...phosphodiester linkages, which permanently join the nucleotides together. After ligation, the insert DNA is physically...

... on a rocking platform. (Optional) Counterstain nuclei with 500 µL of 300 nM DAPI working solution for...

...genetic code for all life is based upon four nucleotides, 64 codons, and 20 amino acids. Yet in the past...

...the plasmids should also be propagated in an endonuclease negative E. coli strain such as NEB stable. ...

...use a HEK293 line stable expressing Epstein Barr nuclear antigen (EBNA). Safety Warnings HEK293 cells are...

...multiple guide RNA expression cassettes and a Cas9 nuclease/nickase expression cassette using the Golden Gate...

... hard to extract proteins such as those in the nucleus. Incubate on ice for 30 min . Centrifuge lysate...

...Reverse ITR Primer (10 µM) 1.8 µL 16.2 µL 900 nM Nuclease-free water 5.9 µL 71.55 µL Total Volume 20 µL ...

... ~18 mL from steps 12 & 13). Benzonase is an endonuclease that will degrade any residual DNA carried over...

...like zinc finger nucleases (ZFNs) or transcription-activator-like effector nucleases (TALENs) required...popular Cas endonuclease, with many modified versions engineered to increase specificity and nuclease activity...increase nuclease fidelity SpCas9-NG - NG; increase in vitro activity SpG - NGN; increase nuclease activity...break; the wild type function of most Cas nucleases dCas9 Nuclease dead Cas9, an enzymatically inactive form...required scientists to design and generate a new nuclease pair for every genomic target. Due to its comparative...RNA ( gRNA or sgRNA ) and a CRISPR-associated endonuclease (Cas enzyme). The gRNA is a short synthetic ...necessary for Cas-binding and a user-defined ∼20-nucleotide spacer that defines the genomic target to be ...

... transcribed from DNA. DNA is comprised of 4 nucleotides or bases, adenine, thymine, cytosine, and guanine... double stranded helix. The order of these 4 nucleotides makes up the genetic code and provides the instructions...amino acids. Each amino acid is encoded for by 3 nucleotides termed a codon. As there are only 20 natural ...CRISPR technology. CRISPR is composed of a DNA endonuclease and guide RNAs that target specific locations...sequence DNA” to identify the order of these four nucleotide bases in a particular DNA strand. Sequencing ...migrates down the single stranded DNA adding nucleotides according to the sequence of the complementary...interest. In vitro DNA replication requires the 4 nucleotides, a DNA polymerase enzyme, the template DNA to...

...molecular biology enzymes: 5' exonuclease, polymerase and ligase. 5' exonuclease digests the 5' end of dsDNA...properties of type IIS restriction endonucleases. These endonucleases cut dsDNA at a specified distance...) relies on the 3'-5' exonuclease activity of T4 DNA polymerase. An exonuclease is an enzyme which removes...removes nucleotides from the end of a DNA strand. In LIC, the T4 DNA polymerase’s exonuclease activity ...Restriction Enzyme Cloning Restriction enzyme (endonuclease) based molecular cloning is the "classic" cloning...polymerases synthesize DNA molecules using the 4 nucleotides and lastly DNA ligase fuses DNA strands together...together in one isothermal reaction. First the 5’ exonuclease chews back the 5’ ends of your DNA fragments ...

...are only able to enter the nucleus during mitotic breakdown of the nuclear envelope. Lentiviruses can ...componentes are small enough to enter the nucleus through nuclear pore complex. This feature may be useful...retroviral particle containing Matrix, Capsid, and Nucleocapsid components. Pol in trans Precursor protein containing... (Ψ) in cis RNA target site for packaging by Nucleocapsid. WPRE in cis Woodchuck hepatitis virus post‐...stimulates the expression of transgenes via increased nuclear export. LTR Subcomponents: U3 R U5 in cis LTR; ...

...off-target editing is a major concern, Cas9-gRNA ribonucleoprotein (RNP) complexes are advantageous due to the... to generate transgenic embryos Cas9-gRNA ribonucleoprotein (RNP) complexes Purified Cas9 protein and ...increase or decrease depending upon the specific nucleotides within the selected target sequence. For example...example, gRNA targeting sequences containing a G nucleotide at position 20 (1 bp upstream of the PAM) may...be more efficacious than gRNAs containing a C nucleotide at the same position in spite of being a perfect...products are denatured-renatured, treated with a nuclease that cleaves DNA heteroduplexes, and run on an...restriction digest (for HDR repaired DSBs): For small nucleotide edits that introduce a novel restriction site...

...Ther. 2005. 5(4): 387-398. PubMed . HIV-1 Genome Nuclear Import Is Mediated by a Central DNA Flap. Zennou...lentiviral particle containing Matrix, Capsid, and Nucleocapsid components. Pol in trans Precursor protein containing...and partially spliced transcripts to facilitate nuclear export. Tat in trans Second generation only; Trans-activator... (Ψ) in cis RNA target site for packaging by Nucleocapsid. RRE in cis Rev Response Element; sequence to...stimulates the expression of transgenes via increased nuclear export. LTR Sub-components: U3 R Sub-element: TAR...

...genetically-encoded nuclear Cry2 fusion can thus direct a CIB1-protein fusion to the nucleus upon light activiation...voltage (LOV) domain of FKF1 LINUS light-inducible nuclear localization signal based on the LOV2 domain of...photosensitive kinase, which uses the ubiquitous flavin mononucleotide as its chromophore, and phosphorylates the ...

...interactions between DNA and protein. Micrococcal nuclease digestion is used to shear the DNA in these assays...specific to your assay. Most IF techniques use a nuclear stain, such as DAPI, to identify individual cells...cells. Immunofluorescence experiments often use a nuclei marker to identify individual cells and additional... PE, and APC 4-color microscopy panel: DAPI for nuclei, AF488, AF555, and AF647 Antibody Selection Selecting...

...found in eukaryotic cells that package DNA into nucleosomes. Histone binding prevents the initiation of transcription...Pribnow box which usually consists of the six nucleotides, TATAAT. Proximal Promoter Further upstream from...response element promoter U6 Constitutive Human U6 nuclear promoter for small RNA expression UAS Specific ...

...transfection of suspension-growing human 293-EBNA1 cells. Nucleic Acids Res . 30:e9. PMID: 11788735 (Link opens in...

...septicum alpha-toxin , and resistance to to the nucleotide analog 6-thioguanine . Next-generation sequencing...