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CRISPR CAS9 human

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https://www.readbyqxmd.com/read/29932473/detailed-analysis-of-htt-repeat-elements-in-human-blood-using-targeted-amplification-free-long-read-sequencing
#1
Ida Höijer, Yu-Chih Tsai, Tyson A Clark, Paul Kotturi, Niklas Dahl, Eva-Lena Stattin, Marie-Louise Bondeson, Lars Feuk, Ulf Gyllensten, Adam Ameur
Amplification of DNA is required as a mandatory step during library preparation in most targeted sequencing protocols. This can be a critical limitation when targeting regions that are highly repetitive or with extreme guanine-cytosine (GC) content, including repeat expansions associated with human disease. Here we used an amplification-free protocol for targeted enrichment utilizing the CRISPR/Cas9 system (No-Amp Targeted sequencing) in combination with Single Molecule, Real-Time (SMRT) sequencing for studying repeat elements in the huntingtin (HTT) gene, where an expanded CAG repeat is causative for Huntington disease...
June 22, 2018: Human Mutation
https://www.readbyqxmd.com/read/29928486/novel-fluorinated-carbonic-anhydrase-ix-inhibitors-reduce-hypoxia-induced-acidification-and-clonogenic-survival-of-cancer-cells
#2
Justina Kazokaitė, Raymon Niemans, Virginija Dudutienė, Holger M Becker, Jānis Leitāns, Asta Zubrienė, Lina Baranauskienė, Gabor Gondi, Reinhard Zeidler, Jurgita Matulienė, Kaspars Tārs, Ala Yaromina, Philippe Lambin, Ludwig J Dubois, Daumantas Matulis
Human carbonic anhydrase (CA) IX has emerged as a promising anticancer target and a diagnostic biomarker for solid hypoxic tumors. Novel fluorinated CA IX inhibitors exhibited up to 50 pM affinity towards the recombinant human CA IX, selectivity over other CAs, and direct binding to Zn(II) in the active site of CA IX inducing novel conformational changes as determined by X-ray crystallography. Mass spectrometric gas-analysis confirmed the CA IX-based mechanism of the inhibitors in a CRISPR/Cas9-mediated CA IX knockout in HeLa cells...
June 1, 2018: Oncotarget
https://www.readbyqxmd.com/read/29925663/the-role-of-mxb-in-interferon-%C3%AE-mediated-inhibition-of-hiv-1-infection
#3
Bin Xu, Qinghua Pan, Chen Liang
Type I interferon inhibits viruses through inducing the expression of antiviral proteins, including the myxovirus resistance (Mx) proteins. Compared to human MxA protein that inhibits a wide range of viruses, the MxB protein has been reported to specifically inhibit primate lentiviruses including HIV-1, and herpesviruses. Further, the role of endogenous MxB in interferon-α mediated inhibition of HIV-1 infection was questioned by a recent study showing that MxB knockout did not increase the infection of HIV-1 which carried the G protein of vesicular stomatitis virus (VSV), and thus was able to infect CD4-negative HT1080 cells...
June 20, 2018: Journal of Virology
https://www.readbyqxmd.com/read/29925504/ldha-in-neuroblastoma-is-associated-with-poor-outcome-and-its-depletion-decreases-neuroblastoma-growth-independent-of-aerobic-glycolysis
#4
Carmen Dorneburg, Matthias Fischer, Thomas F E Barth, Wolfgang Mueller-Klieser, Barbara Hero, Judith Gecht, Daniel R Carter, Katleen De Preter, Benjamin Mayer, Lisa Christner, Frank Speleman, Glenn M Marshall, Klaus-Michael Debatin, Christian Beltinger
PURPOSE: To investigate whether lactate dehydrogenase A (LDHA), an important component of the LDH tetramer crucial for aerobic glycolysis, is associated with patient outcome and constitutes a therapeutic target in neuroblastoma (NB). EXPERIMENTAL DESIGN: Expression of LDHA mRNA and protein was determined in 709 and 110 NB patient samples, respectively, and correlated to survival and risk factors. LDHA and LDHB were depleted in human NB cell lines by CRISPR/Cas9 and shRNA, respectively, and aerobic glycolysis, clonogenicity and tumorigenicity were determined...
