5,134 research outputs found
Ultraaccurate genome sequencing and haplotyping of single human cells.
Accurate detection of variants and long-range haplotypes in genomes of single human cells remains very challenging. Common approaches require extensive in vitro amplification of genomes of individual cells using DNA polymerases and high-throughput short-read DNA sequencing. These approaches have two notable drawbacks. First, polymerase replication errors could generate tens of thousands of false-positive calls per genome. Second, relatively short sequence reads contain little to no haplotype information. Here we report a method, which is dubbed SISSOR (single-stranded sequencing using microfluidic reactors), for accurate single-cell genome sequencing and haplotyping. A microfluidic processor is used to separate the Watson and Crick strands of the double-stranded chromosomal DNA in a single cell and to randomly partition megabase-size DNA strands into multiple nanoliter compartments for amplification and construction of barcoded libraries for sequencing. The separation and partitioning of large single-stranded DNA fragments of the homologous chromosome pairs allows for the independent sequencing of each of the complementary and homologous strands. This enables the assembly of long haplotypes and reduction of sequence errors by using the redundant sequence information and haplotype-based error removal. We demonstrated the ability to sequence single-cell genomes with error rates as low as 10-8 and average 500-kb-long DNA fragments that can be assembled into haplotype contigs with N50 greater than 7 Mb. The performance could be further improved with more uniform amplification and more accurate sequence alignment. The ability to obtain accurate genome sequences and haplotype information from single cells will enable applications of genome sequencing for diverse clinical needs
Recommended from our members
The Spitzer C2D Survey of Nearby Dense Cores: Jet and Molecular Outflow Associated With A Young Stellar Object in Core A of L1251
A long infrared jet has been discovered by the Spitzer c2d Legacy Program in core A of L1251. It is associated with a very embedded Class 0 object with an accretion luminosity of about 0.9 L(circle dot) derived by radiative transfer model fitting to the observed spectral energy distribution. Comparing the observed Infrared Array Camera colors along the infrared jet with those calculated from a model of an admixture of gas with a power-law temperature distribution indicates that the jet is possibly created by a paraboloidal bow shock propagating into the ambient medium of n(H(2)) = 10(5) cm(-3). In addition, the variation of the power-law index along the jet suggests that the portion of hot gas decreases with distance from the jet engine. The molecular outflow in this region has been mapped for the first time using CO data. From the calculated outflow momentum flux, a very strong lower limit to the average accretion luminosity is 3.6 sin i/cos(3) i L(circle dot), indicative of a decrease in the accretion rate with time.Korean government (MEST) 2009-0062865NRF R01-2007-000-20336-0NASA 1407, 1224608Astronom
Post-tetanic increase in the fast-releasing synaptic vesicle pool at the expense of the slowly releasing pool
