Efficient TALEN-mediated gene targeting of chicken primordial germ cells.
In this work we use TALE nucleases (TALENs) to target a reporter construct to theDDX4(vasa) locus in chicken primordial germ cells (PGCs). Vasa is a key germ cell determinant in many animal species and is posited to control avian germ cell formation. We show that TALENs mediate homology-directed repair of theDDX4locus on the Z sex chromosome at high (8.1%) efficiencies. Large genetic deletions of 30 kb encompassing the entireDDX4locus were also created using a single TALEN pair. The targeted PGCs were germline competent and were used to produceDDX4null offspring. InDDX4knockout chickens, PGCs are initially formed but are lost during meiosis in the developing ovary, leading to adult female sterility. TALEN-mediated gene targeting in avian PGCs is therefore an efficient process.
The Roslin Institute and Royal Dick School of Veterinary Studies, University of Edinburgh, Easter Bush Campus, Midlothian EH25 9RG, UK.
Lorna
L
Taylor
Recombinetics Inc, 1246 University Avenue West, Suite 300, Saint Paul, MN 55104, USA.
Daniel F
DF
Carlson
The Roslin Institute and Royal Dick School of Veterinary Studies, University of Edinburgh, Easter Bush Campus, Midlothian EH25 9RG, UK.
Sunil
S
Nandi
The Roslin Institute and Royal Dick School of Veterinary Studies, University of Edinburgh, Easter Bush Campus, Midlothian EH25 9RG, UK.
Adrian
A
Sherman
Recombinetics Inc, 1246 University Avenue West, Suite 300, Saint Paul, MN 55104, USA.
Scott C
SC
Fahrenkrug
The Roslin Institute and Royal Dick School of Veterinary Studies, University of Edinburgh, Easter Bush Campus, Midlothian EH25 9RG, UK mike.mcgrew@roslin.ed.ac.uk.
Michael J
0000-0001-8213-4632
MJ
McGrew
D000483:Alleles; D000818:Animals; D030801:Animals, Genetically Modified; D002645:Chickens / Q000235:genetics; D003433:Crosses, Genetic; D053487:DEAD-box RNA Helicases / Q000378:metabolism*; D005260:Female; D017353:Gene Deletion; D055786:Gene Knockout Techniques; D018390:Gene Targeting; D005854:Germ Cells / Q000166:cytology*; D049452:Green Fluorescent Proteins / Q000378:metabolism; D008297:Male; D008540:Meiosis; D000069896:Transcription Activator-Like Effector Nucleases; D019076:Transgenes
D049452:Green Fluorescent Proteins; D000069896:Transcription Activator-Like Effector Nucleases; D053487:DEAD-box RNA Helicases
Avian; Chicken; DDX4; Primordial germ cell; TALEN; Transgenic knockout