Name: non-invasive chromosome screening (nics) of human preimplantation embryos: sample collection and chromosomal ploidy analysis by malbac-ngs
Uploaded: 2017-05-12T11:36:31
Description: Chromosomal abnormalities are common in human embryos and cause implantation failure, early pregnancy losses, and birth defects in practice of assisted ...

The work was supported by The Natural Science Foundation of Jiangsu province, China, No. BK20131094, the Joint Research Program of Medical Science and Technology Development Fund of the Medical Control Center in Wuxi City, No. YGZX1204, the National natural science Foundation of China (No. 81503655) and the State Key Development Program for Basic Research of China (Grant No. 2013CB945200). This work was supported as a major scientific research project of the Wuxi Commission of Reproduction Health&#65288;z201602).

Ethics statement: Institutional review board (IRB) approvals (Nanjing Jinlin: 2014NZKY-005; Wuxi Maternity: 2014-04-0515-02) were obtained. All of the embryos were voluntarily donated by patients, with informed consent obtained before performing the experiments on each embryo.
1. Preparation
<ol>
	<li>Prepare the following reagents
	<ol>
		<li>Prepare fertilization medium plus 10% serum protein substitute (SPS) (IM) by adding 1 mL SPS to 9 mL Quinn&#39;s fertilization medium.</li>
		<li>Make...

