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Kim, Heon Seok,Lee, Kyungjin,Bae, Sangsu,Park, Jeongbin,Lee, Chong-Kyo,Kim, Meehyein,Kim, Eunji,Kim, Minju,Kim, Seokjoong,Kim, Chonsaeng,Kim, Jin-Soo American Society for Biochemistry and Molecular Bi 2017 The Journal of biological chemistry Vol.292 No.25
<P>Several groups have used genome-wide libraries of lentiviruses encoding small guide RNAs (sgRNAs) for genetic screens. In most cases, sgRNA expression cassettes are integrated into cells by using lentiviruses, and target genes are statistically estimated by the readout of sgRNA sequences after targeted sequencing. We present a new virus-free method for human gene knockout screens using a genome-wide library of CRISPR/Cas9 sgRNAs based on plasmids and target gene identification via whole-genome sequencing (WGS) confirmation of authentic mutations rather than statistical estimation through targeted amplicon sequencing. We used 30,840 pairs of individually synthesized oligonucleotides to construct the genome-scale sgRNA library, collectively targeting 10,280 human genes (<I>i.e.</I> three sgRNAs per gene). These plasmid libraries were co-transfected with a Cas9-expression plasmid into human cells, which were then treated with cytotoxic drugs or viruses. Only cells lacking key factors essential for cytotoxic drug metabolism or viral infection were able to survive. Genomic DNA isolated from cells that survived these challenges was subjected to WGS to directly identify CRISPR/Cas9-mediated causal mutations essential for cell survival. With this approach, we were able to identify known and novel genes essential for viral infection in human cells. We propose that genome-wide sgRNA screens based on plasmids coupled with WGS are powerful tools for forward genetics studies and drug target discovery.</P>
Digenome-seq: genome-wide profiling of CRISPR-Cas9 off-target effects in human cells
Kim, Daesik,Bae, Sangsu,Park, Jeongbin,Kim, Eunji,Kim, Seokjoong,Yu, Hye Ryeong,Hwang, Jinha,Kim, Jong-Il,Kim, Jin-Soo Nature Publishing Group, a division of Macmillan P 2015 NATURE METHODS Vol.12 No.3
Although RNA-guided genome editing via the CRISPR-Cas9 system is now widely used in biomedical research, genome-wide target specificities of Cas9 nucleases remain controversial. Here we present Digenome-seq, in vitro Cas9-digested whole-genome sequencing, to profile genome-wide Cas9 off-target effects in human cells. This in vitro digest yields sequence reads with the same 5′ ends at cleavage sites that can be computationally identified. We validated off-target sites at which insertions or deletions were induced with frequencies below 0.1%, near the detection limit of targeted deep sequencing. We also showed that Cas9 nucleases can be highly specific, inducing off-target mutations at merely several, rather than thousands of, sites in the entire genome and that Cas9 off-target effects can be avoided by replacing 'promiscuous' single guide RNAs (sgRNAs) with modified sgRNAs. Digenome-seq is a robust, sensitive, unbiased and cost-effective method for profiling genome-wide off-target effects of programmable nucleases including Cas9.
Energetics of Z-DNA Binding Protein-Mediated Helicity Reversals in DNA, RNA, and DNA–RNA Duplexes
Bae, Sangsu,Kim, Yuyoung,Kim, Doyoun,Kim, Kyeong Kyu,Kim, Yang-Gyun,Hohng, Sungchul American Chemical Society 2013 The journal of physical chemistry. B, Condensed ma Vol.117 No.44
<P>Z-DNA binding proteins (ZBPs) specifically recognize and stabilize left-handed double helices, including Z-DNA and Z-RNA. However, the energetics of Z-form stabilization by ZBPs have never been characterized due to the technical limitations of bulk studies, resulting in an unclear understanding of the ZBP operational mechanism at the molecular level. Here, we use single-molecule fluorescence resonance energy transfer (FRET) to determine the energetics of Z-form stabilization by ZBP for DNA, RNA, and DNA–RNA duplexes, revealing that the formation of B–Z or A–Z junctions dominates the thermodynamics and kinetics of Z-form stabilization. Furthermore, in contrast to general assumptions, the Z-form is most efficiently and most rapidly formed in the DNA–RNA hybrid duplex due to the greatly reduced junction energy in the DNA–RNA hybrid.</P><P><B>Graphic Abstract</B> <IMG SRC='http://pubs.acs.org/appl/literatum/publisher/achs/journals/content/jpcbfk/2013/jpcbfk.2013.117.issue-44/jp409862j/production/images/medium/jp-2013-09862j_0006.gif'></P><P><A href='http://pubs.acs.org/doi/suppl/10.1021/jp409862j'>ACS Electronic Supporting Info</A></P>
Characteristics of Ballistic Tansport in Short-Channel MOSFETs
Nambin KIM,Hyungsang KIM,Hyunjung KIM,임현식,Sangsu PARK,Yongmin KIM 한국물리학회 2004 THE JOURNAL OF THE KOREAN PHYSICAL SOCIETY Vol.45 No.3
The Si MOSFET is the single most important unit of any digital or analog circuit today. As it is scaled down to the nanometer region, charge carriers (electrons and holes) in the channel are expected to ow with fewer scattering events, leading to ballistic transport. In this work, the characteristics of the quasi-ballistic MOSFET are systematically investigated by taking into account scattering phenomena of the carriers. For a practical viewpoint, a conventional MOSFET structure is exploited. Numerical calculations of potential prole are carried out, in order to understand and quantify the transport as functions of gate and drain voltages under various conditions.
A New Record of Proreus simulans (Dermaptera: Chelisochidae) in Korea
Kim, Taewoo,Kim, Sangsu The Korean Society of Systematic Zoology 2019 Animal Systematics, Evolution and Diversity Vol.35 No.1
The earwig Proreus simulans($St{\aa}l$, 1860) has been recently reported from Jeollanam-do in the far southern section of the Korean Peninsula. Natural photographs of this species were obtained from Gwangyang Province for the first time in 2013, with specimens being physically collected in light traps from Suncheon Province in 2018. The species is widely distributed throughout the Oriental region, where its main habitat is lowland paddy fields. The earwig belongs to family Chelisochidae, subfamily Chelisochinae, and genus Proreus, which are newly recognized in Korea. The diversity of the order Dermaptera is currently estimated to be 6 families, 14 genera, and 24 species in Korea.
Experimental Study on Freezing Phenomena of Water Saturated Square Cavity with Inclined Cold Surface
Kim, Jongjun,An, Sangsu,Kim, Byungchul CHOSUN UNIVERSITY 1997 Basic Science and Engineering Vol.1 No.1
It was studied the phenomena of transient freezing of an inclined water-saturated enclosure. One side of the test section was cooled and the other sides were insulated. The effects of the initial temperature, the inclination angle on the temperature field and the shape of the ice-water interface were observed. In the beginning f freezing, when inclination angle increased, the freezing rate was increased and when the water in test section formed the more stable density layer, freezing was faster because the convective fluid flow became small. When the initial temperature was above the 4℃, the frozen thickness in the upper part of inclined surface was thinner than that of the lower part, but with time the frozen thickness of upper part was thicker than that of lower part, below the 4℃, the frozen thickness in the upper part was thicker than that of lower part from the begining, and above the 8℃, upper part was thinner with concave in the beginning.