Molecular cloning of translocation breakpoints in a case of constitutional translocation t(11;22)(q23;q11) and preparation of probes for preimplantation genetic diagnosis

1999 ◽  
Vol 11 (1) ◽  
pp. 17 ◽  
Author(s):  
Jingly Fung ◽  
Santiago Munné ◽  
Jaqueline Garcia ◽  
Ung-Jin Kim ◽  
Heinz-Ulli G. Weier
Keyword(s):  
Preimplantation Genetic Diagnosis ◽  
Yeast Artificial Chromosome ◽  
Chromosomal Abnormalities ◽  
Genetic Diagnosis ◽  
Artificial Chromosome ◽  
Preimplantation Embryos ◽  
Cytogenetic Map ◽  
Additional Time ◽  
Interphase Cell ◽  
Translocation Breakpoints

In vitro fertilization (IVF) centres with preimplantation genetic diagnosis (PGD) programmes are often confronted with the problem of identifying chromosomal abnormalities in interphase cells biopsied from preimplantation embryos of carriers of a reciprocal translocation. The present authors have developed a DNA testing based approach to analyse embryos from translocation carriers, and this report describes breakpoint-spanning probes to detect abnormalities in cases of the most common human translocation (i.e. the t(11;22)(q23;q11)). Screening a yeast artificial chromosome (YAC) library for probes covering the respective breakpoint regions in the patient lead to probes for the breakpoint on chromosome 11q23. The physically mapped YAC and bacterial artificial chromosome (BAC) clones from chromosome 22 were then integrated with the cytogenetic map, which allowed localization of the breakpoint on chromosome 22q11 to an interval of less than 84 kb between markers D22S184 and KI457 and to prepare probes suitable for interphase cell analysis. In summary, breakpoint localization could be accomplished in about 4 weeks with additional time needed to optimize probes for use in PGD.

scholarly journals Role of Preimplantation Genetic Diagnosis (PGD) in Current Infertility Practice

2010 ◽  
Vol 1 (1) ◽  
pp. 1-10 ◽  
Author(s):  
Helen Ghislaine Tempest ◽  
Joe Leigh Simpson
Keyword(s):  
Preimplantation Genetic Diagnosis ◽  
Chromosomal Abnormalities ◽  
Polar Body ◽  
Genetic Diagnosis ◽  
Clinical Trial Data ◽  
Preimplantation Embryos ◽  
Novel Technologies ◽  
Chromosome Translocations ◽  
Chromosome Imbalances

ABSTRACT Chromosome imbalances are the leading cause of pregnancy loss in humans and play major roles in male and female infertility. Within the past two decades, the development and application of preimplantation genetic diagnosis (PGD) has played an important role in infertility practices worldwide. The purpose of this review is to discuss, how PGD may be applied in combating numerical chromosomal abnormalities and in Robertsonian and reciprocal chromosome translocations. We shall consider prevalence and risk of each aberration, interchromosomal effects and rationale behind use of PGD in each case. Numerical chromosome abnormalities (aneuploidy and polyploidy) in particular affect a very high proportion of preimplantation embryos (~ 50%). Given that a majority of preimplantation embryos are aneuploid, PGD can be used to screen embryos and transfer euploid embryos to improve pregnancy rates and reduce spontaneous abortions. The rationale of utilize PGD to transfer only euploid embryos would seem sound, but controversies exist surrounding application of PGD for aneuploidy detection. To this end, we will discuss the dichotomy between favorable descriptive reports and less favorable randomized clinical trial data. This review will discuss the trend towards differing sources of embryonic DNA (e.g. polar body vs blastomere vs blastocyst) as well as development of novel technologies for 24 chromosomes analysis.

Sample preparations are important for fluorescence in situ hybridization of cells biopsied from preimplantation embryos

Zygote ◽  
2013 ◽  
Vol 22 (3) ◽  
pp. 300-304
Author(s):  
Lifei Li ◽  
Xuehong Zhang ◽  
Weihua Wang
Keyword(s):  
Sample Preparation ◽  
Chromosomal Abnormalities ◽  
Genetic Diagnosis ◽  
Preimplantation Embryos ◽  
Human Embryos ◽  
Fixation Time ◽  
Preparation Technique ◽  
Hypotonic Solution ◽  
Modified Method