<ol>
	<li>Barlow, P. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Early+pregnancy+loss+and+obstetrical+risk+after+in-vitro+fertilization+and+embryo+replacement.">Early pregnancy loss and obstetrical risk after in-vitro fertilization and embryo replacement.</a> Hum Reprod. 3 (5), 671-675 (1988).</li><li>Munne, S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Chromosome+abnormalities+and+their+relationship+to+morphology+and+development+of+human+embryos.">Chromosome abnormalities and their relationship to morphology and development of human embryos.</a> Reprod Biomed Online. 12 (2), 234-253 (2006).</li><li>Harton, G. L. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Diminished+effect+of+maternal+age+on+implantation+after+preimplantation+genetic+diagnosis+with+array+comparative+genomic+hybridization.">Diminished effect of maternal age on implantation after preimplantation genetic diagnosis with array comparative genomic hybridization.</a> Fertil Steril. 100 (6), 1695-1703 (2013).</li><li>Hodes-Wertz, B., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Idiopathic+recurrent+miscarriage+is+caused+mostly+by+aneuploid+embryos.">Idiopathic recurrent miscarriage is caused mostly by aneuploid embryos.</a> Fertil Steril. 98 (3), 675-680 (2012).</li><li>Keltz, M. D., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Preimplantation+Genetic+Screening+(PGS)+with+Comparative+Genomic+Hybridization+(CGH)+following+day+3+single+cell+blastomere+biopsy+markedly+improves+IVF+outcomes+while+lowering+multiple+pregnancies+and+miscarriages.">Preimplantation Genetic Screening (PGS) with Comparative Genomic Hybridization (CGH) following day 3 single cell blastomere biopsy markedly improves IVF outcomes while lowering multiple pregnancies and miscarriages.</a> J Assist Reprod Genet. 30 (10), 1333-1339 (2013).</li><li>Scott, R. T. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Blastocyst+biopsy+with+comprehensive+chromosome+screening+and+fresh+embryo+transfer+significantly+increases+in+vitro+fertilization+implantation+and+delivery+rates:+a+randomized+controlled+trial.">Blastocyst biopsy with comprehensive chromosome screening and fresh embryo transfer significantly increases in vitro fertilization implantation and delivery rates: a randomized controlled trial.</a> Fertil Steril. 100 (3), 697-703 (2013).</li><li>Forman, E. J., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=In+vitro+fertilization+with+single+euploid+blastocyst+transfer:+a+randomized+controlled+trial.">In vitro fertilization with single euploid blastocyst transfer: a randomized controlled trial.</a> Fertil Steril. 100 (1), 100-107 (2013).</li><li>Yang, Z., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Selection+of+single+blastocysts+for+fresh+transfer+via+standard+morphology+assessment+alone+and+with+array+CGH+for+good+prognosis+IVF+patients:+results+from+a+randomized+pilot+study.">Selection of single blastocysts for fresh transfer via standard morphology assessment alone and with array CGH for good prognosis IVF patients: results from a randomized pilot study.</a> Mol Cytogenet. 5 (1), 24 (2012).</li><li>Cimadomo, D., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+Impact+of+Biopsy+on+Human+Embryo+Developmental+Potential+during+Preimplantation+Genetic+Diagnosis.">The Impact of Biopsy on Human Embryo Developmental Potential during Preimplantation Genetic Diagnosis.</a> BioMed Res Intl. 2016, 1-10 (2016).</li><li>Wu, Y., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Blastomere+biopsy+influences+epigenetic+reprogramming+during+early+embryo+development,+which+impacts+neural+development+and+function+in+resulting+mice.">Blastomere biopsy influences epigenetic reprogramming during early embryo development, which impacts neural development and function in resulting mice.</a> Cell Mol Life Sci. 71 (9), 1761-1774 (2013).</li><li>Zhao, H. C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Aberrant+Epigenetic+Modification+in+Murine+Brain+Tissues+of+Offspring+from+Preimplantation+Genetic+Diagnosis+Blastomere+Biopsies.">Aberrant Epigenetic Modification in Murine Brain Tissues of Offspring from Preimplantation Genetic Diagnosis Blastomere Biopsies.</a> Biol Reprod. 89 (5), 117 (2013).</li><li>Zeng, Y. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Preimplantation+genetic+diagnosis+(PGD)+influences+adrenal+development+and+response+to+cold+stress+in+resulting+mice.">Preimplantation genetic diagnosis (PGD) influences adrenal development and response to cold stress in resulting mice.</a> Cell Tissue Res. 354 (3), 729-741 (2013).</li><li>Palini, S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Genomic+DNA+in+human+blastocoele+fluid.">Genomic DNA in human blastocoele fluid.</a> Reprod BioMed Online. 26 (6), 603-610 (2013).</li><li>Gianaroli, L. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Blastocentesis:+a+source+of+DNA+for+preimplantation+genetic+testing.+Results+from+a+pilot+study.">Blastocentesis: a source of DNA for preimplantation genetic testing. Results from a pilot study.</a> Fertil Steril. 102 (6), 1692-1699 (2014).</li><li>Stigliani, S., Anserini, P., Venturini, P. L., Scaruffi, P. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Mitochondrial+DNA+content+in+embryo+culture+medium+is+significantly+associated+with+human+embryo+fragmentation.">Mitochondrial DNA content in embryo culture medium is significantly associated with human embryo fragmentation.</a> Human Reprod. 28 (10), 2652-2660 (2013).</li><li>Stigliani, S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Mitochondrial+DNA+in+Day+3+embryo+culture+medium+is+a+novel,+non-invasive+biomarker+of+blastocyst+potential+and+implantation+outcome.">Mitochondrial DNA in Day 3 embryo culture medium is a novel, non-invasive biomarker of blastocyst potential and implantation outcome.</a> Mol Hum Reprod. 20 (12), 1238-1246 (2014).</li><li>Wu, H., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Medium-Based+Noninvasive+Preimplantation+Genetic+Diagnosis+for+Human+&#945;-Thalassemias-SEA.">Medium-Based Noninvasive Preimplantation Genetic Diagnosis for Human &#945;-Thalassemias-SEA.</a> Medicine. 94 (12), e669 (2015).</li><li>Xu, J., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Noninvasive+chromosome+screening+of+human+embryos+by+genome+sequencing+of+embryo+culture+medium+for+in+vitro+fertilization.">Noninvasive chromosome screening of human embryos by genome sequencing of embryo culture medium for in vitro fertilization.</a> Proc Nat Acad Sci. 113 (42), 11907-11912 (2016).</li><li>Palermo, G. D., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Births+after+intracytoplasmic+injection+of+sperm+obtained+by+testicular+extraction+from+men+with+nonmosaic+Klinefelter's+syndrome.">Births after intracytoplasmic injection of sperm obtained by testicular extraction from men with nonmosaic Klinefelter's syndrome.</a> N Engl J Med. 338 (9), 588-590 (1998).</li><li>Alpha Scientists in Reproductive Medicine and ESHRE Special Interest Group of Embryology. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+Istanbul+consensus+workshop+on+embryo+assessment:+proceedings+of+an+expert+meeting.">The Istanbul consensus workshop on embryo assessment: proceedings of an expert meeting.</a> Hum Reprod. 26 (6), 1270-1283 (2011).</li><li>. Qubit dsDNA HS Assay Kit Available from: <a target="_blank" href="https://www.thermofisher.com/order/catalog/product/Q32851?ICID=search-product">https://www.thermofisher.com/order/catalog/product/Q32851?ICID=search-product</a> (2015)</li><li>. Miseq system use guide Available from: <a target="_blank" href="https://support.illumina.com/downloads/miseq_system_user_guide_15027617.html">https://support.illumina.com/downloads/miseq_system_user_guide_15027617.html</a> (2015)</li></ol>

If the NICS results are contaminated with parental genetic material, then make sure all cumulus-corona radiata cells are removed and ensure that ICSI is performed for fertilization. Avoid inappropriate storage of the culture medium or template preparation processes that can potentially degrade DNA. Decontaminate the workspace thoroughly by DNase and RNase decontamination reagents. To avoid the contamination from other embryos, always culture one embryo in single droplet of medium to avoid cross-contamination from Day 3. ...