June 20, 2018: Clinical Cancer Research: An Official Journal of the American Association for Cancer Research
https://www.readbyqxmd.com/read/29925039/a-splice-junction-targeted-crispr-approach-spjcrispr-reveals-human-foxo3b-to-be-a-protein-coding-gene
#5
Evan E Santo, Jihye Paik
The rapid development of CRISPR technology is revolutionizing molecular approaches to the dissection of complex biological phenomena. Here we describe an alternative generally applicable implementation of the CRISPR-Cas9 system that allows for selective knockdown of extremely homologous genes. This strategy employs the lentiviral delivery of paired sgRNAs and nickase Cas9 (Cas9D10A) to achieve targeted deletion of splice junctions. This general strategy offers several advantages over standard single-guide exon-targeting CRISPR-Cas9 such as greatly reduced off-target effects, more restricted genomic editing, routine disruption of target gene mRNA expression and the ability to differentiate between closely related genes...
June 17, 2018: Gene
https://www.readbyqxmd.com/read/29924937/reversing-mechanoinductive-desmoplakin-dsp-expression-by-crispr-dcas9-mediated-epigenome-editing
#6
Jing Qu, Lanyan Zhu, Zijing Zhou, Ping Chen, Shuyan Liu, Morgan L Locy, Victor J Thannickal, Yong Zhou
RATIONALE: Desmoplakin (DSP), the most abundant component of desmosomes which maintain the mechanical integrity of epithelium, is a GWAS-identified genetic risk locus in human idiopathic pulmonary fibrosis (IPF). IPF subjects express a significantly higher level of DSP than control subjects. OBJECTIVES: Determine potential mechanisms by which DSP is regulated in lung fibrosis Methods: Matrigel-coated soft and stiff polyacrylamide gels were made to simulate the stiffness of normal and fibrotic lungs...
June 20, 2018: American Journal of Respiratory and Critical Care Medicine
https://www.readbyqxmd.com/read/29924933/straightforward-delivery-of-linearized-double-stranded-dna-encoding-sgrna-and-donor-dna-for-the-generation-of-single-nucleotide-variants-based-on-the-crispr-cas9-system
#7
Soyeong Jun, Hyeonseob Lim, Hoon Jang, Wookjae Lee, Jinwoo Ahn, Ji Hyun Lee, Duhee Bang
CRISPR/Cas9 for genome editing requires delivery of a guide RNA sequence and donor DNA for targeted homologous recombination. Typically, single-stranded oligodeoxynucleotide, serving as the donor template, and a plasmid encoding guide RNA are delivered as two separate components. However, in the multiplexed generation of single nucleotide variants, this two-component delivery system is limited by difficulty of delivering a matched pair of sgRNA and donor DNA to the target cell. Here, we describe a novel co-delivery system called "sgR-DNA" that uses a linearized double-stranded DNA consisting of donor DNA component and a component encoding sgRNA...
June 20, 2018: ACS Synthetic Biology
https://www.readbyqxmd.com/read/29923822/investigation-of-the-role-of-gbf1-in-the-replication-of-positive-sense-single-stranded-rna-viruses
#8
Juliette Ferlin, Rayan Farhat, Sandrine Belouzard, Laurence Cocquerel, Antoine Bertin, Didier Hober, Jean Dubuisson, Yves Rouillé
GBF1 has emerged as a host factor required for the replication of positive-sense single-stranded RNA viruses of different families, but its mechanism of action is still unknown. GBF1 is a guanine nucleotide exchange factor for Arf family members. Recently, we identified Arf4 and Arf5 (class II Arfs) as host factors required for the replication of hepatitis C virus (HCV), a GBF1-dependent virus. To assess whether a GBF1/class II Arf pathway is conserved among positive-sense single-stranded RNA viruses, we investigated yellow fever virus (YFV), Sindbis virus (SINV), coxsackievirus B4 (CVB4) and human coronavirus 229E (HCoV-229E)...
