keyword
MENU ▼
Read by QxMD icon Read
search

Cas9

keyword
https://www.readbyqxmd.com/read/28346402/large-scale-design-of-robust-genetic-circuits-with-multiple-inputs-and-outputs-for-mammalian-cells
#1
Benjamin H Weinberg, N T Hang Pham, Leidy D Caraballo, Thomas Lozanoski, Adrien Engel, Swapnil Bhatia, Wilson W Wong
Engineered genetic circuits for mammalian cells often require extensive fine-tuning to perform as intended. We present a robust, general, scalable system, called 'Boolean logic and arithmetic through DNA excision' (BLADE), to engineer genetic circuits with multiple inputs and outputs in mammalian cells with minimal optimization. The reliability of BLADE arises from its reliance on recombinases under the control of a single promoter, which integrates circuit signals on a single transcriptional layer. We used BLADE to build 113 circuits in human embryonic kidney and Jurkat T cells and devised a quantitative, vector-proximity metric to evaluate their performance...
March 27, 2017: Nature Biotechnology
https://www.readbyqxmd.com/read/28346401/targeted-base-editing-in-rice-and-tomato-using-a-crispr-cas9-cytidine-deaminase-fusion
#2
Zenpei Shimatani, Sachiko Kashojiya, Mariko Takayama, Rie Terada, Takayuki Arazoe, Hisaki Ishii, Hiroshi Teramura, Tsuyoshi Yamamoto, Hiroki Komatsu, Kenji Miura, Hiroshi Ezura, Keiji Nishida, Tohru Ariizumi, Akihiko Kondo
We applied a fusion of CRISPR-Cas9 and activation-induced cytidine deaminase (Target-AID) for point mutagenesis at genomic regions specified by single guide RNAs (sgRNAs) in two crop plants. In rice, we induced multiple herbicide-resistance point mutations by multiplexed editing using herbicide selection, while in tomato we generated marker-free plants with homozygous heritable DNA substitutions, demonstrating the feasibility of base editing for crop improvement.
March 27, 2017: Nature Biotechnology
https://www.readbyqxmd.com/read/28345006/genome-wide-specificity-of-highly-efficient-talens-and-crispr-cas9-for-t-cell-receptor-modification
#3
Friederike Knipping, Mark J Osborn, Karl Petri, Jakub Tolar, Hanno Glimm, Christof von Kalle, Manfred Schmidt, Richard Gabriel
In T cells with transgenic high-avidity T cell receptors (TCRs), endogenous and transferred TCR chains compete for surface expression and may pair inappropriately, potentially causing autoimmunity. To knock out endogenous TCR expression, we assembled 12 transcription activator-like effector nucleases (TALENs) and five guide RNAs (gRNAs) from the clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPR-associated (Cas9) system. Using TALEN mRNA, TCR knockout was successful in up to 81% of T cells...
March 17, 2017: Molecular Therapy. Methods & Clinical Development
https://www.readbyqxmd.com/read/28344889/loss-of-tapasin-in-human-lung-and-colon-cancer-cells-and-escape-from-tumor-associated-antigen-specific-ctl-recognition
#4
Yosuke Shionoya, Takayuki Kanaseki, Sho Miyamoto, Serina Tokita, Ayumi Hongo, Yasuhiro Kikuchi, Vitaly Kochin, Kazue Watanabe, Ryota Horibe, Hiroshi Saijo, Tomohide Tsukahara, Yoshihiko Hirohashi, Hiroki Takahashi, Noriyuki Sato, Toshihiko Torigoe
Cytotoxic T-lymphocytes (CTLs) lyse target cells after recognizing the complexes of peptides and MHC class I molecules (pMHC I) on cell surfaces. Tapasin is an essential component of the peptide-loading complex (PLC) and its absence influences the surface repertoire of MHC class I peptides. In the present study, we assessed tapasin expression in 85 primary tumor lesions of non-small cell lung cancer (NSCLC) patients, demonstrating that tapasin expression positively correlated with patient survival. CD8(+) T-cell infiltration of tumor lesions was synergistically observed with tapasin expression and correlated positively with survival...
2017: Oncoimmunology
