Redkmer: An assembly-free pipeline for the identification of abundant and specific X-chromosome target sequences for X-shredding by CRISPR endonucleases
File(s)
Published version
Author(s)
Papathanos, Philippos Aris
Windbichler, Nikolai
Type
Journal Article
Abstract
CRISPR-based synthetic sex ratio distorters, which operate by shredding the X-chromosome during male meiosis, are promising tools for the area-wide control of harmful insect pest or disease vector species. X-shredders have been proposed as tools to suppress insect populations by biasing the sex ratio of the wild population toward males, thus reducing its natural reproductive potential. However, to build synthetic X-shredders based on CRISPR, the selection of gRNA targets, in the form of high-copy sequence repeats on the X chromosome of a given species, is difficult, since such repeats are not accurately resolved in genome assemblies and cannot be assigned to chromosomes with confidence. We have therefore developed the redkmer computational pipeline, designed to identify short and highly abundant sequence elements occurring uniquely on the X chromosome. Redkmer was designed to use as input minimally processed whole genome sequence data from males and females. We tested redkmer with short- and long-read whole genome sequence data of Anopheles gambiae, the major vector of human malaria, in which the X-shredding paradigm was originally developed. Redkmer established long reads as chromosomal proxies with excellent correlation to the genome assembly and used them to rank X-candidate kmers for their level of X-specificity and abundance. Among these, a high-confidence set of 25-mers was identified, many belonging to previously known X-chromosome repeats of Anopheles gambiae, including the ribosomal gene array and the selfish elements harbored within it. Data from a control strain, in which these repeats are shared with the Y chromosome, confirmed the elimination of these kmers during filtering. Finally, we show that redkmer output can be linked directly to gRNA selection and off-target prediction. In addition, the output of redkmer, including the prediction of chromosomal origin of single-molecule long reads and chromosome specific kmers, could also be used for the characterization of other biologically relevant sex chromosome sequences, a task that is frequently hampered by the repetitiveness of sex chromosome sequence content.
Date Issued
2018-02-01
Date Acceptance
2018-02-01
Citation
The Crispr Journal, 2018, 1 (1), pp.88-98
ISSN
2573-1599
Publisher
Mary Ann Liebert, Inc.
Start Page
88
End Page
98
Journal / Book Title
The Crispr Journal
Volume
1
Issue
1
Copyright Statement
© 2018 Philippos Aris Papathanos and Nikolai Windbichler; Published by Mary Ann Liebert, Inc. This article is available under the Creative Commons License CC-BY-NC (http://creativecommons.org/licenses/by-nc/4.0). This license permits non-commercial use, distribution and reproduction in any medium, provided the original work isproperly cited. Permission only needs to be obtained for commercial use and can be done via RightsLink.
Sponsor
Biotechnology and Biological Sciences Research Council (BBSRC)
Identifier
https://www.ncbi.nlm.nih.gov/pubmed/30627701
PII: 10.1089/crispr.2017.0012
Grant Number
BB/P000843/1
Publication Status
Published
Coverage Spatial
United States
Date Publish Online
2018-02-01