Design and Generation of Drosophila Single Guide RNA Expression Constructs

2016 ◽  
Vol 2016 (9) ◽  
pp. pdb.prot090779 ◽  
Author(s):  
Benjamin E. Housden ◽  
Yanhui Hu ◽  
Norbert Perrimon
Keyword(s):  
Rna Expression ◽  
Guide Rna

scholarly journals CRISPR/Cas9 with single guide RNA expression driven by small tRNA promoters showed reduced editing efficiency compared to a U6 promoter

RNA ◽  
2016 ◽  
Vol 23 (1) ◽  
pp. 1-5 ◽  
Author(s):  
Yuda Wei ◽  
Yan Qiu ◽  
Yanhao Chen ◽  
Gaigai Liu ◽  
Yongxian Zhang ◽  
...  
Keyword(s):  
Rna Expression ◽  
Editing Efficiency ◽  
Guide Rna ◽  
U6 Promoter

scholarly journals Decoupling tRNA promoter and processing activities enables specific Pol-II Cas9 guide RNA expression

2018 ◽  
Author(s):  
David JHF Knapp ◽  
Yale S Michaels ◽  
Max Jamilly ◽  
Quentin RV Ferry ◽  
Hector Barbosa ◽  
...  
Keyword(s):  
Rational Design ◽  
Rna Expression ◽  
Temporal Control ◽  
High Background ◽  
Pol Ii ◽  
Trna Processing ◽  
Guide Rna ◽  
Promising Strategy ◽  
Screening Platform

ABSTRACTSpatial/temporal control of Cas9 guide RNA expression could considerably expand the utility of CRISPR-based technologies. Current approaches based on tRNA processing offer a promising strategy but suffer from high background. Here we developed a variant screening platform to identify differential sequence determinants of human tRNA promoter and processing activities. Rational design based on the ensuing principles allowed us to engineer an improved tRNA scaffold that enabled highly specific guide RNA production from a Pol-II promoter.

scholarly journals Highly multiplex guide RNA expression units of CRISPR/Cas9 were completely stable using cosmid amplification in a novel polygonal structure

2019 ◽  
Vol 21 (11) ◽  
Author(s):  
Tomoko Nakanishi ◽  
Aya Maekawa ◽  
Hirotaka Tabata ◽  
Takashi Yoshioka ◽  
Zheng Pei ◽  
...  
Keyword(s):  
Rna Expression ◽  
Guide Rna ◽  
Polygonal Structure

scholarly journals A multiplex guide RNA expression system and its efficacy for plant genome engineering

Plant Methods ◽  
2020 ◽  
Vol 16 (1) ◽  
Author(s):  
Youngbin Oh ◽  
Bora Lee ◽  
Hyeonjin Kim ◽  
Sang-Gyu Kim
Keyword(s):  
Genome Engineering ◽  
Expression System ◽  
Plant Genome ◽  
Rna Expression ◽  
Guide Rna

scholarly journals Decoupling tRNA promoter and processing activities enables specific Pol-II Cas9 guide RNA expression

2019 ◽  
Vol 10 (1) ◽  
Author(s):  
David J. H. F. Knapp ◽  
Yale S. Michaels ◽  
Max Jamilly ◽  
Quentin R. V. Ferry ◽  
Hector Barbosa ◽  
...  
Keyword(s):  
Rna Expression ◽  
Pol Ii ◽  
Guide Rna

scholarly journals Inhibition of CRISPR/Cas9-Mediated Genome Engineering by a Type I Interferon-Induced Reduction in Guide RNA Expression

2017 ◽  
Vol 40 (3) ◽  
pp. 272-277 ◽  
Author(s):  
Mitsuhiro Machitani ◽  
Fuminori Sakurai ◽  
Keisaku Wakabayashi ◽  
Kosuke Nakatani ◽  
Kazuo Takayama ◽  
...  
Keyword(s):  
Genome Engineering ◽  
Type I Interferon ◽  
Rna Expression ◽  
Type I ◽  
Guide Rna

5S rRNA Promoter for Guide RNA Expression Enabled Highly Efficient CRISPR/Cas9 Genome Editing in Aspergillus niger

