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Erik Borgström, Marta Paterlini, Jeff E Mold, Jonas Frisen, Joakim Lundeberg
BACKGROUND: Whole genome amplification (WGA) is currently a prerequisite for single cell whole genome or exome sequencing. Depending on the method used the rate of artifact formation, allelic dropout and sequence coverage over the genome may differ significantly. RESULTS: The largest difference between the evaluated protocols was observed when analyzing the target coverage and read depth distribution. These differences also had impact on the downstream variant calling...
2017: PloS One
Jiawei Xu, Wenbin Niu, Zhaofeng Peng, Xiao Bao, Meixiang Zhang, Linlin Wang, Linqing Du, Nan Zhang, Yingpu Sun
Triploidy occurred about 2-3% in human pregnancies and contributed to approximately 15% of chromosomally caused human early miscarriage. It is essential for preimplantation genetic diagnosis and screen to distinct triploidy sensitively. Here, we performed comparative investigations between MALBAC-NGS and MDA-SNP array sensitivity on triploidy detection. Self-correction and reference-correction algorism were used to analyze the NGS data. We identified 5 triploid embryos in 1198 embryos of 218 PGD and PGS cycles using MDA-SNP array, the rate of tripoidy was 4...
November 9, 2016: Oncotarget
Juanjuan Xu, Rui Fang, Li Chen, Daozhen Chen, Jian-Ping Xiao, Weimin Yang, Honghua Wang, Xiaoqing Song, Ting Ma, Shiping Bo, Chong Shi, Jun Ren, Lei Huang, Li-Yi Cai, Bing Yao, X Sunney Xie, Sijia Lu
Preimplantation genetic screening (PGS) is widely used to select in vitro-fertilized embryos free of chromosomal abnormalities and to improve the clinical outcome of in vitro fertilization (IVF). A disadvantage of PGS is that it requires biopsy of the preimplantation human embryo, which can limit the clinical applicability of PGS due to the invasiveness and complexity of the process. Here, we present and validate a noninvasive chromosome screening (NICS) method based on sequencing the genomic DNA secreted into the culture medium from the human blastocyst...
October 18, 2016: Proceedings of the National Academy of Sciences of the United States of America
Jin Huang, Liying Yan, Sijia Lu, Nan Zhao, X Sunney Xie, Jie Qiao
OBJECTIVE: To validate a 24-chromosome aneuploidy preimplantation genetic screening protocol based on multiple annealing and looping-based amplification cycle (MALBAC) and next-generation sequencing (NGS). DESIGN: Single-nucleotide polymorphism (SNP) array and MALBAC-NGS analysis. SETTING: University-affiliated in vitro fertilization (IVF) center. PATIENT(S): Fifteen women from whom 30 blastocysts were obtained for genotyping...
June 2016: Fertility and Sterility
Liying Yan, Lei Huang, Liya Xu, Jin Huang, Fei Ma, Xiaohui Zhu, Yaqiong Tang, Mingshan Liu, Ying Lian, Ping Liu, Rong Li, Sijia Lu, Fuchou Tang, Jie Qiao, X Sunney Xie
In vitro fertilization (IVF), preimplantation genetic diagnosis (PGD), and preimplantation genetic screening (PGS) help patients to select embryos free of monogenic diseases and aneuploidy (chromosome abnormality). Next-generation sequencing (NGS) methods, while experiencing a rapid cost reduction, have improved the precision of PGD/PGS. However, the precision of PGD has been limited by the false-positive and false-negative single-nucleotide variations (SNVs), which are not acceptable in IVF and can be circumvented by linkage analyses, such as short tandem repeats or karyomapping...
December 29, 2015: Proceedings of the National Academy of Sciences of the United States of America
Yong Hou, Kui Wu, Xulian Shi, Fuqiang Li, Luting Song, Hanjie Wu, Michael Dean, Guibo Li, Shirley Tsang, Runze Jiang, Xiaolong Zhang, Bo Li, Geng Liu, Niharika Bedekar, Na Lu, Guoyun Xie, Han Liang, Liao Chang, Ting Wang, Jianghao Chen, Yingrui Li, Xiuqing Zhang, Huanming Yang, Xun Xu, Ling Wang, Jun Wang
BACKGROUND: Single-cell resequencing (SCRS) provides many biomedical advances in variations detection at the single-cell level, but it currently relies on whole genome amplification (WGA). Three methods are commonly used for WGA: multiple displacement amplification (MDA), degenerate-oligonucleotide-primed PCR (DOP-PCR) and multiple annealing and looping-based amplification cycles (MALBAC). However, a comprehensive comparison of variations detection performance between these WGA methods has not yet been performed...