Post-tetanic potentiation (PTP) at the calyx of Held synapse is caused by increases not only in release probability (P(r)) but also in the readily releasable pool size estimated from a cumulative plot of excitatory post-synaptic current amplitudes (RRP(cum)), which contribute to the augmentation phase and the late phase of PTP, respectively. The vesicle pool dynamics underlying the latter has not been investigated, because PTP is abolished by presynaptic whole-cell patch clamp. We found that supplement of recombinant calmodulin to the presynaptic pipette solution rescued the increase in the RRP(cum) after high-frequency stimulation (100 Hz for 4-s duration, HFS), but not the increase in P(r). Release-competent synaptic vesicles (SVs) are heterogeneous in their releasing kinetics. To investigate post-tetanic changes of fast and slowly releasing SV pool (FRP and SRP) sizes, we estimated quantal release rates before and 40 s after HFS using the deconvolution method. After HFS, the FRP size increased by 19.1% and the SRP size decreased by 25.4%, whereas the sum of FRP and SRP sizes did not increase. Similar changes in the RRP were induced by a single long depolarizing pulse (100 ms). The post-tetanic complementary changes of FRP and SRP sizes were abolished by inhibitors of myosin II or myosin light chain kinase. The post-tetanic increase in the FRP size coupled to a decrease in the SRP size provides the first line of evidence for the idea that a slowly releasing SV can be converted to a fast releasing one.Jordan R, 2005, BIOPHYS J, V89, P2091, DOI 10.1529/biophysj.105.061663Kim MH, 2005, J NEUROSCI, V25, P6057, DOI 10.1523/JNEUROSCI.0454-05.2005Korogod N, 2005, J NEUROSCI, V25, P5127, DOI 10.1523/JNEUROSCI.1295-05.2005Moulder KL, 2005, J NEUROSCI, V25, P3842, DOI 10.1523/JINEUROSC.5231-04.2005Habets RLP, 2005, J PHYSIOL-LONDON, V564, P173, DOI 10.1113/jphysiol.2004.079160Felmy F, 2003, NEURON, V37, P801Satzler K, 2002, J NEUROSCI, V22, P10567Wadel K, 2007, NEURON, V53, P563, DOI 10.1016/j.neuron.2007.01.021Wolfel M, 2007, J NEUROSCI, V27, P3198, DOI 10.1523/JNEUROSCI.4471-06.2007Habets RLP, 2007, J PHYSIOL-LONDON, V581, P467, DOI 10.1113/jphysiol.2006.127365Korogod N, 2007, P NATL ACAD SCI USA, V104, P15923, DOI 10.1073/pnas.0704603104Hosoi N, 2007, J NEUROSCI, V27, P14286, DOI 10.1523/JNEUROSCI.4122-07.2007Zikich D, 2008, J NEUROSCI, V28, P1949, DOI 10.1523/JNEUROSCI.5096-07.2008Mongillo G, 2008, SCIENCE, V319, P1543, DOI 10.1126/science.1150769Srinivasan G, 2008, J NEUROPHYSIOL, V99, P1810, DOI 10.1152/jn.00949.2007Lee JS, 2008, J NEUROSCI, V28, P7945, DOI 10.1523/JNEUROSCI.2165-08.2008Awatramani GB, 2005, NEURON, V48, P109, DOI 10.1016/j.neuron.2005.08.038Kushmerick C, 2006, J NEUROSCI, V26, P1366, DOI 10.1523/JNEUROSCI.3889-05.2006Sakaba T, 2006, J NEUROSCI, V26, P5863, DOI 10.1523/JNEUROSCI.0182-06.2006Tokuoka H, 2006, J NEUROSCI, V26, P11606, DOI 10.1523/JNEUROSCI.3400-06.2006Habets RLP, 2006, J NEUROPHYSIOL, V96, P2868, DOI 10.1152/jn.00427.2006Lou XL, 2008, J NEUROSCI, V28, P8257, DOI 10.1523/JNEUROSCI.0550-08.2008Neher E, 2008, NEURON, V59, P861, DOI 10.1016/j.neuron.2008.08.019He LM, 2009, NATURE, V459, P93, DOI 10.1038/nature07860Schneggenburger R, 2002, TRENDS NEUROSCI, V25, P206Tsujimoto T, 2002, SCIENCE, V295, P2276, DOI 10.1126/science.1068278Meinrenken CJ, 2002, J NEUROSCI, V22, P1648Sakaba T, 2001, NEURON, V32, P1119Neher E, 2001, J NEUROSCI, V21, P444Sakaba T, 2001, P NATL ACAD SCI USA, V98, P331Dinkelacker V, 2000, J NEUROSCI, V20, P8377Wu LG, 1999, NEURON, V23, P821Peersen OB, 1997, PROTEIN SCI, V6, P794Borst JGG, 1996, NATURE, V383, P431PUSCH M, 1988, PFLUG ARCH EUR J PHY, V411, P204
Development of a hybrid magnetic resonance/computed tomography-compatible phantom for magnetic resonance guided radiotherapy
The purpose of the present study was to develop a hybrid magnetic resonance/computed tomography (MR/CT)-compatible phantom and tissue-equivalent materials for each MR and CT image. Therefore, the essential requirements necessary for the development of a hybrid MR/CT-compatible phantom were determined and the development process is described. A total of 12 different tissue-equivalent materials for each MR and CT image were developed from chemical components. The uniformity of each sample was calculated. The developed phantom was designed to use 14 plugs that contained various tissue-equivalent materials. Measurement using the developed