Assisted reproductive technologies (ART) have been increasingly used for treatment of infertility. However, the success rate of ART, such as in-vitro fertilization (IVF) has been limited and the pregnancy loss rate has been significantly higher than the normal population 1. The main cause of these problems are chromosomal abnormalities, which commonly exist in preimplantation human embryos 2. PGS came as an effective way to screen the embryos for chromosomal balance before implantation 3,4. And some studies have proved PGS can reduce the rate of abortion and improve the rate of pregnancy 5,6,7,8. However, PGS requires complex technical expertise, which requires specific training and experience. The invasive embryo biopsy procedure could also potentially cause damage to the embryos 9,10. Although long-term biosafety issue of embryo biopsy has not been evaluated thoroughly in human yet, animal studies have shown its negative influences on embryo development 11,12.
It was previously reported that trace amount of DNA materials are secreted into the culture medium during embryo development, and efforts have been made to perform comprehensive chromosome screening (CCS) using spent embryo culture medium 13,14,15,16,17. However, the detection rate and the accuracy of the tests have not met the requirement of extensive clinical use. We reported an improvement of using MALBAC for increasing the detection rate as well as the accuracy of the NICS test 18. Here we report a detailed protocol including spent media sample preparation, NGS preparation and data analysis. By carefully removing cumulus cells from the oocytes, we perform intracytoplasmic single sperm injection (ICSI) and blastocyst culture. We then collect the Day 3-Day 5/Day 6 spent medium for MALBAC WGA and NGS library preparation. By using the NICS technology, we streamline the WGA and NGS library preparation steps in about 3 h, which enables obtaining CCS results noninvasively in about 9 h.

