SSNIP-seq: A simple and rapid method for isolation of single-sperm nucleic acid for high-throughput sequencing

Stevan Novakovic; Vanessa Tsui; Tim Semple; Luciano Martelotto; Davis J McCarthy; Wayne Crismani

doi:10.1371/journal.pone.0275168

SSNIP-seq: A simple and rapid method for isolation of single-sperm nucleic acid for high-throughput sequencing

PLoS One. 2022 Sep 29;17(9):e0275168. doi: 10.1371/journal.pone.0275168. eCollection 2022.

Authors

Stevan Novakovic¹, Vanessa Tsui^{1

2}, Tim Semple³, Luciano Martelotto³, Davis J McCarthy^{4

5}, Wayne Crismani^{1

2}

Affiliations

¹ DNA Repair and Recombination Laboratory, St Vincent's Institute of Medical Research, Melbourne, Victoria, Australia.
² The Faculty of Medicine, Dentistry and Health Science, The University of Melbourne, Melbourne, Victoria, Australia.
³ Single Cell Innovation Laboratory, Centre for Cancer Research, The University of Melbourne, Melbourne, Victoria, Australia.
⁴ Bioinformatics and Cellular Genomics, St. Vincent's Institute of Medical Research, Melbourne, Victoria, Australia.
⁵ Melbourne Integrative Genomics, Faculty of Science, The University of Melbourne, Melbourne, Victoria, Australia.

Abstract

We developed a simple and reliable method for the isolation of haploid nuclei from fresh and frozen testes. The described protocol uses readily available reagents in combination with flow cytometry to separate haploid and diploid nuclei. The protocol can be completed within 1 hour and the resulting individual haploid nuclei have intact morphology. The isolated nuclei are suitable for library preparation for high-throughput DNA and RNA sequencing using bulk or single nuclei. The protocol was optimised with mouse testes and we anticipate that it can be applied for the isolation of mature sperm from other mammals including humans.

Publication types

Research Support, Non-U.S. Gov't

MeSH terms

Animals
Gene Library
High-Throughput Nucleotide Sequencing
Humans
Male
Mammals
Mice
Nucleic Acids*
Semen
Spermatozoa

Substances

Nucleic Acids

Grants and funding

WC and DJM received funding related to this work from the Australian National Health and Medical Research Council (GNT1129757, GNT1185387). WC is a fellow of the Victorian Cancer Agency (MCRF21006).