Kermarrec ◽  
Anouchka Guyon-Debast ◽  
...  
Keyword(s):  
Genome Editing ◽  
First Generation ◽  
Genome Engineering ◽  
Crop Improvement ◽  
Point Mutations ◽  
Acetolactate Synthase ◽  
Agrobacterium Mediated Transformation ◽  
Dicotyledonous Plants ◽  
Function Discovery ◽  
Vegetatively Propagated Plants

Genome editing tools have rapidly been adopted by plant scientists for gene function discovery and crop improvement. The current technical challenge is to efficiently induce precise and predictable targeted point mutations valuable for crop breeding purposes. Cytidine base editors (CBEs) are CRISPR/Cas9 derived tools recently developed to direct a C-to-T base conversion. Stable genomic integration of CRISPR/Cas9 components through Agrobacterium-mediated transformation is the most widely used approach in dicotyledonous plants. However, elimination of foreign DNA may be difficult to achieve, especially in vegetatively propagated plants. In this study, we targeted the acetolactate synthase (ALS) gene in tomato and potato by a CBE using Agrobacterium-mediated transformation. We successfully and efficiently edited the targeted cytidine bases, leading to chlorsulfuron-resistant plants with precise base edition efficiency up to 71% in tomato. More importantly, we produced 12.9% and 10% edited but transgene-free plants in the first generation in tomato and potato, respectively. Such an approach is expected to decrease deleterious effects due to the random integration of transgene(s) into the host genome. Our successful approach opens up new perspectives for genome engineering by the co-edition of the ALS with other gene(s), leading to transgene-free plants harboring new traits of interest.

Application of Functional Genomics Tools for Crop Improvement

2007 ◽  
pp. 411-427 ◽  
Author(s):  
Motoyuki Ashikari ◽  
Makoto Matsuoka ◽  
Masahiro Yano
Keyword(s):  
Functional Genomics ◽  
Crop Improvement ◽  
Genomics Tools

scholarly journals Genotyping strategies for detecting CRISPR mutations in polyploid species: a case study-based approach in hexaploid wheat

2021 ◽  
Author(s):  
Ajay Gupta ◽  
Wanlong Li
Keyword(s):  
Genome Engineering ◽  
Crop Improvement ◽  
Cost Effective ◽  
Significant Advance ◽  
Polyploid Species ◽  
Small Indels ◽  
Cleavage Assay ◽  
Versatile Tool ◽  
Mismatch Cleavage

As a versatile tool for genome engineering, CRISPR-Cas9 has been revolutionizing the field of molecular biology, biotechnology, and crop improvement. By precisely targeting pre-selected genomic sites, CRISPR-Cas9 primarily induces insertions or deletions (indels) of variable size. Despite the significant advance in the technology per se, detecting these indels is the major and difficult part of the CRISPR program in polyploid species, like wheat, with relatively low mutation rates. A plethora of methods are available for detecting mutations, but no method is perfect for all mutation types. In this case study, we demonstrated a new, protocol for capturing length polymorphism from small indels using a nested PCR approach. This new method is tractable, efficient, and cost-effective in detecting and genotyping indels >3-bp. We also discussed the major genotyping platforms used in our wheat CRISPR projects, such as mismatch cleavage assay, restriction enzyme assay, ribonucleoprotein assay, and Sanger sequencing, for their advantages and pitfalls in wheat CRISPR mutation detection.

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