Unexpected insertion of carrier DNA sequences into the fission yeast genome during CRISPR–Cas9 mediated gene deletion

Sophie Longmuir; Nabihah Akhtar; Stuart Andrew MacNeill

doi:10.1186/s13104-019-4228-x

Unexpected insertion of carrier DNA sequences into the fission yeast genome during CRISPR–Cas9 mediated gene deletion

Sophie Longmuir, Nabihah Akhtar, Stuart Andrew MacNeill

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9 Citations (Scopus)

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Abstract

Objectives: The fission yeast Schizosaccharomyces pombe is predicted to encode ~ 200 proteins of < 100 amino acids, including a number of previously uncharacterised proteins that are found conserved in related Schizosaccharomyces species only. To begin an investigation of the function of four of these so-called microproteins (designated Smp1–Smp4), CRISPR–Cas9 genome editing technology was used to delete the corresponding genes in haploid fission yeast cells.

Results: None of the four microprotein-encoding genes was essential for viability, meiosis or sporulation, and the deletion cells were no more sensitive to a range of cell stressors than wild-type, leaving the function of the proteins unresolved. During CRISPR–Cas9 editing however, a number of strains were isolated in which additional sequences were inserted into the target loci at the Cas9 cut site. Sequencing of the inserts revealed these to be derived from the chum salmon Oncorhynchus keta, the source of the carrier DNA used in the S. pombe transformation.

Original language	English
Article number	191
Number of pages	5
Journal	BMC Research Notes
Volume	12
DOIs	https://doi.org/10.1186/s13104-019-4228-x
Publication status	Published - 29 Mar 2019

Keywords

Microprotein
Fission yeast
Schizosaccharomyces pombe
Oncorhynchus keta
CRISPR–Cas9

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10.1186/s13104-019-4228-xLicence: CC BY

Longmuir et al. (2019)
© The Author(s) 2019. This article is distributed under the terms of the Creative Commons Attribution 4.0 International License (http://creativecommons.org/licen ses/by/4.0/), which permits unrestricted use, distribution, and reproduction in any medium, provided you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The Creative Commons Public Domain Dedication waiver (http://creativecommons.org/publicdomain/zero/1.0/) applies to the data made available in this article, unless otherwise stated.
Final published version, 1.04 MBLicence: CC BY

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title = "Unexpected insertion of carrier DNA sequences into the fission yeast genome during CRISPR–Cas9 mediated gene deletion",

abstract = "Objectives: The fission yeast Schizosaccharomyces pombe is predicted to encode ~ 200 proteins of < 100 amino acids, including a number of previously uncharacterised proteins that are found conserved in related Schizosaccharomyces species only. To begin an investigation of the function of four of these so-called microproteins (designated Smp1–Smp4), CRISPR–Cas9 genome editing technology was used to delete the corresponding genes in haploid fission yeast cells.Results: None of the four microprotein-encoding genes was essential for viability, meiosis or sporulation, and the deletion cells were no more sensitive to a range of cell stressors than wild-type, leaving the function of the proteins unresolved. During CRISPR–Cas9 editing however, a number of strains were isolated in which additional sequences were inserted into the target loci at the Cas9 cut site. Sequencing of the inserts revealed these to be derived from the chum salmon Oncorhynchus keta, the source of the carrier DNA used in the S. pombe transformation.",

keywords = "Microprotein, Fission yeast, Schizosaccharomyces pombe, Oncorhynchus keta, CRISPR–Cas9",

author = "Sophie Longmuir and Nabihah Akhtar and MacNeill, {Stuart Andrew}",

note = "Funding: This work was funded by the School of Biology, University of St Andrews, and by the Nuffield Foundation. ",

year = "2019",

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doi = "10.1186/s13104-019-4228-x",

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T1 - Unexpected insertion of carrier DNA sequences into the fission yeast genome during CRISPR–Cas9 mediated gene deletion

AU - Longmuir, Sophie

AU - Akhtar, Nabihah

AU - MacNeill, Stuart Andrew

N1 - Funding: This work was funded by the School of Biology, University of St Andrews, and by the Nuffield Foundation.

PY - 2019/3/29

Y1 - 2019/3/29

N2 - Objectives: The fission yeast Schizosaccharomyces pombe is predicted to encode ~ 200 proteins of < 100 amino acids, including a number of previously uncharacterised proteins that are found conserved in related Schizosaccharomyces species only. To begin an investigation of the function of four of these so-called microproteins (designated Smp1–Smp4), CRISPR–Cas9 genome editing technology was used to delete the corresponding genes in haploid fission yeast cells.Results: None of the four microprotein-encoding genes was essential for viability, meiosis or sporulation, and the deletion cells were no more sensitive to a range of cell stressors than wild-type, leaving the function of the proteins unresolved. During CRISPR–Cas9 editing however, a number of strains were isolated in which additional sequences were inserted into the target loci at the Cas9 cut site. Sequencing of the inserts revealed these to be derived from the chum salmon Oncorhynchus keta, the source of the carrier DNA used in the S. pombe transformation.

AB - Objectives: The fission yeast Schizosaccharomyces pombe is predicted to encode ~ 200 proteins of < 100 amino acids, including a number of previously uncharacterised proteins that are found conserved in related Schizosaccharomyces species only. To begin an investigation of the function of four of these so-called microproteins (designated Smp1–Smp4), CRISPR–Cas9 genome editing technology was used to delete the corresponding genes in haploid fission yeast cells.Results: None of the four microprotein-encoding genes was essential for viability, meiosis or sporulation, and the deletion cells were no more sensitive to a range of cell stressors than wild-type, leaving the function of the proteins unresolved. During CRISPR–Cas9 editing however, a number of strains were isolated in which additional sequences were inserted into the target loci at the Cas9 cut site. Sequencing of the inserts revealed these to be derived from the chum salmon Oncorhynchus keta, the source of the carrier DNA used in the S. pombe transformation.

KW - Microprotein

KW - Fission yeast

KW - Schizosaccharomyces pombe

KW - Oncorhynchus keta

KW - CRISPR–Cas9

U2 - 10.1186/s13104-019-4228-x

DO - 10.1186/s13104-019-4228-x

M3 - Article

SN - 1756-0500

VL - 12

JO - BMC Research Notes

JF - BMC Research Notes

M1 - 191

ER -

Unexpected insertion of carrier DNA sequences into the fission yeast genome during CRISPR–Cas9 mediated gene deletion

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