https://www.readbyqxmd.com/read/28344037/methods-for-decoding-cas9-protospacer-adjacent-motif-pam-sequences-a-brief-overview
#5
REVIEW
Tautvydas Karvelis, Giedrius Gasiunas, Virginijus Siksnys
Recently the Cas9, an RNA guided DNA endonuclease, emerged as a powerful tool for targeted genome manipulations. Cas9 protein can be reprogrammed to cleave, bind or nick any DNA target by simply changing crRNA sequence, however a short nucleotide sequence, termed PAM, is required to initiate crRNA hybridization to the DNA target. PAM sequence is recognized by Cas9 protein and must be determined experimentally for each Cas9 variant. Exploration of Cas9 orthologs could offer a diversity of PAM sequences and novel biochemical properties that may be beneficial for genome editing applications...
March 23, 2017: Methods: a Companion to Methods in Enzymology
https://www.readbyqxmd.com/read/28342760/correction-of-hirschsprung-associated-mutations-in-human-induced-pluripotent-stem-cells-via-crispr-cas9-restores-neural-crest-cell-function
#6
Frank Pui-Ling Lai, Sin-Ting Lau, John Kwong-Leong Wong, Hongsheng Gui, Reeson Xu Wang, Tingwen Zhou, Wing Hon Lai, Hung-Fat Tse, Paul Kwong-Hang Tam, Maria-Mercedes Garcia-Barcelo, Elly Sau-Wai Ngan
BACKGROUND & AIM: Hirschsprung disease is caused by failure of enteric neural crest cells (ENCCs) to fully colonize the bowel, leading to bowel obstruction and megacolon. Heterozygous mutations in the coding region of the RET gene cause a severe form of Hirschsprung disease (total colonic aganglionosis). However, 80% of HSCR patients have short-segment Hirschsprung disease (S-HSCR), which has not been associated with genetic factors. We sought to identify mutations associated S-HSCR, and used the CRISPR/Cas9 gene editing system to determine how mutations affect ENCC function...
March 22, 2017: Gastroenterology
https://www.readbyqxmd.com/read/28342290/verapamil-treatment-induces-cytoprotective-autophagy-by-modulating-cellular-metabolism
#7
Elzbieta Kania, Beata Pajak, Jim O'Prey, Pablo Sierra Gonzalez, Anna Litwiniuk, Kaja Urbańska, Kevin M Ryan, Arkadiusz Orzechowski
Verapamil, an L-type calcium channel blocker, has been used successfully to treat cardiovascular diseases. Interestingly, we have recently shown that treatment of cancer cells with Verapamil causes an effect on autophagy. As autophagy is known to modulate chemotherapy responses, this prompted us to explore the impact of Verapamil on autophagy and cell viability in greater detail. We report here that Verapamil causes an induction of autophagic flux in a number or tumor cells and immortalized normal cells. Moreover, we found that inhibition of autophagy in COLO 205 cells, via treatment with the chloroquine or by CRISPR/Cas9-mediated disruption of the autophagy genes Atg7 and Atg5, causes an upregulation of apoptotic markers in response to Verapamil...
March 25, 2017: FEBS Journal
https://www.readbyqxmd.com/read/28341841/regulation-of-membrane-ruffling-by-polarized-stim1-and-orai1-in-cortactin-rich-domains
#8
Aida M Lopez-Guerrero, Patricia Tomas-Martin, Carlos Pascual-Caro, Thomas Macartney, Alejandro Rojas-Fernandez, Graeme Ball, Dario R Alessi, Eulalia Pozo-Guisado, Francisco Javier Martin-Romero
Cell motility and migration requires the reorganization of the cortical cytoskeleton at the leading edge of cells and extracellular Ca(2+) entry is essential for this reorganization. However the molecular nature of the regulators of this pathway is unknown. This work contributes to understanding the role of STIM1 and ORAI1 in the promotion of membrane ruffling by showing that phospho-STIM1 localizes at the leading edge of cells, and that both phospho-STIM1 and ORAI1 co-localize with cortactin (CTTN), a regulator of the cytoskeleton at membrane ruffling areas...