<table>
	<tbody>
		<tr>
			<td>Hyaluronidase solution, 80 U/mL</td>
			<td>SAGE</td>
			<td>ART4007-A</td>
			<td>Digest oocyte-corona-cumulus complex</td>
		</tr>
		<tr>
			<td>Quinn&#39;s Advantage m-HTF Medium with HEPES</td>
			<td>SAGE</td>
			<td>ART-1023</td>
			<td>For embryo clutrure</td>
		</tr>
		<tr>
			<td>Quinn&#39;s Advantage Fertilization Medium</td>
			<td>SAGE</td>
			<td>ART-1020</td>
			<td>For oocyte and sperm fertilization</td>
		</tr>
		<tr>
			<td>Quinn&#39;s Advantage Cleavage Medium</td>
			<td>SAGE</td>
			<td>ART-1026</td>
			<td>For embryo cleavage stage culture</td>
		</tr>
		<tr>
			<td>Quinn&#39;s Advantage Blastocyst Medium</td>
			<td>SAGE</td>
			<td>ART-1029</td>
			<td>For embryo blastocyst stage culture</td>
		</tr>
		<tr>
			<td>Quinn&#39;s Advantage SPS Serum protein Substitute Kit</td>
			<td>SAGE</td>
			<td>ART-3010</td>
			<td>To denude the oocyte</td>
		</tr>
		<tr>
			<td>Quinn&#39;s Advantage Tissue culture mineral oil</td>
			<td>SAGE</td>
			<td>ART-4008P</td>
			<td>To cover the culture medium</td>
		</tr>
		<tr>
			<td>STRIPPER TIPS</td>
			<td>ORIGIO</td>
			<td>MXL3-IND-135</td>
			<td>For denudating granulosa cells</td>
		</tr>
		<tr>
			<td>Pasteur pipettes</td>
			<td>ORIGIO</td>
			<td>PP-9-1000</td>
			<td>For IVF laboratory</td>
		</tr>
		<tr>
			<td>ZILOS-tk Laser System</td>
			<td>Hamilton Thorne</td>
			<td>CLASS 1 laser</td>
			<td>For artificial blastocoele collapse</td>
		</tr>
		<tr>
			<td>ICSI</td>
			<td>ORIGIO</td>
			<td>MPH-35-35</td>
			<td>For ICSI</td>
		</tr>
		<tr>
			<td>HOLDNIG</td>
			<td>ORIGIO</td>
			<td>MPH-MED-35</td>
			<td>For ICSI</td>
		</tr>
		<tr>
			<td>9&#34;IVF Pasteur Pipette</td>
			<td>Oirgio</td>
			<td>MXL3-IND-135</td>
			<td>For embryo tansfer</td>
		</tr>
		<tr>
			<td>microscope</td>
			<td>OLYMPUS</td>
			<td>1X71</td>
			<td>For embryo observation</td>
		</tr>
		<tr>
			<td>incubator</td>
			<td>Labotect</td>
			<td>Inkubator C16</td>
			<td>For embryo culture</td>
		</tr>
		<tr>
			<td>Vitrification kit</td>
			<td>KITAZATO BioPharma</td>
			<td>VT101</td>
			<td>For embryo vitrification</td>
		</tr>
		<tr>
			<td>ES (Vitrification kit)</td>
			<td>KITAZATO BioPharma</td>
			<td>Reagent inVitrification kit</td>
			<td>For embryo vitrification</td>
		</tr>
		<tr>
			<td>VS (Vitrification kit)</td>
			<td>KITAZATO BioPharma</td>
			<td>Reagent inVitrification kit</td>
			<td>For embryo vitrification</td>
		</tr>
		<tr>
			<td>Cryotop open systerm</td>
			<td>KITAZATO BioPharma</td>
			<td>81110</td>
			<td>For embryo vitrification</td>
		</tr>
		<tr>
			<td>BD Falcon Tissue culture Dishes, Sterile</td>
			<td>BD Bioscience</td>
			<td>353002</td>
			<td>For embryo culture</td>
		</tr>
		<tr>
			<td>BD Falcon Tissue culture Dishes&#65288;Easy Grip&#65289;, Sterile</td>
			<td>BD Bioscience</td>
			<td>353001</td>
			<td>For embryo culture</td>
		</tr>
		<tr>
			<td>BD Falcon Organ Culture Dish, Sterile</td>
			<td>BD Bioscience</td>
			<td>363037</td>
			<td>For embryo culture</td>
		</tr>
		<tr>
			<td>Nunc IVF 4-Well Dish</td>
			<td>Thermo Scientific</td>
			<td>144444</td>
			<td>For embryo washing and blastocyst culture</td>
		</tr>
		<tr>
			<td>Vitrification Cryotop Open systerm</td>
			<td>KIZTAZATO</td>
			<td>81111</td>
			<td>For embryo vitrification</td>
		</tr>
		<tr>
			<td>NICSInst library preparation kit</td>
			<td>Yikon Genomics</td>
			<td>KT1000800324</td>
			<td>Whole genome amplification and library construction</td>
		</tr>
		<tr>
			<td>MT Enzyme Mix</td>
			<td>Yikon Genomics</td>
			<td>Reagent in NICSInst library preparation kit</td>
			<td>For culture medium pre-treatment</td>
		</tr>
		<tr>
			<td>Cell Lysis Buffer</td>
			<td>Yikon Genomics</td>
			<td>Reagent in NICSInst library preparation kit</td>
			<td>For culture medium pre-treatment</td>
		</tr>
		<tr>
			<td>Cell Lysis Enzyme</td>
			<td>Yikon Genomics</td>
			<td>Reagent in NICSInst library preparation kit</td>
			<td>For culture medium pre-treatment</td>
		</tr>
		<tr>
			<td>Pre-Lib Buffer</td>
			<td>Yikon Genomics</td>
			<td>Reagent in NICSInst library preparation kit</td>
			<td>Pre-library preparation</td>
		</tr>
		<tr>
			<td>Pre-Lib Enzyme</td>
			<td>Yikon Genomics</td>
			<td>Reagent in NICSInst library preparation kit</td>
			<td>Pre-library preparation</td>
		</tr>
		<tr>
			<td>Barcode Primer1-48</td>
			<td>Yikon Genomics</td>
			<td>Reagent in NICSInst library preparation kit</td>
			<td>For library amplificaton</td>
		</tr>
		<tr>
			<td>Library buffer</td>
			<td>Yikon Genomics</td>
			<td>Reagent in NICSInst library preparation kit</td>
			<td>For library amplificaton</td>
		</tr>
		<tr>
			<td>Library Enzyme Mix</td>
			<td>Yikon Genomics</td>
			<td>Reagent in NICSInst library preparation kit</td>
			<td>For library amplificaton</td>
		</tr>
		<tr>
			<td>CMPure Magbeads</td>
			<td>Yikon Genomics</td>
			<td>Reagent in NICSInst library preparation kit</td>
			<td>For library purification</td>
		</tr>
		<tr>
			<td>Distill water</td>
			<td>Yikon Genomics</td>
			<td>Reagent in NICSInst library preparation kit</td>
			<td>To dissolve DNA</td>
		</tr>
		<tr>
			<td>NICSInst Sample Prep Station</td>
			<td>Yikon Genomics</td>
			<td>ME1001003</td>
			<td>Amplificate DNA</td>
		</tr>
		<tr>
			<td>Illumina MiSeq System</td>
			<td>Illumina</td>
			<td>SY-410-1001</td>
			<td>For library sequencing</td>
		</tr>
		<tr>
			<td>Vortexer</td>
			<td>Qilinbeier</td>
			<td>DNYS8</td>
			<td>Sample mix</td>
		</tr>
		<tr>
			<td>Mini-centrifuge</td>
			<td>ESSENSCIEN</td>
			<td>ELF6</td>
			<td>For separation</td>
		</tr>
		<tr>
			<td>Magnetic Stand</td>
			<td>DynaMagTM-2</td>
			<td>12321D</td>
			<td>For library purification</td>
		</tr>
		<tr>
			<td>100 % ethanol</td>
			<td>Sinopharm Chemical</td>
			<td>10009218</td>
			<td>For DNA library purification</td>
		</tr>
		<tr>
			<td>Qubit 3.0 Fluorometer</td>
			<td>Thermo Scientific</td>
			<td>Q33216</td>
			<td>For library quantification</td>
		</tr>
		<tr>
			<td>10 &#181;L, 200 &#181;L,1000 &#181;L DNase /RNase Free Tips</td>
			<td>Axygen</td>
			<td>T-300-R-S, 
			T-200-Y-R-S, 
			T-1000-B-R-S</td>
			<td>For sample transfer</td>
		</tr>
		<tr>
			<td>1.5 mL EP tube, 0.2 mL PCR tube</td>
			<td>Axygen</td>
			<td>MCT-150-C, 
			PCR-02-C</td>
			<td>DNase/RNase free, Low Binding PCR tubes and 1.5 mL micro-centrifuge tubes are recommended.</td>
		</tr>
	</tbody>
</table>

