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

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...

Descripción completa

Detalles Bibliográficos
Autores principales: Novakovic, Stevan, Tsui, Vanessa, Semple, Tim, Martelotto, Luciano, McCarthy, Davis J., Crismani, Wayne
Formato: Online Artículo Texto
Lenguaje:English
Publicado: Public Library of Science 2022
Materias:
Lab Protocol
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC9521801/
https://www.ncbi.nlm.nih.gov/pubmed/36173986
http://dx.doi.org/10.1371/journal.pone.0275168
_version_ 1784799921203838976
author Novakovic, Stevan
Tsui, Vanessa
Semple, Tim
Martelotto, Luciano
McCarthy, Davis J.
Crismani, Wayne
author_facet Novakovic, Stevan
Tsui, Vanessa
Semple, Tim
Martelotto, Luciano
McCarthy, Davis J.
Crismani, Wayne
author_sort Novakovic, Stevan
collection PubMed
description 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.
format Online
Article
Text
id pubmed-9521801
institution National Center for Biotechnology Information
language English
publishDate 2022
publisher Public Library of Science
record_format MEDLINE/PubMed
spelling pubmed-95218012022-09-30 SSNIP-seq: A simple and rapid method for isolation of single-sperm nucleic acid for high-throughput sequencing Novakovic, Stevan Tsui, Vanessa Semple, Tim Martelotto, Luciano McCarthy, Davis J. Crismani, Wayne PLoS One Lab Protocol 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. Public Library of Science 2022-09-29 /pmc/articles/PMC9521801/ /pubmed/36173986 http://dx.doi.org/10.1371/journal.pone.0275168 Text en © 2022 Novakovic et al https://creativecommons.org/licenses/by/4.0/This is an open access article distributed under the terms of the Creative Commons Attribution License (https://creativecommons.org/licenses/by/4.0/) , which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.
spellingShingle Lab Protocol
Novakovic, Stevan
Tsui, Vanessa
Semple, Tim
Martelotto, Luciano
McCarthy, Davis J.
Crismani, Wayne
SSNIP-seq: A simple and rapid method for isolation of single-sperm nucleic acid for high-throughput sequencing
title SSNIP-seq: A simple and rapid method for isolation of single-sperm nucleic acid for high-throughput sequencing
title_full SSNIP-seq: A simple and rapid method for isolation of single-sperm nucleic acid for high-throughput sequencing
title_fullStr SSNIP-seq: A simple and rapid method for isolation of single-sperm nucleic acid for high-throughput sequencing
title_full_unstemmed SSNIP-seq: A simple and rapid method for isolation of single-sperm nucleic acid for high-throughput sequencing
title_short SSNIP-seq: A simple and rapid method for isolation of single-sperm nucleic acid for high-throughput sequencing
title_sort ssnip-seq: a simple and rapid method for isolation of single-sperm nucleic acid for high-throughput sequencing
topic Lab Protocol
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC9521801/
https://www.ncbi.nlm.nih.gov/pubmed/36173986
http://dx.doi.org/10.1371/journal.pone.0275168
work_keys_str_mv AT novakovicstevan ssnipseqasimpleandrapidmethodforisolationofsinglespermnucleicacidforhighthroughputsequencing
AT tsuivanessa ssnipseqasimpleandrapidmethodforisolationofsinglespermnucleicacidforhighthroughputsequencing
AT sempletim ssnipseqasimpleandrapidmethodforisolationofsinglespermnucleicacidforhighthroughputsequencing
AT martelottoluciano ssnipseqasimpleandrapidmethodforisolationofsinglespermnucleicacidforhighthroughputsequencing
AT mccarthydavisj ssnipseqasimpleandrapidmethodforisolationofsinglespermnucleicacidforhighthroughputsequencing
AT crismaniwayne ssnipseqasimpleandrapidmethodforisolationofsinglespermnucleicacidforhighthroughputsequencing

Ejemplares similares