In another PGS case, in which array CGH had been used to detect aneuploidy by copy number analysis of both polar bodies, the WGA products (Sureplex; Illumina, San Diego, CA, USA) from both polar bodies were SNP genotyped along with parental genomic DNAs and, with patients informed consent, WGA products (SureMDA; Illumina, San Diego, CA, USA) of nine corresponding fertilised embryos which had all been diagnosed as aneuploid.
Sureplex
Manufactured by Illumina
Sourced in United States
Sureplex is a library preparation system designed for use with Illumina sequencing platforms. It is a rapid and automated method for generating multiplexed sequencing libraries from DNA samples. The Sureplex system enables the preparation of multiple libraries simultaneously, improving efficiency and throughput.
Automatically generated - may contain errors
Lab products found in correlation
7 protocols using sureplex
1
Polar Body Aneuploidy Screening
2
PGT-M and PGT-SR/A Testing Methods
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Patient counselling and genetic testing methods for PGT-M have been described previously [1 (link)]. Testing for monogenic diseases was either indirect, relying on haplotyping of genetic markers (STR/SNP) across the locus of interest and flanking regions, or direct, coupling pathogenic variant detection to genetic marker analysis. Chromosomal testing for PGT-SR/A has been performed by FISH or by WGA (Sureplex, Illumina) followed by either 24Sure array-CGH (Illumina) or by library preparation (KAPA HyperPlus, Roche), sequencing (Illumina) and an in-house-developed interpretation pipeline. Briefly, laser energy is used to open the zona pellucida on day 4 and then allow the embryo to grow to blastocyst. During the study period, the laser was used to remove cells (‘pulling method’). In the majority (84%) of the biopsies performed in cleavage-stage embryos, one cell was removed. Details on hormonal stimulation, oocyte collections, ICSI, embryo biopsy and transfer can be found in De Rycke et al. [11 (link)].
3
Polar Body Aneuploidy Screening
4
Retrospective PGT-A Data Analysis
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
This study was approved by the Ethics Committee of the First Affiliated Hospital of Anhui Medical University, China (No. 20211217). This study was a retrospective analysis of archived data using clinical samples obtained from patients referred to the Reproductive Medical Center of Anhui Medical University for the purpose of PGT-A. Written informed consent for the analysis of archived data was obtained from patients with the approval of the Ethics Committee.
Embryos in this study were subjected to TE biopsy at the request of the patients for chromosomal aneuploidy assessment. For this purpose, the standard procedure in our laboratory involved whole-genome amplification (WGA; SurePlex, Illumina, CA, USA) followed by next-generation sequencing (NGS) on the Ion Proton (Thermo Fisher Scientific, MA, USA) or NextSeq550 (Illumina, CA, USA) platform. The experimental procedure described herein involved the analysis of archived data following the completion of PGT-A. The embryos were not subjected to additional manipulations, and the clinical treatment of the patients was not changed as a result of their participation in this study.
Embryos in this study were subjected to TE biopsy at the request of the patients for chromosomal aneuploidy assessment. For this purpose, the standard procedure in our laboratory involved whole-genome amplification (WGA; SurePlex, Illumina, CA, USA) followed by next-generation sequencing (NGS) on the Ion Proton (Thermo Fisher Scientific, MA, USA) or NextSeq550 (Illumina, CA, USA) platform. The experimental procedure described herein involved the analysis of archived data following the completion of PGT-A. The embryos were not subjected to additional manipulations, and the clinical treatment of the patients was not changed as a result of their participation in this study.
5
Whole Genome Amplification of Embryo Samples
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
All embryo samples including biopsies, whole embryos and single cells disaggregated from arrested embryos were washed through several drops of phosphate buffered saline (PBS) (Gibco; Life technologies, USA) supplemented with 0.1% polyvinyl alcohol (Sigma-Aldrich, USA) and placed into PCR tubes in 10 µl of PBS. DNA extraction from buccal cell swabs from both parents and close relatives and lysis of embryo samples was as described previously17 (link), 18 (link). Whole genome amplification (WGA) of embryo samples was by a PCR library based method according to manufacturer’s instructions (SurePlex, Illumina, San Diego, USA).
6
Trophoblast Biopsy for Genetic Analysis
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Firstly, blastocysts were graded according to the Gardner blastocyst morphologic scoring system [11 (link)]. Approximately 4–6 trophoblast cells from high quality blastocysts were biopsied 5–6 days after fertilization by an experienced embryologist. In our center, blastocysts were simply shrunk by proper energy at first, then minor laser energy was used to slot the zona pellucida at the position of biopsy. The trophoblast cells to be biopsied were sucked into the biopsy needle through negative pressure, and then were cut by laser along the interface between zona pellucida and biopsy needle. Finally, trophoblast cells were completely segregated. Generally, the biopsied trophoblasts were lysed, and DNA was amplified by whole genome amplification (WGA, SurePlex, Illumina, Inc., San Diego, CA, USA) following the manufacturer’s protocol.
7
Comparative NGS Aneuploidy Detection
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
The concept of utilizing NGS for aneuploidy detection in embryos depends upon WGA in order to provide enough DNA from embryo biopsies for subsequent library construction. After it has been generated the WGA product is fragmented into a library of small fragments (100 to 200 bp), which are sequenced in parallel. These sequences or 'reads' are then aligned to the current human reference genome. The relative number of fragments of DNA sequence reads that map to each chromosome provides an indication of ploidy (e.g. trisomy is associated with an increase in the relative number of reads for the affected chromosome, while a monosomy results in a decrease).
Two different WGA methods were compared; Sureplex (Rubicon) and multiple displacement amplification (MDA) using Repli-G Midi (Qiagen). For comparison with NGS, embryo samples amplified using Sureplex were tested via aCGH (24Sure, Illumina). These embryos were then either rebiopsied and amplified using the Repli-G, or aliquots of the original Sureplex WGA were processed by NGS, all samples analysed both blinded and non-blinded.
WGA using MDA was performed according to the manufacturer's instructions, except for a modification of the incubation time (reduced to 2 h). The WGA using Sureplex was processed as directed by the manufacturer.
Two different WGA methods were compared; Sureplex (Rubicon) and multiple displacement amplification (MDA) using Repli-G Midi (Qiagen). For comparison with NGS, embryo samples amplified using Sureplex were tested via aCGH (24Sure, Illumina). These embryos were then either rebiopsied and amplified using the Repli-G, or aliquots of the original Sureplex WGA were processed by NGS, all samples analysed both blinded and non-blinded.
WGA using MDA was performed according to the manufacturer's instructions, except for a modification of the incubation time (reduced to 2 h). The WGA using Sureplex was processed as directed by the manufacturer.
About PubCompare
Our mission is to provide scientists with the largest repository of trustworthy protocols and intelligent analytical tools, thereby offering them extensive information to design robust protocols aimed at minimizing the risk of failures.
We believe that the most crucial aspect is to grant scientists access to a wide range of reliable sources and new useful tools that surpass human capabilities.
However, we trust in allowing scientists to determine how to construct their own protocols based on this information, as they are the experts in their field.
Ready to get started?
Sign up for free.
Registration takes 20 seconds.
Available from any computer
No download required
Revolutionizing how scientists
search and build protocols!