March 24, 2017: Scientific Reports
https://www.readbyqxmd.com/read/28341547/evaluation-and-rational-design-of-guide-rnas-for-efficient-crispr-cas9-mediated-mutagenesis-in-ciona
#9
Shashank Gandhi, Maximilian Haeussler, Florian Razy-Krajka, Lionel Christiaen, Alberto Stolfi
The CRISPR/Cas9 system has emerged as an important tool for various genome engineering applications. A current obstacle to high throughput applications of CRISPR/Cas9 is the imprecise prediction of highly active single guide RNAs (sgRNAs). We previously implemented the CRISPR/Cas9 system to induce tissue-specific mutations in the tunicate Ciona. In the present study, we designed and tested 83 single guide RNA (sgRNA) vectors targeting 23 genes expressed in the cardiopharyngeal progenitors and surrounding tissues of Ciona embryo...
March 21, 2017: Developmental Biology
https://www.readbyqxmd.com/read/28339613/a-nonsense-mutation-in-ccdc62-gene-is-responsible-for-spermiogenesis-defects-and-male-infertility-in-repro29-repro29-mice%C3%A2
#10
Yuchi Li, Cailing Li, Shouren Lin, Bo Yang, Weiren Huang, Hanwei Wu, Yuanbin Chen, Lihua Yang, Manling Luo, Huan Guo, Jianbo Chen, Tiantian Wang, Qian Ma, Yanli Gu, Lisha Mou, Zhimao Jiang, Jun Xia, Yaoting Gui
Phenotype-driven mutagenesis is an unbiased method to identify novel genes involved in spermatogenesis and other reproductive processes. Male repro29/repro29 mice generated by the Reproductive Genomics Program at the Jackson Laboratory were infertile with deformed sperm and poor motility. Using selected exonic capture and massively parallel sequencing technologies, we identified a nonsense mutation in the exon 6 of coiled-coil domain-containing 62 gene (Ccdc62), which results in a formation of a premature stop codon and a truncated protein...
January 28, 2017: Biology of Reproduction
https://www.readbyqxmd.com/read/28338870/stmyb44-negatively-regulates-phosphate-transport-by-suppressing-expression-of-phosphate1-in-potato
#11
Xiangjun Zhou, Manrong Zha, Jing Huang, Li Li, Muhammad Imran, Cankui Zhang
Phosphorus is an important macronutrient for plant growth, but often deficient in soil. To understand the molecular basis of the complex responses of potato (Solanum tuberosum L.) to phosphate (Pi) deficiency stress, the RNA-Seq approach was taken to identify genes responding to Pi starvation in potato roots. A total of 359 differentially expressed genes were identified, among which the Solanum tuberosum transcription factor gene MYB44 (StMYB44) was found to be down-regulated by Pi starvation. StMYB44 was ubiquitously expressed in potato tissues and organs, and StMYB44 protein was exclusively localized in the nucleus...
March 1, 2017: Journal of Experimental Botany
https://www.readbyqxmd.com/read/28337968/crispr-cas9-mutagenesis-invalidates-a-putative-cancer-dependency-targeted-in-on-going-clinical-trials
#12
Ann Lin, Christopher J Giuliano, Nicole M Sayles, Jason M Sheltzer
The Maternal Embryonic Leucine Zipper Kinase (MELK) has been reported to be a genetic dependency in several cancer types. MELK RNAi and small-molecule inhibitors of MELK block the proliferation of various cancer cell lines, and MELK knockdown has been described as particularly effective against the highly-aggressive basal/triple-negative subtype of breast cancer. Based on these preclinical results, the MELK inhibitor OTS167 is currently being tested as a novel chemotherapy agent in several clinical trials. Here, we report that mutagenizing MELK with CRISPR/Cas9 has no effect on the fitness of basal breast cancer cell lines or cell lines from six other cancer types...
March 24, 2017: ELife
https://www.readbyqxmd.com/read/28337265/pulmonary-surfactant-synthesis-in-mirna-26a-1-mirna-26a-2-double-knockout-mice-generated-using-the-crispr-cas9-system
#13
