PMID- 20809332
OWN - NLM
STAT- MEDLINE
DCOM- 20101214
LR  - 20211028
IS  - 1940-6029 (Electronic)
IS  - 1064-3745 (Linking)
VI  - 659
DP  - 2010
TI  - Fluorescence in situ hybridization on early porcine embryos.
PG  - 427-36
LID - 10.1007/978-1-60761-789-1_33 [doi]
AB  - Insight into the normal and abnormal function of an interphase nucleus can be 
      revealed by using fluorescence in situ hybridization (FISH) to determine 
      chromosome copy number and/or the nuclear position of loci or chromosome 
      territories. FISH has been used extensively in studies of mouse and human early 
      embryos, however, translation of such methods to domestic species have been 
      hindered by the presence of high levels of intracytoplasmic lipid in these 
      embryos which can impede the efficiency of FISH. This chapter describes in detail 
      a FISH protocol for overcoming this problem. Following extensive technical 
      development, the protocol was derived and optimized for IVF porcine embryos to 
      enable investigation of whole chromosome and subchromosomal regions by FISH 
      during these early stages of development. Porcine embryos can be generated 
      in-vitro using semen samples from commercial companies and oocytes retrieved from 
      discarded abattoir material. According to our method, porcine embryos are lyzed 
      and immobilized on slides using Hydrochloric acid and "Tween 20" detergent, prior 
      to pretreatment with RNase A and pepsin before FISH. The method described has 
      been optimized for subsequent analysis of FISH in two dimensions since organic 
      solvents, which are necessary to remove the lipid, have the effect of flattening 
      the nuclear structure. The work in this chapter has focussed on the pig; however, 
      such methods could be applied to bovine, ovine, and canine embryos, all of which 
      are rich in lipid.
FAU - Foster, Helen A
AU  - Foster HA
AD  - Division of Biosciences, Brunel University, Uxbridge, Middlesex, UK.
FAU - Sturmey, Roger G
AU  - Sturmey RG
FAU - Stokes, Paula J
AU  - Stokes PJ
FAU - Leese, Henry J
AU  - Leese HJ
FAU - Bridger, Joanna M
AU  - Bridger JM
FAU - Griffin, Darren K
AU  - Griffin DK
LA  - eng
GR  - BB/E024211/1/BB_/Biotechnology and Biological Sciences Research Council/United 
      Kingdom
PT  - Journal Article
PL  - United States
TA  - Methods Mol Biol
JT  - Methods in molecular biology (Clifton, N.J.)
JID - 9214969
RN  - 0 (DNA Probes)
SB  - IM
MH  - Animals
MH  - DNA Probes/metabolism
MH  - Embryo, Mammalian/cytology/*metabolism
MH  - In Situ Hybridization, Fluorescence/*methods
MH  - Swine
EDAT- 2010/09/03 06:00
MHDA- 2010/12/16 06:00
CRDT- 2010/09/03 06:00
PHST- 2010/09/03 06:00 [entrez]
PHST- 2010/09/03 06:00 [pubmed]
PHST- 2010/12/16 06:00 [medline]
AID - 10.1007/978-1-60761-789-1_33 [doi]
PST - ppublish
SO  - Methods Mol Biol. 2010;659:427-36. doi: 10.1007/978-1-60761-789-1_33.