non-invasive chromosome screening (nics) of human preimplantation embryos: sample collection and chromosomal ploidy analysis by malbac-ngs

Chromosomal abnormalities are common in human embryos and cause implantation failure, early pregnancy losses, and birth defects in practice of assisted reproductive technology (ART). Non-invasive chromosome screening (NICS) is an emerging technology that enables the selection of chromosomal-balanced embryos, without performing invasive embryo biopsy. Here we report the full protocol of NICS, which includes culture medium pretreatment, whole genome amplification (WGA) by multiple annealing and looping-based amplification cycles (MALBAC), library preparation for next generation sequencing (NGS) and NGS data analysis. To validate the reliability and efficiency of NICS, we have already performed NICS on 27 transfer cycles in 23 couples with balanced translocation, azoospermia, recurrent pregnancy loss (RPL), or recurrent implantation failure (RIF), 17 of them have achieved successful clinical pregnancies, and 9 among them have already obtained healthy live births. No pregnancy loss has been reported thus far. The NICS method avoids the need for embryo biopsy and therefore substantially increases the safety of its use.

We applied the method on a patient with a balanced translocation. IRB approval and informed consents were obtained before applying the NICS assay on the patient. Karyotype analysis of the patient showed a balanced translocation (1; 18) (p13.3; q21). We obtained a total of six blastocysts from the patient and performed NICS on Day 3-Day 5 culture medium of all six embryos. Chromosome abnormalities caused by the parents&#39; balanced translocation were detected in five of them with the NICS...

We report a protocol for chromosome screening of human embryos by using spent culture medium, which avoids embryo biopsy and enables reporting chromosome ploidy using next generation sequencing (NGS). We present the detailed procedure including the preparation of culture medium, whole genome amplification (WGA), NGS library preparation, and data analysis.

Non-invasive Chromosome Screening (NICS) of Human Preimplantation Embryos: Sample Collection and Chromosomal Ploidy Analysis by MALBAC-NGS

Chromosomal abnormalities are common in human embryos and cause implantation failure, early pregnancy losses, and birth defects in practice of assisted ...