June 20, 2018: Journal of General Virology
https://www.readbyqxmd.com/read/29921768/compound-k-induces-endoplasmic-reticulum-stress-and-apoptosis-in-human-liver-cancer-cells-by-regulating-stat3
#9
Xuan Zhang, Silin Zhang, Qitong Sun, Wenjun Jiao, Yan Yan, Xuewu Zhang
The ginsenoside compound K (20- O -β-d-glucopyranosyl-20( S )-protopanaxadiol; CK) is an intestinal bacterial metabolite of ginseng protopanaxadiol saponin that has been reported to induce apoptosis in many cancer cells; however, the precise mechanisms of its activity in human hepatocellular carcinoma (HCC) cells remain unclear. Herein, we demonstrated that CK inhibited the growth and colony formation of HepG2 and SMMC-7721 cells, phenotypes that were mediated by inducing apoptosis. Meanwhile, CK showed lower toxicity in normal hepatoma cells...
June 19, 2018: Molecules: a Journal of Synthetic Chemistry and Natural Product Chemistry
https://www.readbyqxmd.com/read/29917197/crispr-technology-new-paradigm-to-target-the-infectious-disease-pathogens
#10
M A Bakhrebah, M S Nassar, M S Alsuabeyl, W A Zaher, S A Meo
OBJECTIVE: Infectious diseases are one of the prime causes of death worldwide. An innovative sequence specific editing technology "Clustered Regularly Interspaced Short Palindromic Repeats (CRISPR)" has been tested on a broad range of microorganisms to target and destroy invading foreign DNA to human cells or tissues. This study aimed to discuss the mechanism and therapeutic usage of CRISPR/Cas9 genome editing technology in the management of various infectious disease pathogens...
June 2018: European Review for Medical and Pharmacological Sciences
https://www.readbyqxmd.com/read/29915997/synthetic-biology-advances-and-applications-in-the-biotechnology-industry-a-perspective
#11
Leonard Katz, Yvonne Y Chen, Ramon Gonzalez, Todd C Peterson, Huimin Zhao, Richard H Baltz
Synthetic biology is a logical extension of what has been called recombinant DNA (rDNA) technology or genetic engineering since the 1970s. As rDNA technology has been the driver for the development of a thriving biotechnology industry today, starting with the commercialization of biosynthetic human insulin in the early 1980s, synthetic biology has the potential to take the industry to new heights in the coming years. Synthetic biology advances have been driven by dramatic cost reductions in DNA sequencing and DNA synthesis; by the development of sophisticated tools for genome editing, such as CRISPR/Cas9; and by advances in informatics, computational tools, and infrastructure to facilitate and scale analysis and design...
June 18, 2018: Journal of Industrial Microbiology & Biotechnology
https://www.readbyqxmd.com/read/29914939/transcriptional-up-regulation-of-bag3-a-chaperone-assisted-selective-autophagy-factor-in-animal-models-of-ky-deficient-hereditary-myopathy
#12
Elliot J Jokl, Gideon L Hughes, Tobias Cracknell, Mary E Pownall, Gonzalo Blanco
The importance of kyphoscoliosis peptidase (KY) in skeletal muscle physiology has recently been emphasised by the identification of novel human myopathies associated with KY deficiency. Neither the pathogenic mechanism of KY deficiency nor a specific role for KY in muscle function have been established. However, aberrant localisation of FLNC in muscle fibers has been shown in humans and mice with loss of function mutations in the KY gene. FLNC turnover has been proposed to be controlled by Chaperone Assisted Selective Autophagy (CASA), a client-specific and tension-induced pathway that is required for muscle maintenance...