2015: GigaScience
Lieselot Deleye, Dieter De Coninck, Christodoulos Christodoulou, Tom Sante, Annelies Dheedene, Björn Heindryckx, Etienne Van den Abbeel, Petra De Sutter, Björn Menten, Dieter Deforce, Filip Van Nieuwerburgh
Current whole genome amplification (WGA) methods lead to amplification bias resulting in over- and under-represented regions in the genome. Nevertheless, certain WGA methods, such as SurePlex and subsequent arrayCGH analysis, make it possible to detect copy number alterations (CNAs) at a 10 Mb resolution. A more uniform WGA combined with massive parallel sequencing (MPS), however, could allow detection at higher resolution and lower cost. Recently, MALBAC, a new WGA method, claims unparalleled performance...
2015: Scientific Reports
Luwen Ning, Zhoufang Li, Guan Wang, Wen Hu, Qingming Hou, Yin Tong, Meng Zhang, Yao Chen, Li Qin, Xiaoping Chen, Heng-Ye Man, Pinghua Liu, Jiankui He
Single-cell genomic analysis has grown rapidly in recent years and finds widespread applications in various fields of biology, including cancer biology, development, immunology, pre-implantation genetic diagnosis, and neurobiology. To date, the amplification bias, amplification uniformity and reproducibility of the three major single cell whole genome amplification methods (GenomePlex WGA4, MDA and MALBAC) have not been systematically investigated using mammalian cells. In this study, we amplified genomic DNA from individual hippocampal neurons using three single-cell DNA amplification methods, and sequenced them at shallow depth...
2015: Scientific Reports
Lei Huang, Fei Ma, Alec Chapman, Sijia Lu, Xiaoliang Sunney Xie
We present a survey of single-cell whole-genome amplification (WGA) methods, including degenerate oligonucleotide-primed polymerase chain reaction (DOP-PCR), multiple displacement amplification (MDA), and multiple annealing and looping-based amplification cycles (MALBAC). The key parameters to characterize the performance of these methods are defined, including genome coverage, uniformity, reproducibility, unmappable rates, chimera rates, allele dropout rates, false positive rates for calling single-nucleotide variations, and ability to call copy-number variations...
2015: Annual Review of Genomics and Human Genetics
Na Li, Li Wang, Hui Wang, Minyue Ma, Xiaohong Wang, Yi Li, Wenke Zhang, Jianguang Zhang, David S Cram, Yuanqing Yao
Reliable and accurate pre-implantation genetic diagnosis (PGD) of patient's embryos by next-generation sequencing (NGS) is dependent on efficient whole genome amplification (WGA) of a representative biopsy sample. However, the performance of the current state of the art WGA methods has not been evaluated for sequencing. Using low template DNA (15 pg) and single cells, we showed that the two PCR-based WGA systems SurePlex and MALBAC are superior to the REPLI-g WGA multiple displacement amplification (MDA) system in terms of consistent and reproducible genome coverage and sequence bias across the 24 chromosomes, allowing better normalization of test to reference sequencing data...
April 20, 2015: Journal of Genetics and Genomics, Yi Chuan Xue Bao
Cheng-Zhong Zhang, Viktor A Adalsteinsson, Joshua Francis, Hauke Cornils, Joonil Jung, Cecile Maire, Keith L Ligon, Matthew Meyerson, J Christopher Love
Artifacts introduced in whole-genome amplification (WGA) make it difficult to derive accurate genomic information from single-cell genomes and require different analytical strategies from bulk genome analysis. Here, we describe statistical methods to quantitatively assess the amplification bias resulting from whole-genome amplification of single-cell genomic DNA. Analysis of single-cell DNA libraries generated by different technologies revealed universal features of the genome coverage bias predominantly generated at the amplicon level (1-10 kb)...
April 16, 2015: Nature Communications
Minfeng Chen, Pengfei Song, Dan Zou, Xuesong Hu, Shancen Zhao, Shengjie Gao, Fei Ling
[This corrects the article DOI: 10.1371/journal.pone.0114520.].
