...using a sequencing machine that recognizes the different fluorescent labels. The machine detects which fluorescently...templates, are spatially separated from each other. These attached DNA templates are then amplified again...again producing ~1,000 copies of each template. Each template is then replicated using the modified bases...fluorescent color that is emitted each time a base is added. Again, each base (A,C,T, or G) is labelled ...an organism. Proteins are made up of amino acids. Each amino acid is encoded for by 3 nucleotides termed...20 natural amino acids and 64 codon combinations each amino acid is encoded for by multiple codons. Plasmids...fragments, which is far too many to separate from each other for the purposes of isolating one specific...

...applications common to each category, including an outline of the basic steps needed for each application, important...different isotypes for each primary antibody and secondary antibodies specific to each isotype. Overview of... data. Check that your machine has appropriate lasers and filters for each fluorophore you select. Suggested... itself prior to antibody production; therefore, each B cell only produces one type of antibody. Monocultures...enable higher production of a single antibody, but each B cell can only produce one variable region. Isotypes...-antibody units, while others exist as monomers. Each antibody class therefore has a valency, or number...typically the number of antibodies multiplied by two, as each antibody has two binding sites. Isotype Description...

...specificity. Dual-nickase approach increases specificity but is less efficient. Each putative knockout allele...engineering . Prior to CRISPR, genome engineering approaches like zinc finger nucleases (ZFNs) or transcription-activator-like...rapidly become the most popular genome engineering approach. Engineered CRISPR systems contain two components...experiment, you might consider using the dual nickase approach to create a double nick-induced DSB. The nickase...have been engineered using cell-based selection approaches. Yeast-based screening systems resulted in evolved...termed left & right homology arms .) The length of each homology arm is dependent on the size of the change...several gRNAs from the same plasmid ensures that each cell containing the plasmid will express all of ...

...) until peak expression of the opsin is reached. This approach is especially powerful when combined with...including a brief overview of the wild-type opsin of each and some examples of relevant variants. This list...closing of the channel 470(activ.), 590(inact.) ReaChR Red-activatable variant of CrChR2 590 Channelrhodopsins... variant 467 Aurora Red-shifted anion-selective ReaChR variant 517 Halorhodopsins Jaws Red-shifted, light-driven...Generally, the most robust and stable expression is achieved in a transgenic mouse line, such as for example...that bind to the LOV domain only in the dark. attaching only one member of the Zdk/LOV2 pair to the target...KR, Deisseroth K. 2010 Molecular and cellular approaches for diversifying and extending optogenetics. ...

...move around pieces of DNA. Oftentimes several approaches will work for any specific cloning project; however...likely that for any given project there is an ideal approach. This may be due to speed, cost, availability ... of time required for cloning. To utilize this approach, the fragment of DNA that you would like to clone...amplified with specific Gateway attB1 and attB2 sites attached to the 5’ and 3’ ends of DNA sequence. This fragment...fragments generating long overhangs that anneal to each other due to their homology. DNA polymerase then... enzyme cloning. Scientists have utilized this approach to create compatible custom overhangs that can...

...cDNA libraries, each plasmid contains a unique cDNA. In shRNA or gRNA libraries, each plasmid contains...sequence, but there are multiple sequences targeting each gene in the overall library. Barcoding libraries...negative results - for a given plasmid, think of each cell carrying it as being a biological replicate...

...engineered α7 nicotinic acetylcholine receptor (nAChR) domains that respond to specific small molecules...Adeno-associated virus (AAV) is a widely-used tool for achieving in vivo expression of chemogenetic receptors, ... our AAV guide . FLEx Vectors are also used to achieve cell-specific expression of AAV-encoded chemogenetic...Azua IR, Li JH, et al (2009). A chemical-genetic approach to study G protein regulation of β cell function...

...produce AAV in the lab. Addgene’s AAV Guide will teach you about AAV components, serotypes, pseudotyping...molecular weight structures that are maintained extrachromosomally. rAAV are more likely than wild-type AAV ...the virus depends on the cell’s DNA replication machinery to synthesize the complementary strand, transgene...

...creating replication-competent virus increases with each round of amplification. What is RCA? RCA stands ... probability of a crossover event increases with each round of amplification. Where can I find information...

...plasmids can be used in transient transfections to achieve expression of the transgene, and those that can...the insert from a parent vector, some possible approaches to using this site include subcloning or appending...

...guide to viral vectors contains an introduction to each vector and its applications, tips for using vectors...

...RNA polymerase in binding to the promoter region. Each sigma factor recognizes different core promoter ...