June 18, 2018: Disease Models & Mechanisms
https://www.readbyqxmd.com/read/29914921/determining-the-pathogenicity-of-a-genomic-variant-of-uncertain-significance-using-crispr-cas9-and-human-induced-pluripotent-stem-cells
#13
Ning Ma, Joe Zhang, Ilanit Itzhaki, Sophia L Zhang, Haodong Chen, Francois Haddad, Tomoya Kitani, Kitchener D Wilson, Lei Tian, Rajani Shrestha, Haodi Wu, Chi Keung Lam, Nazish Sayed, Joseph C Wu
Background -The progression toward low-cost and rapid next-generation sequencing has uncovered a multitude of variants of uncertain significance (VUS) in both patients and asymptomatic "healthy" individuals. A VUS is a rare or novel variant for which disease pathogenicity has not been conclusively demonstrated or excluded, and thus cannot be definitively annotated. VUS, therefore, pose critical clinical interpretation and risk-assessment challenges, and new methods are urgently needed to better characterize their pathogenicity...
June 18, 2018: Circulation
https://www.readbyqxmd.com/read/29908711/gene-editing-on-center-stage
#14
REVIEW
Rasmus O Bak, Natalia Gomez-Ospina, Matthew H Porteus
Smithies et al. (1985) and Jasin and colleagues (1994) provided proof of concept that homologous recombination (HR) could be applied to the treatment of human disease and that its efficiency could be improved by the induction of double-strand breaks (DSBs). A key advance was the discovery of engineered nucleases, such as zinc-finger nucleases (ZFNs) and transcription activator-like (TAL) effector nucleases (TALENs), that can generate site-specific DSBs. The democratization and widespread use of genome editing was enabled by the discovery of the clustered regularly interspaced short palindromic repeats (CRISPR)-Cas9 nuclease system...
June 13, 2018: Trends in Genetics: TIG
https://www.readbyqxmd.com/read/29907766/efficient-rna-drug-delivery-using-red-blood-cell-extracellular-vesicles
#15
Waqas Muhammad Usman, Tin Chanh Pham, Yuk Yan Kwok, Luyen Tien Vu, Victor Ma, Boya Peng, Yuen San Chan, Likun Wei, Siew Mei Chin, Ajijur Azad, Alex Bai-Liang He, Anskar Y H Leung, Mengsu Yang, Ng Shyh-Chang, William C Cho, Jiahai Shi, Minh T N Le
Most of the current methods for programmable RNA drug therapies are unsuitable for the clinic due to low uptake efficiency and high cytotoxicity. Extracellular vesicles (EVs) could solve these problems because they represent a natural mode of intercellular communication. However, current cellular sources for EV production are limited in availability and safety in terms of horizontal gene transfer. One potentially ideal source could be human red blood cells (RBCs). Group O-RBCs can be used as universal donors for large-scale EV production since they are readily available in blood banks and they are devoid of DNA...
June 15, 2018: Nature Communications
https://www.readbyqxmd.com/read/29906414/guide-rna-selection-for-crispr-cas9-transfections-in-plasmodium-falciparum
#16
Jose M Ribeiro, Meera Garriga, Nicole Potchen, Anna K Crater, Ankit Gupta, Daisuke Ito, Sanjay A Desai
CRISPR-Cas9 mediated genome editing is addressing key limitations in the transfection of malaria parasites. While this method has already simplified the needed molecular cloning and reduced the time required to generate mutants in the human pathogen Plasmodium falciparum, optimal selection of required guide RNAs and guidelines for successful transfections have not been well characterized, leading workers to use time-consuming trial and error approaches. We used a genome-wide computational approach to create a comprehensive and publicly accessible database of possible guide RNA sequences in the P...