2015: PloS One
Alec R Chapman, Zi He, Sijia Lu, Jun Yong, Longzhi Tan, Fuchou Tang, X Sunney Xie
Recently, Multiple Annealing and Looping-Based Amplification Cycles (MALBAC) has been developed for whole genome amplification of an individual cell, relying on quasilinear instead of exponential amplification to achieve high coverage. Here we adapt MALBAC for single-cell transcriptome amplification, which gives consistently high detection efficiency, accuracy and reproducibility. With this newly developed technique, we successfully amplified and sequenced single cells from 3 germ layers from mouse embryos in the early gastrulation stage, and examined the epithelial-mesenchymal transition (EMT) program among cells in the mesoderm layer on a single-cell level...
2015: PloS One
Minfeng Chen, Pengfei Song, Dan Zou, Xuesong Hu, Shancen Zhao, Shengjie Gao, Fei Ling
Single-cell sequencing promotes our understanding of the heterogeneity of cellular populations, including the haplotypes and genomic variability among different generation of cells. Whole-genome amplification is crucial to generate sufficient DNA fragments for single-cell sequencing projects. Using sequencing data from single sperms, we quantitatively compare two prevailing amplification methods that extensively applied in single-cell sequencing, multiple displacement amplification (MDA) and multiple annealing and looping-based amplification cycles (MALBAC)...
2014: PloS One
Jin Huang, Liying Yan, Wei Fan, Nan Zhao, Yan Zhang, Fuchou Tang, X Sunney Xie, Jie Qiao
OBJECTIVE: To validate multiple annealing and looping-based amplification cycle (MALBAC) sequencing for 24-chromosome aneuploidy screening of cleavage embryos and to explore the chromosomal characteristics of embryos at the cleavage stage. DESIGN: The 24-chromosome aneuploidy analyses of the blastomeres included comparative genomic hybridization (CGH), single nucleotide polymorphism (SNP), and MALBAC sequencing. SETTING: University-affiliated IVF center...
December 2014: Fertility and Sterility
Zhilong Yu, Sijia Lu, Yanyi Huang
We developed a microfluidic device to perform multiplex single-cell whole-genome amplification (WGA) using multiple annealing and looping-based amplification cycles (MALBAC). This device, made of polydimethylsiloxane (PDMS), allows us to monitor the whole process of cell loading and single-cell WGA for sequencing. We show that the genome coverage of MALBAC amplifications is reproducible between chambers on a single chip and between different chips, which enables data normalization using standard samples to accurately identify copy number variations (CNVs)...
October 7, 2014: Analytical Chemistry
Charles F A de Bourcy, Iwijn De Vlaminck, Jad N Kanbar, Jianbin Wang, Charles Gawad, Stephen R Quake
Single-cell sequencing is emerging as an important tool for studies of genomic heterogeneity. Whole genome amplification (WGA) is a key step in single-cell sequencing workflows and a multitude of methods have been introduced. Here, we compare three state-of-the-art methods on both bulk and single-cell samples of E. coli DNA: Multiple Displacement Amplification (MDA), Multiple Annealing and Looping Based Amplification Cycles (MALBAC), and the PicoPLEX single-cell WGA kit (NEB-WGA). We considered the effects of reaction gain on coverage uniformity, error rates and the level of background contamination...
2014: PloS One
Yu Hou, Wei Fan, Liying Yan, Rong Li, Ying Lian, Jin Huang, Jinsen Li, Liya Xu, Fuchou Tang, X Sunney Xie, Jie Qiao
Single-cell genome analyses of human oocytes are important for meiosis research and preimplantation genomic screening. However, the nonuniformity of single-cell whole-genome amplification hindered its use. Here, we demonstrate genome analyses of single human oocytes using multiple annealing and looping-based amplification cycle (MALBAC)-based sequencing technology. By sequencing the triads of the first and second polar bodies (PB1 and PB2) and the oocyte pronuclei from same female egg donors, we phase the genomes of these donors with detected SNPs and determine the crossover maps of their oocytes...
December 19, 2013: Cell
Chenghang Zong, Sijia Lu, Alec R Chapman, X Sunney Xie
Kindred cells can have different genomes because of dynamic changes in DNA. Single-cell sequencing is needed to characterize these genomic differences but has been hindered by whole-genome amplification bias, resulting in low genome coverage. Here, we report on a new amplification method-multiple annealing and looping-based amplification cycles (MALBAC)-that offers high uniformity across the genome. Sequencing MALBAC-amplified DNA achieves 93% genome coverage ≥1x for a single human cell at 25x mean sequencing depth...
December 21, 2012: Science
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