Type

Journal Article

Authors

Patrick Lonergan
James Roche
Emma Gallagher
Solomon Mamo
James D Murray
Ciara M O'Meara

Subjects

Biochemistry

Topics
animals cadherins embryology female rna small interfering gene transfer techniques cattle genetics metabolism administration dosage embryo culture techniques transfection gene silencing male cells cultured fertilization in vitro cytology microinjections physiology animals genetically modified zygote antagonists inhibitors embryo mammalian

Gene silencing in bovine zygotes: siRNA transfection versus microinjection. (2010)

Abstract The aim of this study was to compare gene silencing in bovine zygotes when small interfering RNAs (siRNAs) were introduced into bovine zygotes by microinjection or lipid-based transfection. In Experiment 1, E-cadherin siRNA was injected at 100 or 375 µM and compared with PBS-injected and non-injected controls. Embryos were then cultured in vitro for 7 days and periodically assessed for development. For transfection, zona-free zygotes were incubated in transfection medium with siRNA for 1h at 39°C and then cultured to Day 7. Injection of PBS or 375 µM E-cadherin siRNA resulted in a decrease in the number of embryos reaching the 8-cell stage (51.5% and 45.5%) or the blastocyst stage (39.0 and 32.5%) compared with non-injected controls (62.9 and 45.0%, respectively; P<0.05). Messenger RNA abundance was suppressed by 36 and 46% when siRNA targeting E-cadherin was injected at 100 and 375 µM, respectively, compared with controls (P<0.05). Transfection with 100 nM E-cadherin siRNA decreased development to the 8-cell stage (20.3 versus 53.0%) and blastocyst stage (7.2 versus 18.2%) compared with controls (P<0.05). Messenger RNA relative abundance was not different between controls (non-transfected or transfected with GAPDH or scrambled siRNA). However, transfection of zygotes with 100 and 200 nM E-cadherin siRNA led to a 72 and 38% reduction, respectively, in E-cadherin mRNA relative abundance in Day 7 blastocysts compared with controls (P<0.05).
Collections Ireland -> University College Dublin -> PubMed

Full list of authors on original publication

Patrick Lonergan, James Roche, Emma Gallagher, Solomon Mamo, James D Murray, Ciara M O'Meara

Experts in our system

1
P. Lonergan
University College Dublin
Total Publications: 190
 
2
J. F. Roche
University College Dublin
Total Publications: 111
 
3
Solomon Mamo
University College Dublin
Total Publications: 16