ultramer rna oligo standard  (Integrated DNA Technologies)

Journal: Viruses

Article Title: Targeted Chromatinization and Repression of HIV-1 Provirus Transcription with Repurposed CRISPR/Cas9

doi: 10.3390/v12101154

Figure Lengend Snippet: Guide RNA (gRNA) sequence characteristics.

Article Snippet: Three hundred nM of Ultramer RNA Oligos gRNA constructs were used per condition using program X-005 on Nucleofector 2b Device (Lonza, Portsmouth, NH, USA).

Techniques: Sequencing, Activity Assay

Journal: Viruses

Article Title: Targeted Chromatinization and Repression of HIV-1 Provirus Transcription with Repurposed CRISPR/Cas9

doi: 10.3390/v12101154

Figure Lengend Snippet: Disabled Cas9 (dCas9) constructs.

Article Snippet: Three hundred nM of Ultramer RNA Oligos gRNA constructs were used per condition using program X-005 on Nucleofector 2b Device (Lonza, Portsmouth, NH, USA).

Techniques: Construct, Plasmid Preparation, Control

Journal: Viruses

Article Title: Targeted Chromatinization and Repression of HIV-1 Provirus Transcription with Repurposed CRISPR/Cas9

doi: 10.3390/v12101154

Figure Lengend Snippet: Identification of HIV-1 guide RNAs (gRNAs) that mediate disabled Cas9 (dCas9)- Kruppel-associated box (KRAB) repression. ( A ) Cartoon demonstrating locations targeted by gRNAs. ( B ) Repression of HIV transcription by gRNAs. HEK293T cells were cotransfected with NL4-3-Δ env -Luciferase, dCas9-KRAB, and the indicated guide RNAs. Negative controls included transfections with gRNAs only (black bars). HIV expression was monitored after 48 h by luciferase activity, which was normalized to control gRNA. Data represent averages of multiple experiments, with each guide being assessed in at least three independent experiments. Error bars represent standard error and Student t -test were performed to determine significance. * and ** indicate p ≤ 0.05 and p ≤ 0.01, respectively.

Article Snippet: Three hundred nM of Ultramer RNA Oligos gRNA constructs were used per condition using program X-005 on Nucleofector 2b Device (Lonza, Portsmouth, NH, USA).

Techniques: Luciferase, Transfection, Expressing, Activity Assay, Control

Journal: Viruses

Article Title: Targeted Chromatinization and Repression of HIV-1 Provirus Transcription with Repurposed CRISPR/Cas9

doi: 10.3390/v12101154

Figure Lengend Snippet: dCas9-KRAB inhibits reactivation of latent proviral HIV. J-Lat 6.3 cells that stably express dsaCas9-KRAB were electroporated with short RNA gRNA targeting LTR (397) or scrambled control. Cells were activated with PMA + Ionomycin, and HIV expression was monitored by qRT-PCR. These data include results from five independent experiments. Error bars represent standard error, and Student t-test was performed to determine significance. ** indicates p ≤ 0.01.

Article Snippet: Three hundred nM of Ultramer RNA Oligos gRNA constructs were used per condition using program X-005 on Nucleofector 2b Device (Lonza, Portsmouth, NH, USA).

Techniques: Stable Transfection, Control, Expressing, Quantitative RT-PCR

Journal: Viruses

Article Title: Targeted Chromatinization and Repression of HIV-1 Provirus Transcription with Repurposed CRISPR/Cas9

doi: 10.3390/v12101154

Figure Lengend Snippet: dCas9-KRAB mediates post-translational modification of histones that correlate with transcriptional repression. HEK 293T cells were infected with pseudotyped NL4-3-env-Luciferase and 24 hours later transfected in duplicate with dCas9-KRAB construct and gRNA targeting the HIV LTR (397) or a scrambled control. Forty-eight hours post transfection, ChIPs were performed using anti-acH3 and anti-H3K9me3 antibodies. Associated HIV-1 LTR sequences were detected by PCR. These data represent a single experiment of 3 independent experiments (see ). Error bars represent standard deviation, and a student t -test was performed for statistical significance. **** indicates p < 0.001.

Article Snippet: Three hundred nM of Ultramer RNA Oligos gRNA constructs were used per condition using program X-005 on Nucleofector 2b Device (Lonza, Portsmouth, NH, USA).

Techniques: Modification, Infection, Luciferase, Transfection, Construct, Control, Standard Deviation