...For small deletions (as the “deletion band” may still amplify), resolve agarose gel to determine if size...for 20 min). These cycling conditions allow for digestion and ligation to occur in one reaction (see step...conditions for each primer pair designed based on testing the bulk sorted cells. Run samples on 2% agarose...

...later. At a minimum we recommend the use of a diagnostic digest and high-throughput next generation sequencing...plasmid backbone a single time and visualize that digestion on an agarose gel ( see restriction digest protocol... Transform, recover, set up overnight growth (Estimated time 2-3 hours) Transformation should be performed...limited. Day 2: Harvest cells and purify DNA (Estimated time 3-4 hours) Cells should be harvested first...

...Knockout Human Yusa 3rd 5 90,709 Human Interferon-Stimulated Gene CRISPR Knockout Library 125753 Knockout ...Mutation Libraries 229137–229140 Donor Vector Human Stiewe N/A Varies 1,910–3,034 per exon Human Cas13 lncRNA...69,716 SAM v2 - 2 plasmid system 1000000078 (Blasticidin) Activation Human Zhang 3rd 3 70,290 CHyMErA ...

...Fluorescent Protein Resources: Empty Backbones Allen Institute for Cell Science Plasmid Collection FRET Biosensors...-pmCherryC1 Actin Filaments Cortactin mCherry Christien Merrifield 36201 pLifeAct-mTurquoise2 Actin Filaments...Protein Collection . Return to top Do you have suggestions for other plasmids that should be added to this...

...desired restriction sites and add them to your existing vector. It is a couple days of work that will ...the overhangs generated by restriction digest of existing sites in the original vector. Designing oligos...bases to complement the overhangs generated when digesting the vector with EcoRI and SalI (see diagram )....

...Equipment Water Bath 10% bleach or 70% ethanol Distilled water Disinfectant Thermometer Water bath weights...the edge of the slide. Fill the water bath with distilled water, not tap water, to prevent salts from accumulating...water in it from previous uses. However, you will still need to regularly clean out the water bath even ...

...partnership to provide AAV viral preps, includes listing of transfer status for viral preps....Discontinued Penn Vector Core Viral Vectors (Plasmids still available, other serotypes may be available) Based...Wilson Please contact [email protected] with any questions. Return to top...

...percentage of fluorescent cells, these cells can still be used in downstream applications. Safety Warnings...ensure that you are in compliance with your institution’s biosafety regulations. Workflow Timeline Day...with multiple integration events leading to underestimation of the true titer. Calculate the transduction...

...Reactivity Recommended Applications PI Do you have suggestions for other antibodies that should be added to ... represent the official views of the National Institutes of Health....

...- Davidson Allen Institute for Brain Science AAV Enhancer Collection Allen Institute for Cell Science ...

...mammalian cells or zebrafish (can be used as a substitute for pMLM335 described in Foley et al., PLoS One...mammalian cells or zebrafish (can be used as a substitute for pMLM336 described in Foley et al., PLoS One...

...genome-containing virions. Phenol red is added to clearly distinguish the steps. This method will help enrich the prep...ultracentrifugation. The steps are clearly distinguishable thanks to the use of phenol red. Image adapted...cartoon indicating the position of the needle for harvesting of the purified AAV using option #1. Right image...

...phytochrome photoreceptors (BphPs). Spectrally distinct permanently fluorescent iRFP variants have been...photoactivation) 3.0 pPAiRFP2-N1 Do you have suggestions for other plasmids that should be added to this...

...on how it was originally used. Here are a few questions to consider when choosing a gRNA: Does a given...Insert Vector Type PI Publication Do you have suggestions for other plasmids that should be added to this...

... and Cascade on one single plasmid, though many still separate the individual Cas components. Figure 1...Last reviewed on: January 30, 2025 Do you have suggestions for other plasmids that should be added to this...

...Brzezinski Lab CRISPR Collection The Brzezinski lab investigates gene regulation in the context of the developing...cloning to insert the desired guide RNA sequence by digestion of the backbone with BbsI followed by a quick ...

...proteins (FPs) in a variety of plasmids allows for constitutive overexpression of FPs and tagging of genes of...Tags Publication FP Overexpression Plasmids for constitutive overexpression of codon-optimized FPs in yeast...

...please let us know using the Cloning Grade DNA Suggestion Form . Please note that this does not guarantee...us of which plasmids would be most helpful to requesting scientists. Return to top...

...viral vector production is a common step after requesting plasmids from Addgene, and by providing this ...storage. Biosafety Guide Documentation for your institutional biosafety office and your lab, information on...

... bath Pipetman Reagents 37 g pre-mixed powder consisting of: 5.0 g yeast extract 10.0 g peptone from casein...mL of agar and on which you can individually distinguish a maximum of ~100 colonies Pro-Tip If you will...After overnight drying, we place the plates in a plastic bag with an absorbent material to reduce condensation...