June 12, 2018: International Journal for Parasitology
https://www.readbyqxmd.com/read/29905858/optimized-knock-in-of-point-mutations-in-zebrafish-using-crispr-cas9
#17
Sergey V Prykhozhij, Charlotte Fuller, Shelby L Steele, Chansey J Veinotte, Babak Razaghi, Johane M Robitaille, Christopher R McMaster, Adam Shlien, David Malkin, Jason N Berman
We have optimized point mutation knock-ins into zebrafish genomic sites using clustered regularly interspaced palindromic repeats (CRISPR)/Cas9 reagents and single-stranded oligodeoxynucleotides. The efficiency of knock-ins was assessed by a novel application of allele-specific polymerase chain reaction and confirmed by high-throughput sequencing. Anti-sense asymmetric oligo design was found to be the most successful optimization strategy. However, cut site proximity to the mutation and phosphorothioate oligo modifications also greatly improved knock-in efficiency...
June 14, 2018: Nucleic Acids Research
https://www.readbyqxmd.com/read/29905762/leptodb-an-integrated-database-of-genomics-and-proteomics-resource-of-leptospira
#18
Shruti Beriwal, Nikhil Padhiyar, Deven Bhatt, Prabhakar D Pandit, Afzal Ansari, Kumari Snehkant Lata, Zuber M Saiyed, Vibhisha Vaghasia, Priyanka Sharma, Shivarudrappa B Bhairappanavar, Subhash Soni, Jayashankar Das
Leptospirosis is a potentially fatal zoo-anthroponosis caused by pathogenic species of Leptospira belonging to the family of Leptospiraceae, with a worldwide distribution and effect, in terms of its burden and risk to human health. The 'LeptoDB' is a single window dedicated architecture (5 948 311 entries), modeled using heterogeneous data as a core resource for global Leptospira species. LeptoDB facilitates well-structured knowledge of genomics, proteomics and therapeutic aspects with more than 500 assemblies including 17 complete and 496 draft genomes encoding 1...
January 1, 2018: Database: the Journal of Biological Databases and Curation
https://www.readbyqxmd.com/read/29904383/influenza-a-virus-facilitates-its-infectivity-by-activating-p53-to-inhibit-the-expression-of-interferon-induced-transmembrane-proteins
#19
Bei Wang, Tze Hau Lam, Mun Kuen Soh, Zhiyong Ye, Jinmiao Chen, Ee Chee Ren
Human influenza virus (IAV) are among the most common pathogens to cause human respiratory infections. A better understanding on interplay between IAV and host factors may provide clues for disease prevention and control. While many viruses are known to downregulate p53 upon entering the cell to reduce the innate host antiviral response, IAV infection is unusual in that it activates p53. However, it has not been clear whether this process has proviral or antiviral effects. In this study, using human isogenic p53 wild-type and p53null A549 cells generated from the CRISPR/Cas9 technology, we observed that p53null cells exhibit significantly reduced viral propagation when infected with influenza A virus (strain A/Puerto Rico/8/1934 H1N1)...
2018: Frontiers in Immunology
https://www.readbyqxmd.com/read/29903769/yap-tyrosine-phosphorylation-and-nuclear-localization-in-cholangiocarcinoma-cells-is-regulated-by-lck-and-independent-of-lats-activity
#20
Takaaki Sugihara, Nathan W Werneburg, Matthew C Hernandez, Lin Yang, Ayano Kabashima, Petra Hirsova, Lavanya Yohanathan, Carlos Sosa, Mark Joseph Truty, George Vasmatzis, Gregory J Gores, Rory L Smoot
The hippo pathway effector, Yes-associated protein (YAP) is a transcriptional co-activator implicated in cholangiocarcinoma (CCA) pathogenesis. YAP is known to be regulated by a serine/threonine kinase relay module (MST1/2 - LATS1/2) culminating in phosphorylation of YAP at Serine 127 (S127) and cytoplasmic sequestration. However, YAP also undergoes tyrosine phosphorylation, and the role of tyrosine phosphorylation in YAP regulation remains unclear. Herein, YAP regulation by tyrosine phosphorylation was examined in human and mouse CCA cells, as well as patient-derived xenograft (PDX) models...
June 14, 2018: Molecular Cancer Research: MCR
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