Complex preimplantation genetic tests for Robertsonian translocation, HLA, and X-linked hyper IgM syndrome caused by a novel mutation of CD40LG gene

Abstract

Purpose To perform complex preimplantation genetic tests (PGT) for aneuploidy screening, Robertsonian translocation, HLAmatching, and X-linked hyper IgM syndrome (XHIGM) caused by a novel mutation c.156 G>T of CD40LG gene. Methods Reverse transcription PCR (RT-PCR) and Sanger sequencing were carried out to confirm the causative variant of CD40LG gene in the proband and parents. Day 5 and D6 blastocysts, obtained by in vitro fertilization (IVF) with intracytoplasmic sperm injection, underwent trophectoderm (TE) biopsy and whole genomic amplification (WGA) and next generation sequencing (NGS)-based PGT to detect the presence of a maternal CD40LG mutation, aneuploidy, Robertsonian translocation carrier, and human leukocyte antigen (HLA) haplotype. Results Sanger sequencing data of the genomic DNA showed that the proband has a hemizygous variant of c. 156 G>T in the CD40LG gene, while his mother has a heterozygous variant at the same position. Complementary DNA (cDNA) of CD40LG amplification and sequencing displayed that no cDNA of CD40LG was found in proband, while only wild-type cDNA of CD40LG was amplified in the mother. PGT results showed that only one of the six tested embryos is free of the variant c.156 G>T and aneuploidy and having the consistent HLA type as the proband. Meanwhile, the embryo is a Robertsonian translocation carrier. The embryo was transplanted into the mother's uterus. Amniotic fluid testing results are consistent with that of PGT. A healthy baby girl was delivered, and the peripheral blood testing data was also consistent with the testing results of transplanted embryo. Conclusions The novel mutation of c. 156 G>T in CD40LG gene probably leads to XHIGM by nonsense-meditated mRNA decay (NMD), and complex PGT of preimplantation genetic testing for monogenic disease (PGT-M), aneuploidy (PGT-A), structural rearrangement (PGT-SR), and HLA-matching (PGT-HLA) can be performed in pedigree with both X-linked hyper IgM syndrome and Robertsonian translocation.

OGM and WES identifies translocation breakpoints in PKD1 gene in an polycystic kidney patient and healthy baby delivered using PGT

“…The criteria for grading blastocyst were following the recommendation by Schoolcraft et al Totally 3 embryos were biopsied after IVF on day 5 or 6 and biopsied cells were washed and transferred to a PCR tube containing 2.5 μL phosphate buffer saline (PBS) under strictly sterile and DNA-free conditions against contamination. Whole-genome amplification (WGA) was performed following the protocol of the REPLI-g Single Cell Kit (QIAGEN GmbH, Hilden, Germany) [ 16 , 17 ]. The WGA product was used for the subsequent tests, including PKD1 mutation analysis, aneuploidy analysis, and reciprocal translocation carrier screening.…”

OGM and WES Identifies Translocation Breakpoints in PKD1 Gene in an Polycystic Kidney Patient and Healthy Baby Delivered Using PGT

“…The criteria for grading blastocyst were following the recommendation by Schoolcraft et al Totally 3 embryos were biopsied after IVF on day 5 or 6 and biopsied cells were washed and transferred to a PCR tube containing 2.5L phosphate buffer saline (PBS) under strictly sterile and DNAfree conditions against contamination. Whole-genome ampli cation (WGA) was performed following the protocol of the REPLI-g Single Cell Kit (QIAGEN GmbH, Hilden, Germany) [16,17]. The WGA product was used for the subsequent tests, including PKD1 mutation analysis, aneuploidy analysis, and reciprocal translocation carrier screening.…”

Combined Preimplantation Genetic Testing for Genetic Kidney Disease: Genetic Risk Identification, Assisted Reproductive Cycle, and Pregnancy Outcome Analysis

“…This approach thereby greatly reduces the chance of having a pregnancy affected with the genetic disease. Since the initial practice of PGT in the monogenetic disorders in 1990s (5), it has been extensively employed in the diagnosis of monogenic disease, X-linked disorders, aneuploidy, and chromosomal rearrangements (3,(6)(7)(8)(9)(10). Carrier screening is becoming standard practice for egg and sperm donors and couples seeking assisted reproduction, due to the introduction of target panels that screen for multiple variants in low risk Abbreviations: AD, autosomal dominant; AR, autosomal recessive; ART, assisted reproductive technology; ACMG, the American College of Medical Genetics and Genomics; BPR, biochemical pregnancy rate; CAKUT, congenital anomalies of the kidney and the urinary tract; CKD, chronic kidney disease; CNV, copy number variations; COH, controlled ovarian hyperstimulation; CSA, clinical sequence analyser; E2, estradiol; ECS, expanded carrier screening; ESRD, endstage renal disease; FHB, fetal heartbeat; FETs, frozenembryo transfers; GnRH, gonadotrophic releasing hormone; HGVS, human genome variation society; hCG, human chorionic gonadotropin; ICSI, intracytoplasmic sperm injection; IR, implantation rate; IVF, in vitro fertilization; MII, metaphase II stage; NGS, next-generation sequencing; NS, nephrotic syndrome; NPHP, nephronophthisis; OP/LBR, ongoing pregnancy/live birth rate; PGT, preimplantation genetic testing; PGT-A, PGT for aneuploidies; PGT-SR, PGT for structural rearrangements; PGT-M, preimplantation genetic testing for monogenic disease; PHCG, positive-human chorionic gonadotropin; PKD, polycystic kidney disease; PND, invasive prenatal diagnosis; TE, trophectoderm; SAB, spontaneous abortion; SNP, single nucleotide polymorphism; WES, whole exome sequencing; WGS, whole genome sequencing.…”