SummaryFluorescence in situ hybridization (FISH) is a cytogenetic technology used to detect chromosomal abnormalities in preimplantation human embryos. However, its efficiency is not stable due to improper sample preparation. The present study was designed to modify the current sample preparation technique and then to evaluate its efficiency in human preimplantation genetic diagnosis (PGD). Day 3 cleavage embryos as well as day 5 and 6 blastocysts were biopsied by mechanical aspiration method. In the present study, two methods were used for sample preparation of the biopsied cells. Method I was the traditional method, in which each blastomere was placed in a hypotonic solution for 5 min and then fixed on glass slides. The slides were kept at room temperature before the FISH procedures. Method II was a modified method, in which all blastomeres were placed individually in hypotonic solution drops covered by oil for at least 5 min and then fixed on slides with 0.1% Tween/HCl. After fixation, the slides were kept at –20°C for at least 30 min before the FISH procedures. The two methods were compared in terms of time consumption and proportions of blastomeres with FISH signals. In total, 329 blastomeres from day 3 embryos were fixed by Method I with an average fixation time of 8–10 min for each blastomere. By contrast, with Method II, 362 blastomeres were fixed and the average time was 3–4 min for each blastomere. After FISH, more nuclei had signals with Method II (97.2%) than with Method I (86.9%). All cells that were biopsied from blastocysts and prepared with Method II had FISH signals. However, Method I was not suitable for the fixation of multiple cells biopsied from blastocysts as cells were not traceable during the fixation. The present study indicates that proper sample preparation is critical for obtaining FISH signals in cells biopsied from preimplantation human embryos; hence these modifications can increase the efficiency of human PGD.

scholarly journals A comparison of NGS and FISH technologies: comprehensive 24-chromosome screening of embryo

1970 ◽  
Vol 21 ◽  
pp. 311-315
Author(s):  
Yu. V. Gontar ◽  
O. Yu. Verlynskyi ◽  
A. Kyrpyi ◽  
I. E. Ylyn ◽  
A. M. Fedota
Keyword(s):  
Preimplantation Genetic Diagnosis ◽  
Assisted Reproductive Technologies ◽  
Genetic Diagnosis ◽  
Reproductive Technologies ◽  
The Other ◽  
Preimplantation Embryos ◽  
Art Programs ◽  
Chromosomal Aneuploidy ◽  
Investigation Methods

Aim. Optimization of the algorithm of complex 24 chromosomes screening in programs of assisted reproductive technologies. Methods. Research of non-disjunction chromosomes in preimplantation embryos based on the results of trophectoderm nucleus diagnostics using FISH and NGS-based CCS. During the preimplantation genetic diagnosis (PGD) on the nucleus by FISH were used probes for chromosomes 13, 16, 18, 21, 22, X, Y. Results. Among the demonstrated cases of embryo diagnosis there was only one embryo that showed a coincidence in the results obtained by different investigation methods. In the other sample, where was diagnosed non-mosaic 18 and 22 monosomy by FISH, the NGS-based CCS showed only monosomy 18. The other embryo had ploidy mosaicism indicated by FISH, but according to NGS results it was evaluated as euploid. Conclusions. Embryos obtained in ART programs must be screened for chromosomal aneuploidy in the preimplantation period to increase the effectiveness in the programs of assisted reproductive technologies, using combination of FISH and NGS methods. Keywords: preimplantation genetic diagnosis, assisted reproductive technologies, aneuploid embryos, NGS, FISH.

Clinical outcome of treatment cycles using preimplantation genetic diagnosis for structural chromosomal abnormalities

2001 ◽  
Vol 21 (9) ◽  
pp. 781-787 ◽  
Author(s):  
Margareta Fridström ◽  
Lars Ährlund-Richter ◽  
Erik Iwarsson ◽  
Helena Malmgren ◽  
José Inzunza ◽  
...  
Keyword(s):  
Clinical Outcome ◽  
Preimplantation Genetic Diagnosis ◽  
Chromosomal Abnormalities ◽  
Genetic Diagnosis

Preimplantation genetic diagnosis

2002 ◽  
Vol 10 (1) ◽  
pp. 3-20
Author(s):  
JDA Delhanty ◽  
JC Harper
Keyword(s):  
Preimplantation Genetic Diagnosis ◽  
Chromosomal Abnormalities ◽  
Recurrent Miscarriage ◽  
Pregnancy Termination ◽  
Genetic Disorder ◽  
Single Gene ◽  
Genetic Diagnosis ◽  
Early Embryo ◽  
Knowing That

The aim of preimplantation genetic diagnosis (PGD) is to give couples at risk of passing on a genetic disorder an alternative to standard prenatal diagnosis by enabling them to start a pregnancy that is known to be free of the familial disease. This can be achieved by generating embryos in vitro by standard in vitro fertilization (IVF) techniques and then removing one to two of the cells from the early embryo (embryo biopsy). Single cell polymerase chain reaction (PCR) or fluorescence in situ hybridization (FISH) can then be used to diagnose single gene defects or chromosomal abnormalities respectively. Those embryos diagnosed as free from disease can then be considered for transfer to the womb and so the pregnancy is started knowing that the fetus is unaffected. This avoids the need to consider pregnancy termination in the quest for a healthy child. Originally it was thought that the major reason for referral would be the risk of passing on a single abnormal gene but an increasing proportion of couples are requesting PGD because of recurrent miscarriage due to parental chromosomal abnormality.

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