Tissue-specific and ubiquitous gene knockouts by TALEN electroporation provide new approaches to investigating gene function in Ciona

Nicholas Treen, Keita Yoshida, Tetsushi Sakuma, Haruka Sasaki, Narudo Kawai, Takashi Yamamoto, Yasunori Sasakura

Research output: Contribution to journalArticle

49 Citations (Scopus)

Abstract

Custom designed nucleases can simplify gene targeting experiments and have the potential to allow these techniques to be performed in a wide range of organisms. Transcriptional activator-like effector nucleases (TALENs) are starting to fulfill this potential with the advantages of low cost and fast construction times. Here, we report that TALENs are highly effective at inducing mutations in specific genomic loci in the ascidian chordate Ciona intestinalis. In Ciona there are well-established methods to introduce exogenous DNA by electroporation, and we show that this method can be used to introduce constructs that can express TALENs ubiquitously or in specific tissues. Our current protocols enable the rapid analysis of hundreds of TALEN-induced mutants. TALEN electroporations result in a high rate of mutations. These mutations can result in gene knockouts that recapitulate previously described functions of Fgf3 and Hox12. We show that TALENs can work efficiently to cause tissue-specific knockouts and demonstrate this by knocking out Hox12 in the epidermis and Fgf3 in neural tissues. We also use tissue-specific knockouts to reveal a new function of Fgf3 during ascidian larval metamorphosis.

Original languageEnglish
Pages (from-to)481-487
Number of pages7
JournalDevelopment (Cambridge)
Volume141
Issue number2
DOIs
Publication statusPublished - 2014 Jan 15

Fingerprint

Gene Knockout Techniques
Electroporation
Urochordata
Genes
Ciona intestinalis
Chordata
Mutation
Gene Targeting
Mutation Rate
Epidermis
Costs and Cost Analysis
DNA

Keywords

  • Gene targeting
  • Reverse genetics
  • TALENs

ASJC Scopus subject areas

  • Developmental Biology
  • Molecular Biology

Cite this

Tissue-specific and ubiquitous gene knockouts by TALEN electroporation provide new approaches to investigating gene function in Ciona. / Treen, Nicholas; Yoshida, Keita; Sakuma, Tetsushi; Sasaki, Haruka; Kawai, Narudo; Yamamoto, Takashi; Sasakura, Yasunori.

In: Development (Cambridge), Vol. 141, No. 2, 15.01.2014, p. 481-487.

Research output: Contribution to journalArticle

Treen, Nicholas ; Yoshida, Keita ; Sakuma, Tetsushi ; Sasaki, Haruka ; Kawai, Narudo ; Yamamoto, Takashi ; Sasakura, Yasunori. / Tissue-specific and ubiquitous gene knockouts by TALEN electroporation provide new approaches to investigating gene function in Ciona. In: Development (Cambridge). 2014 ; Vol. 141, No. 2. pp. 481-487.
@article{7cdf610776b04d6a88a4052870c31eb2,
title = "Tissue-specific and ubiquitous gene knockouts by TALEN electroporation provide new approaches to investigating gene function in Ciona",
abstract = "Custom designed nucleases can simplify gene targeting experiments and have the potential to allow these techniques to be performed in a wide range of organisms. Transcriptional activator-like effector nucleases (TALENs) are starting to fulfill this potential with the advantages of low cost and fast construction times. Here, we report that TALENs are highly effective at inducing mutations in specific genomic loci in the ascidian chordate Ciona intestinalis. In Ciona there are well-established methods to introduce exogenous DNA by electroporation, and we show that this method can be used to introduce constructs that can express TALENs ubiquitously or in specific tissues. Our current protocols enable the rapid analysis of hundreds of TALEN-induced mutants. TALEN electroporations result in a high rate of mutations. These mutations can result in gene knockouts that recapitulate previously described functions of Fgf3 and Hox12. We show that TALENs can work efficiently to cause tissue-specific knockouts and demonstrate this by knocking out Hox12 in the epidermis and Fgf3 in neural tissues. We also use tissue-specific knockouts to reveal a new function of Fgf3 during ascidian larval metamorphosis.",
keywords = "Gene targeting, Reverse genetics, TALENs",
author = "Nicholas Treen and Keita Yoshida and Tetsushi Sakuma and Haruka Sasaki and Narudo Kawai and Takashi Yamamoto and Yasunori Sasakura",
year = "2014",
month = "1",
day = "15",
doi = "10.1242/dev.099572",
language = "English",
volume = "141",
pages = "481--487",
journal = "Development (Cambridge)",
issn = "0950-1991",
publisher = "Company of Biologists Ltd",
number = "2",

}

TY - JOUR

T1 - Tissue-specific and ubiquitous gene knockouts by TALEN electroporation provide new approaches to investigating gene function in Ciona

AU - Treen, Nicholas

AU - Yoshida, Keita

AU - Sakuma, Tetsushi

AU - Sasaki, Haruka

AU - Kawai, Narudo

AU - Yamamoto, Takashi

AU - Sasakura, Yasunori

PY - 2014/1/15

Y1 - 2014/1/15

N2 - Custom designed nucleases can simplify gene targeting experiments and have the potential to allow these techniques to be performed in a wide range of organisms. Transcriptional activator-like effector nucleases (TALENs) are starting to fulfill this potential with the advantages of low cost and fast construction times. Here, we report that TALENs are highly effective at inducing mutations in specific genomic loci in the ascidian chordate Ciona intestinalis. In Ciona there are well-established methods to introduce exogenous DNA by electroporation, and we show that this method can be used to introduce constructs that can express TALENs ubiquitously or in specific tissues. Our current protocols enable the rapid analysis of hundreds of TALEN-induced mutants. TALEN electroporations result in a high rate of mutations. These mutations can result in gene knockouts that recapitulate previously described functions of Fgf3 and Hox12. We show that TALENs can work efficiently to cause tissue-specific knockouts and demonstrate this by knocking out Hox12 in the epidermis and Fgf3 in neural tissues. We also use tissue-specific knockouts to reveal a new function of Fgf3 during ascidian larval metamorphosis.

AB - Custom designed nucleases can simplify gene targeting experiments and have the potential to allow these techniques to be performed in a wide range of organisms. Transcriptional activator-like effector nucleases (TALENs) are starting to fulfill this potential with the advantages of low cost and fast construction times. Here, we report that TALENs are highly effective at inducing mutations in specific genomic loci in the ascidian chordate Ciona intestinalis. In Ciona there are well-established methods to introduce exogenous DNA by electroporation, and we show that this method can be used to introduce constructs that can express TALENs ubiquitously or in specific tissues. Our current protocols enable the rapid analysis of hundreds of TALEN-induced mutants. TALEN electroporations result in a high rate of mutations. These mutations can result in gene knockouts that recapitulate previously described functions of Fgf3 and Hox12. We show that TALENs can work efficiently to cause tissue-specific knockouts and demonstrate this by knocking out Hox12 in the epidermis and Fgf3 in neural tissues. We also use tissue-specific knockouts to reveal a new function of Fgf3 during ascidian larval metamorphosis.

KW - Gene targeting

KW - Reverse genetics

KW - TALENs

UR - http://www.scopus.com/inward/record.url?scp=84891509001&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=84891509001&partnerID=8YFLogxK

U2 - 10.1242/dev.099572

DO - 10.1242/dev.099572

M3 - Article

AN - SCOPUS:84891509001

VL - 141

SP - 481

EP - 487

JO - Development (Cambridge)

JF - Development (Cambridge)

SN - 0950-1991

IS - 2

ER -