The Experts below are selected from a list of 223593 Experts worldwide ranked by ideXlab platform
Christopher W. Dick - One of the best experts on this subject based on the ideXlab platform.
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Target Sequence capture in the Brazil nut family (Lecythidaceae): Marker selection and in silico capture from genome skimming data.
Molecular Phylogenetics and Evolution, 2019Co-Authors: Oscar M. Vargas, Myriam Heuertz, Stephen A. Smith, Christopher W. DickAbstract:Abstract Reconstructing species trees from multi-loci datasets is becoming a standard practice in phylogenetics. Nevertheless, access to high-throughput sequencing may be costly, especially with studies of many samples. The potential high cost makes a priori assessments desirable in order to make informed decisions about sequencing. We generated twelve transcriptomes for ten species of the Brazil nut family (Lecythidaceae), identified a set of putatively orthologous nuclear loci and evaluated, in silico, their phylogenetic utility using genome skimming data of 24 species. We designed the markers using MarkerMiner, and developed a script, GoldFinder, to efficiently sub-select the best makers for sequencing. We captured, in silico, all designed 354 nuclear loci and performed a maximum likelihood phylogenetic analysis on the concatenated Sequence matrix. We also calculated individual gene trees with maximum likelihood and used them for a coalescent-based species tree inference. Both analyses resulted in almost identical topologies. However, our nuclear-loci phylogenies were strongly incongruent with a published plastome phylogeny, suggesting that plastome data alone is not sufficient for species tree estimation. Our results suggest that using hundreds of nuclear markers (i.e. 354) will significantly improve the Lecythidaceae species tree. The framework described here will be useful, generally, for developing markers for species tree inference.
Myungjae Song - One of the best experts on this subject based on the ideXlab platform.
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???????????????, 2018Co-Authors: Myungjae SongAbstract:Programmable nuclease is useful tools for genome editing at site-specific locus. For high efficiency of genome editing, it is important that programmable nuclease has high activity. Because Cpf1 is a newly reported nuclease protein of the class 2 CRISPR-Cas system (Zetsche, Gootenberg et al. 2015), it has limited information about Cpf1 activity profiles in mammalian cells. For this reason, Cpf1 preclude its wide use for genome editing. Here, we developed a new high-throughput method for evaluating Cpf1 activity, based on Target Sequence composition in mammalian cells. A library of >11,000 guide RNA and Target Sequence pairs was delivered into human cells by lentiviral vectors. Subsequent delivery of Cpf1 in various methods into this cell library induced indels at the integrated synthetic Target Sequences, which allowed en masse evaluation of guide RNA and Cpf1 activity using deep sequencing. Regardless of delivery method of Cpf1, our hight-throughput method showed similar results and the indel frequencies measured at the integrated Target Sequences correlated with those at the corresponding endogenous sites. In this high-throughput approach, we determined Target Sequence-dependent activity profiles and newly protospacer adjacent motif Sequences of Cpf1 in mammalian cells. And we found that Sequence features of high activity AsCpf1 guide RNAs are distinct from those of SpCas9. Evaluation of activity at mismatched Target Sequences showed that Cpf1 Target Sequences can be divided into seed, trunk, and promiscuous regions depending on mismatch tolerability. Both the Cpf1 characterization profile and the in vivo high-throughput evaluation method will greatly facilitate Cpf1-based genome editing.Docto
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deep learning improves prediction of crispr cpf1 guide rna activity
Nature Biotechnology, 2018Co-Authors: Soobin Jung, Jae Woo Choi, Myungjae Song, Sungroh YoonAbstract:Using deep learning to combine Target Sequence and chromatin accessibility data boosts the accuracy of CRISPR–Cpf1 guide RNA activity
Oscar M. Vargas - One of the best experts on this subject based on the ideXlab platform.
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Target Sequence capture in the Brazil nut family (Lecythidaceae): Marker selection and in silico capture from genome skimming data.
Molecular Phylogenetics and Evolution, 2019Co-Authors: Oscar M. Vargas, Myriam Heuertz, Stephen A. Smith, Christopher W. DickAbstract:Abstract Reconstructing species trees from multi-loci datasets is becoming a standard practice in phylogenetics. Nevertheless, access to high-throughput sequencing may be costly, especially with studies of many samples. The potential high cost makes a priori assessments desirable in order to make informed decisions about sequencing. We generated twelve transcriptomes for ten species of the Brazil nut family (Lecythidaceae), identified a set of putatively orthologous nuclear loci and evaluated, in silico, their phylogenetic utility using genome skimming data of 24 species. We designed the markers using MarkerMiner, and developed a script, GoldFinder, to efficiently sub-select the best makers for sequencing. We captured, in silico, all designed 354 nuclear loci and performed a maximum likelihood phylogenetic analysis on the concatenated Sequence matrix. We also calculated individual gene trees with maximum likelihood and used them for a coalescent-based species tree inference. Both analyses resulted in almost identical topologies. However, our nuclear-loci phylogenies were strongly incongruent with a published plastome phylogeny, suggesting that plastome data alone is not sufficient for species tree estimation. Our results suggest that using hundreds of nuclear markers (i.e. 354) will significantly improve the Lecythidaceae species tree. The framework described here will be useful, generally, for developing markers for species tree inference.