phantom was performed using a 3.0-T scanner with 32 channels and a Somatom Sensation 64. The maximum percentage difference of the signal intensity (SI) value on MR images after adding K2CO3 was 3.31%. Additionally, the uniformity of each tissue was evaluated by calculating the percent image uniformity (%PIU) of the MR image, which was 82.18 ±1.87% with 83% acceptance, and the average circular-shaped regions of interest (ROIs) on CT images for all samples were within ±5 Hounsfield units (HU). Also, dosimetric evaluation was performed. The percentage differences of each tissue-equivalent sample for average dose ranged from -0.76 to 0.21%. A hybrid MR/CT-compatible phantom for MR and CT was investigated as the first trial in this field of radiation oncology and medical physics
Activation of CD147 with Cyclophilin A Induces the Expression of IFITM1 through ERK and PI3K in THP-1 Cells
CD147, as a receptor for Cyclophilins, is a multifunctional transmembrane glycoprotein. In order to identify genes that are induced by activation of CD147, THP-1 cells were stimulated with Cyclophilin A and differentially expressed genes were detected using PCR-based analysis. Interferon-induced transmembrane 1 (IFITM1) was detected to be induced and it was confirmed by RT-PCR and Western blot analysis. CD147-induced expression of IFITM1 was blocked by inhibitors of ERK, PI3K, or NF-κB, but not by inhibitors of p38, JNK, or PKC. IFITM1 appears to mediate inflammatory activation of THP-1 cells since cross-linking of IFITM1 with specific monoclonal antibody against it induced the expression of proinflammatory mediators such as IL-8 and MMP-9. These data indicate that IFITM1 is one of the pro-inflammatory mediators that are induced by signaling initiated by the activation of CD147 in macrophages and activation of ERK, PI3K, and NF-κB is required for the expression of IFITM1
Donor-site closure using absorbable dermal staple for deep inferior epigastric artery perforator flaps: its efficacy and cosmetic outcomes
Cyclic ADP Ribose-Dependent Ca2+ Release by Group I Metabotropic Glutamate Receptors in Acutely Dissociated Rat Hippocampal Neurons
Group I metabotropic glutamate receptors (group I mGluRs; mGluR1 and mGluR5) exert diverse effects on neuronal and synaptic functions, many of which are regulated by intracellular Ca2+. In this study, we characterized the cellular mechanisms underlying Ca2+ mobilization induced by (RS)-3,5-dihydroxyphenylglycine (DHPG; a specific group I mGluR agonist) in the somata of acutely dissociated rat hippocampal neurons using microfluorometry. We found that DHPG activates mGluR5 to mobilize intracellular Ca2+ from ryanodine-sensitive stores via cyclic adenosine diphosphate ribose (cADPR), while the PLC/IP3 signaling pathway was not involved in Ca2+ mobilization. The application of glutamate, which depolarized the membrane potential by 28.5±4.9 mV (n = 4), led to transient Ca2+ mobilization by mGluR5 and Ca2+ influx through L-type Ca2+ channels. We found no evidence that mGluR5-mediated Ca2+ release and Ca2+ influx through L-type Ca2+ channels interact to generate supralinear Ca2+ transients. Our study provides novel insights into the mechanisms of intracellular Ca2+ mobilization by mGluR5 in the somata of hippocampal neurons
N-(Pyridin-2-ylmethyl)pyridin-2-amine
The title compound, C11H11N3, crystallizes with two molecules (A and B) in the asymmetric unit. The geometries of both molecules are very similar, with the exception of the torsion angles of the inter-ring chains; the values for C—N—C—C are 67.4 (5) and −69.3 (5)° for molecules A and B, respectively. The dihedral angles between the pyridyl ring planes are 84.0 (2) and 83.2 (2)° for molecules A and B, respectively. In the crystal, weak intermolecular N—H⋯N hydrogen bonds and C—H⋯π interactions contribute to the stabilization of the packing
- …