ABSTRACT
Background: A decrease in aneuploidy rate following a prolonged co-culture of human blastocysts has been reported. As co-culture is not routinely used in assisted reproductive technology, the present study aimed to evaluate the effect of the prolonged single culture on the rate of diploid cells in human embryos with aneuploidies
Materials and Methods: In this cohort study, we used fluorescence in situ hybridization [FISH] to reanalyze surplus blastocysts undergoing preimplantation genetic diagnosis [PGD] on day 3 postfertilization. They were randomly studied on days 6 or 7 following fertilization
Results: Of the 30 analyzed blastocysts, mosaicism was observed in 26[86.6%], while 2[6.7%] were diploid, and 2[6.7%] were triploid. Of those with mosaicism, 23[88.5%] were determined to be diploid-aneuploid and 3[11.5%] were aneuploid mosaic. The total frequency of embryos with more than 50% diploid cells was 33.3% that was lower on day 7 in comparison with the related value on day 6 [P<0.05]; however, there were no differences when the embryos were classified according to maternal age, blastocyst developmental stage, total cell number on day 3, and embryo quality
Conclusion: Although mosaicism is frequently observed in blastocysts, the prolonged single culture of blastocysts does not seem to increase the rate of normal cells
ABSTRACT
Electroporation is the most common method used for the transfection of DNA. Although this method has been attributed for various cell using different buffer compositions, the effects of DNA concentration on the transfection efficiency has not yet been studied. Here the correlation between the efficiency of electroporation reaction and increments of DNA concentration has been investigated. Following this investigation, a study was set out to produce a transgenic goat which is capable of secreting recombinant human coagulation factor IX in its milk. In this experimental study, a linearized recombinant vector [pBC1] entailing human coagulation factor IX [5.7 kb] cDNA was introduced into goat fetal fibroblast cells [three sub passages] via electroporation in four separate experiments [while other variables were kept constant], beginning with 10 micro g DNA per pulse in the first experiment and increments of 10 micro g/pulse for the next three reactions. Thereafter, polymerase chain reaction [PCR]-positive cell culture plates were diluted by several factors for further analysis of the transfected wells. Subsequently, positive cells were isolated for fluorescence in situ hybridization. Logistic regression model was used for data analyzing. Significance was defined as p< 0.05. The results showed no significant difference among three first concentrations except for group 1 [10 micro g/pulse] and group 3 [30 micro g/pulse], but there was a significant difference between these groups and the fourth group [p<0.05], as this group [40 micro g/pulse] statistically showed the highest rate of transfection. As the fluorescence in situ hybridization [FISH] results indicated the transgene was integrated in a single position in PCR positive cells. Increasing amount of using the vector 40micro g/pulse efficiently increased the number of transfected cells. Besides of voltage and buffer strength which had been previously shown to play a critical role in electroporation efficiency, our results showed an increase in DNA concentration could affect an exponential surge in the electroporation efficiency