Ying-Hui Zhang, Li-Zhi Wu, Hong-Lu Liang, Yang Yang, Jie Qiu, Qing Kan, Wen Zhu, Cheng-Ling Ma, Xiao-Yu Zhou
Pulmonary surfactant (PS), which is synthesized by type II alveolar epithelial cells (AECIIs), maintains alveolar integrity by reducing surface tension. Many premature neonates who lack adequate PS are predisposed to developing respiratory distress syndrome (RDS), one of the leading causes of neonatal morbidity and mortality. PS synthesis is influenced and regulated by various factors, including microRNAs. Previous in vitro studies have shown that PS synthesis is regulated by miR-26a in fetal rat AECIIs. This study aimed to investigate the role of miR-26a in PS synthesis in vivo...
2017: American Journal of Translational Research
https://www.readbyqxmd.com/read/28336775/nelf-e-is-recruited-to-dna-double-strand-break-sites-to-promote-transcriptional-repression-and-repair
#14
Samah W Awwad, Enas R Abu-Zhayia, Noga Guttmann-Raviv, Nabieh Ayoub
Double-strand breaks (DSBs) trigger rapid and transient transcription pause to prevent collisions between repair and transcription machineries at damage sites. Little is known about the mechanisms that ensure transcriptional block after DNA damage. Here, we reveal a novel role of the negative elongation factor NELF in blocking transcription activity nearby DSBs. We show that NELF-E and NELF-A are rapidly recruited to DSB sites. Furthermore, NELF-E recruitment and its repressive activity are both required for switching off transcription at DSBs...
March 23, 2017: EMBO Reports
https://www.readbyqxmd.com/read/28335032/allele-specific-ablation-rescues-electrophysiological-abnormalities-in-a-human-ips-cell-model-of-long-qt-syndrome-with-a-calm2-mutation
#15
Yuta Yamamoto, Takeru Makiyama, Takeshi Harita, Kenichi Sasaki, Yimin Wuriyanghai, Mamoru Hayano, Suguru Nishiuchi, Hirohiko Kohjitani, Sayako Hirose, Jiarong Chen, Fumika Yokoi, Taisuke Ishikawa, Seiko Ohno, Kazuhisa Chonabayashi, Hideki Motomura, Yoshinori Yoshida, Minoru Horie, Naomasa Makita, Takeshi Kimura
Background: Calmodulin is a ubiquitous Ca 2+ sensor molecule encoded by three distinct calmodulin genes, CALM1-3 . Recently, mutations in CALM1-3 have been reported to be associated with severe early-onset long-QT syndrome (LQTS). However, the underlying mechanism through which heterozygous calmodulin mutations lead to severe LQTS remains unknown, particularly in human cardiomyocytes. Objectives: We aimed to establish an LQTS disease model associated with a CALM2 mutation (LQT15) using human induced pluripotent stem cells (hiPSCs) and to assess mutant allele-specific ablation by genome editing for the treatment of LQT15...
March 1, 2017: Human Molecular Genetics
https://www.readbyqxmd.com/read/28334928/functional-assessment-of-ctcf-sites-at-cytokine-sensing-mammary-enhancers-using-crispr-cas9-gene-editing-in-mice
#16
Hye Kyung Lee, Michaela Willi, Chaochen Wang, Chul Min Yang, Harold E Smith, Chengyu Liu, Lothar Hennighausen
The zinc finger protein CTCF has been invoked in establishing boundaries between genes, thereby controlling spatial and temporal enhancer activities. However, there is limited genetic evidence to support the concept that these boundaries restrict the search space of enhancers. We have addressed this question in the casein locus containing five mammary and two non-mammary genes under the control of at least seven putative enhancers. We have identified two CTCF binding sites flanking the locus and two associated with a super-enhancer...
March 16, 2017: Nucleic Acids Research
https://www.readbyqxmd.com/read/28334828/flexible-crispr-library-construction-using-parallel-oligonucleotide-retrieval
#17
Abigail Read, Shaojian Gao, Eric Batchelor, Ji Luo