2018 ◽  
Vol 8 (7) ◽  
pp. 1568-1574 ◽  
Author(s):  
Xiaomei Zheng ◽  
Ping Zheng ◽  
Kun Zhang ◽  
Timothy C. Cairns ◽  
Vera Meyer ◽  
...  
Keyword(s):  
Aspergillus Niger ◽  
Genome Editing ◽  
5S Rrna ◽  
Rna Expression ◽  
Guide Rna ◽  
Highly Efficient

scholarly journals CRISPR/Cas13 effectors have differing extents of off-target effects that limit their utility in eukaryotic cells

2021 ◽  
Author(s):  
Yuxi Ai ◽  
Dongming Liang ◽  
Jeremy E Wilusz
Keyword(s):  
Circular Rna ◽  
Eukaryotic Cells ◽  
Rna Expression ◽  
Collateral Damage ◽  
Cell Type ◽  
Guide Rna ◽  
Perfect Complementarity ◽  
Rna Knockdown ◽  
Target Rna ◽  
Target Effects

CRISPR/Cas13 effectors have garnered increasing attention as easily customizable tools for detecting and depleting RNAs of interest. Near perfect complementarity between a target RNA and the Cas13-associated guide RNA is required for activation of Cas13 ribonuclease activity. Nonetheless, the specificity of Cas13 effectors in eukaryotic cells has been debated as the Cas13 nuclease domains can be exposed on the enzyme surface, providing the potential for promiscuous cleavage of nearby RNAs (so-called collateral damage). Here, using co-transfection assays in Drosophila and human cells, we found that the off-target effects of RxCas13d, a commonly used Cas13 effector, can be as strong as the level of on-target RNA knockdown. The extent of off-target effects is positively correlated with target RNA expression levels, and collateral damage can be observed even after reducing RxCas13d/guide RNA levels. The PspCas13b effector showed improved specificity and, unlike RxCas13d, can be used to deplete a Drosophila circular RNA without affecting the expression of the associated linear RNA. PspCas13b nonetheless still can have off-target effects and we notably found that the extent of off-target effects for Cas13 effectors differs depending on the cell type and target RNA examined. In total, these results highlight the need for caution when designing and interpreting Cas13-based knockdown experiments.

Long non-coding RNA expression profiles in human parathyroid tumors

2017 ◽  
Author(s):  
Annamaria Morotti ◽  
Irene Forno ◽  
Valentina Andre ◽  
Andrea Terrasi ◽  
Chiara Verdelli ◽  
...  
Keyword(s):  
Expression Profiles ◽  
Rna Expression ◽  
Parathyroid Tumors ◽  
Non Coding Rna ◽  
Long Non Coding Rna

RNA expression as a prognostic tool in idiopathic nephrotic syndrome

2007 ◽  
Vol 148 (23) ◽  
pp. 1067-1075
Author(s):  
Krisztina Fischer ◽  
Orsolya Galamb ◽  
Béla Molnár ◽  
Zsolt Tulassay ◽  
András Szabó
Keyword(s):  
Nephrotic Syndrome ◽  
Northern Blot ◽  
Idiopathic Nephrotic Syndrome ◽  
Minimal Change ◽  
Ribonuclease Protection Assay ◽  
Rna Expression ◽  
Rt Pcr ◽  
Protection Assay ◽  
Ribonuclease Protection

A gyermekkori nephrosis 90%-a idiopathiás nephrosis szindróma. Az idetartozó három kórkép, a minimal change betegség, a mesangialis proliferatio és a focalis sclerosis hasonló klinikai képpel jelentkező, eltérő prognózisú és terápiás válaszú betegség. Dolgozatunk célja az idiopathiás nephrosis szindrómába tartozó kórképek kialakulásával, progressziójával összefüggő genetikai ismeretek, génexpressziós változások áttekintése és funkcionális csoportosítása. A génexpressziós változások meghatározásának eszközeként, dolgozatunk röviden összefoglalja a northern blot, a ribonuclease protection assay, az in situ RNS-hibridizáció, a kvantitatív RT-PCR és a microarray módszerek lényegét. Az eddig elvégzett vizsgálatok a DNS-szintézis és repair gének, növekedési faktorok, extracelluláris mátrix, extracelluláris ligandreceptorok, extracelluláris jelátvitel zavarai mellett kiemelik a metabolikus és transzporter gének, illetve az immunszabályozó gének molekuláris eltéréseit, amelyek összefüggésben vannak az idiopathiás nephrosis szindróma eddig megismert molekuláris hátterével. A chiptechnológia fejlődésével és elterjedésével ezek a markerek és a hagyományos vizsgálati módszerek párhuzamos alkalmazása rutindiagnosztikai szempontból is fontossá válhat.

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