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1

Haploidy in early bovine embryos produced in vitro

100%
Lechniak D.
Journal of Applied Genetics
|
1995
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vol. 36
|
issue 4
363-371
EN
Early bovine embryos (two -to sixteen blastomers) produced in vitro were cytogenetically analysed to determine the incidence of haploidy.Follicular oocytes were matured in vitro and inseminated with sperm prepared using the swim up method.After 2-3 days of culture, chromosome slides were prepared according to the air-drying technique and stained with Giemsa.Altogether 202 embryos produced mathaphase spreads.Of these 4.5% were pure haploid embryos and 5.5% displayed a haploid cell line within mosaic embryos (n/2n and n/3n).The occurence of haploid embryos observed in this study was compared to results of other studies and a possible origin of haploidy was discussed.
2

The use of amelogenin gene polymorphism in PCR embryo sexing in bovine IVF embryos

100%
Lechniak D., Cumming I. R.
|
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vol. 38
|
issue 1
45-49
EN
The present study describes a rapid, simple method of bovine IVF embryo sexing by use of PCR technique. A pair of primers corresponding to the bovine amelogenin sequence has been used. The Rapid Cycler (Idaho Technology, USA) used in the current experiment enabled the PCR programme consisting of 55 cycles to be completed in less than 40 minutes. Therefore the total sexing procedure could be performed in less than 90 minutes. The described method succeded in case of 85% analysed embryos.
3

Noninvasive fluorescent screening of microinjected bovine embryos to predict transgene integration

100%
Rosochacki S. J., Kozikova L., Korwin-Kossakowski M., Matejczyk M., Poloszynowicz J., Duszewska A. M.
Folia Biologica
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2003
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vol. 51
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issue 1-2
97-104
EN
Most transgenic domestic animals are generated by direct microinjection of DNA fragments into zygote pronuclei. It has generally been assumed that the majority of integration events should occur prior to the first round of chromosomal DNA replication. The aim of this study was to investigate the expression of GFP in bovine preimplantation embryos by using a gfp reporter gene consisting of chicken beta-actin promoter, the CMV-IE enhancer, gfp cDNA (EGFP) (732 bp) and rabbit beta-globin polyadenylation sequences. In five experiments 302 bovine zygotes were injected while 75 served as a control. The fluorescence intensity was detected at 72 and 168 h following fertilization in bovine embryos injected with 3 ng/mu l in experiments 1-3, and injected with 5 ng/mu l in experiments 4-5. Eight embryos were considered as expressing green fluorescence protein; 2 of them were 100% fluorescent after microinjection of a higher dose of the DNA; one was 75%, two - 50%, and three 25% transgenic. The mosaicism was assumed to be at 75%. The results indicated that the fluorescence could be analyzed at any time of bovine embryo development. It was therefore concluded, that chicken -actin promoter together with the CMV-IE enhancer would confer a strong expression of the gfp reporter gene in preimplantation bovine embryos. Therefore, using GFP that could be simply detected in live bovine (transgenic) embryos would be very promising in establishing transgenic lines of domestic animals producing in their fluids human therapeutic proteins.
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