CRISPR/Cas9-based gene knockout libraries have emerged as a powerful tool for functional screens. We present here a set of pre-designed human and mouse sgRNA sequences that are optimized for both high on-target potency and low off-target effect. To maximize the chance of target gene inactivation, sgRNAs were curated to target both 5΄ constitutive exons and exons that encode conserved protein domains. We describe here a robust and cost-effective method to construct multiple small sized CRISPR library from a single oligo pool generated by array synthesis using parallel oligonucleotide retrieval...
March 16, 2017: Nucleic Acids Research
https://www.readbyqxmd.com/read/28334779/targeted-gene-knock-in-by-homology-directed-genome-editing-using-cas9-ribonucleoprotein-and-aav-donor-delivery
#18
Thomas Gaj, Brett T Staahl, Gonçalo M C Rodrigues, Prajit Limsirichai, Freja K Ekman, Jennifer A Doudna, David V Schaffer
Realizing the full potential of genome editing requires the development of efficient and broadly applicable methods for delivering programmable nucleases and donor templates for homology-directed repair (HDR). The RNA-guided Cas9 endonuclease can be introduced into cells as a purified protein in complex with a single guide RNA (sgRNA). Such ribonucleoproteins (RNPs) can facilitate the high-fidelity introduction of single-base substitutions via HDR following co-delivery with a single-stranded DNA oligonucleotide...
March 2, 2017: Nucleic Acids Research
https://www.readbyqxmd.com/read/28334759/highly-efficient-biallelic-genome-editing-of-human-es-ips-cells-using-a-crispr-cas9-or-talen-system
#19
Kazuo Takayama, Keisuke Igai, Yasuko Hagihara, Rina Hashimoto, Morifumi Hanawa, Tetsushi Sakuma, Masashi Tachibana, Fuminori Sakurai, Takashi Yamamoto, Hiroyuki Mizuguchi
Genome editing research of human ES/iPS cells has been accelerated by clustered regularly interspaced short palindromic repeats/CRISPR-associated 9 (CRISPR/Cas9) and transcription activator-like effector nucleases (TALEN) technologies. However, the efficiency of biallelic genetic engineering in transcriptionally inactive genes is still low, unlike that in transcriptionally active genes. To enhance the biallelic homologous recombination efficiency in human ES/iPS cells, we performed screenings of accessorial genes and compounds...
February 21, 2017: Nucleic Acids Research
https://www.readbyqxmd.com/read/28334154/ethical-issues-of-crispr-technology-and-gene-editing-through-the-lens-of-solidarity
#20
John J Mulvihill, Benjamin Capps, Yann Joly, Tamra Lysaght, Hub A E Zwart, Ruth Chadwick
Background: The avalanche of commentaries on CRISPR-Cas9 technology, a bacterial immune system modified to recognize any short DNA sequence, cut it out, and insert a new one, has rekindled hopes for gene therapy and other applications and raised criticisms of engineering genes in future generations. Sources of data: This discussion draws on articles that emphasize ethics, identified partly through PubMed and Google, 2014-2016. Areas of agreement: CRISPR-Cas9 has taken the pace and prospects for genetic discovery and applications to a high level, stoking anticipation for somatic gene engineering to help patients...
February 23, 2017: British Medical Bulletin
keyword
keyword
163
1
2
Fetch more papers »
Fetching more papers... Fetching...
Read by QxMD. Sign in or create an account to discover new knowledge that matter to you.
Remove bar
Read by QxMD icon Read
×

Search Tips

Use Boolean operators: AND/OR

diabetic AND foot
diabetes OR diabetic

Exclude a word using the 'minus' sign

Virchow -triad

Use Parentheses

water AND (cup OR glass)

Add an asterisk (*) at end of a word to include word stems

Neuro* will search for Neurology, Neuroscientist, Neurological, and so on

Use quotes to search for an exact phrase

"primary prevention of cancer"
(heart or cardiac or cardio*) AND arrest -"American Heart Association"