Shusheng Zhang - One of the best experts on this subject based on the ideXlab platform.
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fluorescence detection for dna using hybridization chain reaction with enzyme amplification
Chemical Communications, 2010Co-Authors: Shuyan Niu, Yu Sherry Jiang, Shusheng ZhangAbstract:A signal amplification based on a combination of hybridization chain reaction (HCR) and enzyme-enhancement was applied in the study of a novel biosensor. The Target Sequence was added as catalyst which was simply transmitted but did not disappear as the DNA double strand grew on the surface of magnetic beads (MBs).
Martin R. Buoncristiani - One of the best experts on this subject based on the ideXlab platform.
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a quadruplex real time qpcr assay for the simultaneous assessment of total human dna human male dna dna degradation and the presence of pcr inhibitors in forensic samples a diagnostic tool for str typing
Forensic Science International-genetics, 2008Co-Authors: William R Hudlow, Mark D. Timken, Katie L. Swango, Mavis Date Chong, Martin R. BuoncristianiAbstract:A quadruplex real-time qPCR assay was developed to simultaneously assess total human DNA, human male DNA, DNA degradation and PCR inhibitors in forensic samples. Specifically, the assay utilizes a approximately 170-190bp Target Sequence that spans the TH01 STR locus to quantify total human DNA (nuTH01), a 137 bp Target Sequence directly adjacent to the SRY gene to quantify human male DNA (nuSRY), a 67 bp Target Sequence flanking the CSF1PO STR locus (nuCSF) to assess degradation (nuCSF:nuTH01 ratio) and a 77 bp synthetic DNA template used as an internal PCR control Target Sequence (IPC) for the assessment of PCR inhibition. Validation studies, performed on an ABI 7500 SDS instrument using TaqMan and TaqManMGB detection, indicate each of the Targets in the quadruplex assay performs effectively and is informative even when challenged with DNase-degraded and hematin-inhibited samples. The nuTH01-nuSRY-nuCSF-IPC quadruplex qPCR assay is envisioned to assist in the choice of the most informative DNA typing system available, which may include standard autosomal STR typing when the results indicate the presence of non-degraded, single gender DNA or non-degraded, male:female mixtures at ratios expected to yield probative alleles; Y STR typing in samples containing a male component that is overwhelmed by the presence of an excess of female DNA; reduced amplicon size STR typing ("MiniSTRs") where the nuCSF:nuTH01 ratio indicates the sample is highly degraded; enhanced STR amplification with additional AmpliTaq Gold/BSA and/or sample clean-up when the presence of PCR inhibitors is suggested by a delayed IPC C(T) value or mitochondrial DNA typing in samples where little to no nuclear DNA is detected. The present study includes evaluations of species specificity, sensitivity, precision, reproducibility, male-female mixtures, population samples and applications to various casework-type samples as indicated by the Scientific Working Group on DNA Analysis Methods (SWGDAM) guidelines.
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A duplex real-time qPCR assay for the quantification of human nuclear and mitochondrial DNA in forensic samples: implications for quantifying DNA in degraded samples.
Journal of Forensic Sciences, 2005Co-Authors: Mark D. Timken, Katie L. Swango, Cristián Orrego, Martin R. BuoncristianiAbstract:A duplex real-time qPCR assay was developed for quantifying human nuclear and mitochondrial DNA in forensic samples. The nuclear portion of the assay utilized amplification of a ∼170–190 bp Target Sequence that spans the repeat region of the TH01 STR locus, and the mitochondrial portion of the assay utilized amplification of a 69 bp Target Sequence in the ND1 region. Validation studies, performed on an ABI 7000 SDS instrument using TaqMan® detection, demonstrated that both portions of the duplex assay provide suitable quantification sensitivity and precision down to 10–15 copies of each genome of interest and that neither portion shows cross-reactivity to commonly encountered non-human genomes. As part of the validation studies, a series of DNase-degraded samples were quantified using three different methods: the duplex nuclearmitochondrial qPCR assay, the ABI Quantifiler™ Human DNA Quantification Kit qPCR assay, which amplifies and detects a 62 bp nuclear Target Sequence, and slot blot hybridization. For non-degraded and moderately degraded samples in the series, all three methods were suitably accurate for quantifying nuclear DNA to achieve successful STR amplifications to yield complete profiles using the ABI AmpFlSTR® Identifiler™ kit. However, for highly degraded samples, the duplex qPCR assay provided better estimates of nuclear template for STR amplification than did either the commercial qPCR assay, which overestimated the quantity of STR-sized DNA fragments, leading to an increased proportion of undetected alleles at the larger STR loci, or slot blot hybridization, which underestimated the quantity of nuclear DNA, leading to an increased proportion of STR amplification artifacts due to amplification of excess template.
