CSIRO Publishing blank image blank image blank image blank imageBooksblank image blank image blank image blank imageJournalsblank image blank image blank image blank imageAbout Usblank image blank image blank image blank imageShopping Cartblank image blank image blank image You are here: Journals > Reproduction, Fertility and Development   
Reproduction, Fertility and Development
Journal Banner
  Vertebrate Reproductive Science & Technology
blank image Search
blank image blank image
blank image
  Advanced Search

Journal Home
About the Journal
Editorial Structure
Online Early
Current Issue
Just Accepted
All Issues
Special Issues
Research Fronts
Virtual Issues
Sample Issue
For Authors
General Information
Submit Article
Author Instructions
Open Access
Awards and Prizes
For Referees
Referee Guidelines
Review an Article
Annual Referee Index
For Subscribers
Subscription Prices
Customer Service
Print Publication Dates

blue arrow e-Alerts
blank image
Subscribe to our email Early Alert or RSS feeds for the latest journal papers.

red arrow Connect with us
blank image
facebook twitter logo LinkedIn

red arrow Connect with SRB
blank image
facebook TwitterIcon

Affiliated Societies

RFD is the official journal of the International Embryo Transfer Society and the Society for Reproductive Biology.


Article << Previous     |     Next >>   Contents Vol 22(1)


M. P. M. Mancini A, B. C. S. Campanha A, D. M. Souza A, C. P. Godoi A, F. Frei A, M. F G. Nogueira A

São Paulo State University (UNESP), Assis, São Paulo, Brazil
 Export Citation


Mixing embryo cells coming from different fertilizations (i.e. embryonic chimera) have been used as a tool to understand embryogenesis, organo- genesis, and pluripotency, as well as a source to obtain transgenic mammals. The objectives of this work were to evaluate the potential of mice demi-embryos, in advanced stage of the development (morulae and blastocysts) to aggregate in chimeras; to compare the chimerism rate of those embryos with the rate of whole 8- to 16-cell-stage embryos; and to measure the genotype composition of the resultant chimera. One-month-old transgenic (C57/BL6/EGFP strain, GFP) or non-transgenic (Swiss Webster strain, SW) mice weighing approximately 35 g were superstimulated with 5 or 10IU of eCG (for GFP or SW mice, respectively) followed with hCG injection of 5 or 10IU (GFP or SW mice, respectively) 48 h later. Embryos were harvested at different stages of development and allocated in 3 groups for aggregation technique. Blastocysts and morulae were bisected (microblade mounted on TransferMan NK-2, Eppendorf), whereas 8- to 16-cell-stage embryos had their zona pellucida mechanically removed (23-gauge needle). Embryos were manipulated in M2 culture medium at room temperature, and aggregation groups consisted of G1 (2 demi-blastocysts, n = 28), G2 (demi-blastocyst and demi-morula, n = 20), and G3 (2 whole 8- to 16-cell-stage embryos, n = 25). All embryos were placed in wells (Embryo GPS dish, SunIVF) containing KSOMaa medium (EmbryoMax, Millipore) under oil (Sigma, St. Louis, MO, USA) and were incubated at 37°C, 5% CO2 in air saturated with humidity. After 24 h of incubation, the presence of chimera was verified, and the percentage of area (square pixel) occupied by each embryonic type (GFP or SW) from both G2 (n = 3) and G3 (n = 3) were measured by the ImageJ program (v. 1.42i, USA). General results of the chimerism rate were 3.6%, 15.0%, and 60.0% (G1, G2, and G3, respectively; P < 0.001, chi-square). The G3 group differed from others (G1, P < 0.001 and G2, P = 0.003), which appeared similar (P = 0.294; Fisher’s exact test). The mean percentage (±SD) of GFP cells in the resultant chimera were 51.3 ± 4.1% and 50.6 ± 10.0% (for G2 and G3, respectively; P = 0.91, t-test). Moreover, the percentages of GFP cells within the same group of G2 or G3 at 0 v. 24 h of culture were not statistically different (data not shown). It was concluded that in our conditions, the embryonic chimerism by aggregation of murine demi-embryos is a feasible procedure, even for embryos in an advanced stage of development (morulae and blastocysts). Nevertheless, the chimerism rate with whole pre-compaction embryos (G3) was higher than that of G1 and G2 groups. Furthermore, the phenotype of embryonic chimera was equally composed, with no effect of strain (GFP or SW cells) or culture (0 or 24 h) on its composition.

Legal & Privacy | Contact Us | Help


© CSIRO 1996-2015