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Identification of Chromosomes and Chromosome Rearrangements in Crop Brassicas and Raphanus sativus: A Cytogenetic Toolkit Using Synthesized Massive Oligonucleotide Libraries

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journal contribution
posted on 13.10.2021, 09:27 by Neha Agrawal, Mehak Gupta, Surinder S Banga, JS Pat Heslop-Harrison
Crop brassicas include three diploid [Brassica rapa (AA; 2n = 2x = 16), B. nigra (BB; 2n = 2x = 18), and B. oleracea (CC; 2n = 2x = 20)] and three derived allotetraploid species. It is difficult to distinguish Brassica chromosomes as they are small and morphologically similar. We aimed to develop a genome-sequence based cytogenetic toolkit for reproducible identification of Brassica chromosomes and their structural variations. A bioinformatic pipeline was used to extract repeat-free sequences from the whole genome assembly of B. rapa. Identified sequences were subsequently used to develop four c. 47-mer oligonucleotide libraries comprising 27,100, 11,084, 9,291, and 16,312 oligonucleotides. We selected these oligonucleotides after removing repeats from 18 identified sites (500–1,000 kb) with 1,997–5,420 oligonucleotides localized at each site in B. rapa. For one set of probes, a new method for amplification or immortalization of the library is described. oligonucleotide probes produced specific and reproducible in situ hybridization patterns for all chromosomes belonging to A, B, C, and R (Raphanus sativus) genomes. The probes were able to identify structural changes between the genomes, including translocations, fusions, and deletions. Furthermore, the probes were able to identify a structural translocation between a pak choi and turnip cultivar of B. rapa. Overall, the comparative chromosomal mapping helps understand the role of chromosome structural changes during genome evolution and speciation in the family Brassicaceae. The probes can also be used to identify chromosomes in aneuploids such as addition lines used for gene mapping, and to track transfer of chromosomes in hybridization and breeding programs.

Funding

This work was supported under the Newton-Bhabha Fund United Kingdom-India Pulses and Oilseeds Research Initiative, with funding from United Kingdom’s Official Development Assistance Newton Fund awarded by United Kingdom Biotechnology and Biological Sciences Research Council (BB/R019819/1). The Indian component of the studies was conducted with financial support from Department of Biotechnology, under the project “Germplasm enhancement for crop architecture and defensive traits in Brassica juncea L. Czern. and Coss.” NA acknowledges financial support from DFID (Department for International Development), United Kingdom for the award of Commonwealth split site fellowship during one year of the studies. SB is also grateful to Indian Council of Agricultural Research for salary support from ICAR National Professor Project “Broadening the genetic base of Indian mustard (Brassica juncea) through alien introgressions and germplasm enhancement.”

History

Citation

Front. Plant Sci. 11:598039. doi: 10.3389/fpls.2020.598039

Author affiliation

Department of Genetics and Genome Biology

Version

VoR (Version of Record)

Published in

Frontiers in Plant Science

Volume

11

Publisher

Frontiers Media

issn

1664-462X

eissn

1664-462X

Copyright date

2020

Available date

13/10/2021

Spatial coverage

Switzerland

Language

English