We narrowed to 1,039 results for: LAS

...cells in a 75 cm 2 flask in a volume of 15 mL. Wednesday: Plate 1x10 6 cells in a T75 flask in a volume of...therefore, plasmid DNA purification should include an endotoxin removal step. For high quality plasmid DNA, ...protocol to transfect mammalian cells with your plasmid of interest. Protocols...harvest cells, or perform your experiment Equipment Class II, Type A2 Biological Safety Cabinet 0.5–10 µL ...of 15 mL. Friday: Plate 8x10 5 cells in a T75 flask in a volume of 15 mL. Do not add antibiotics to the..., the plasmid should also be propagated in an endonuclease negative E. coli strain such as NEB stable....is prepared, transfect cells with a fluorescent plasmid using a variety of ratios. Check the cells 1–2 ...

...Agar Plates Bacterial Transformation Recovering Plasmid DNA from Bacterial Culture Introduction If you ...or stab culture of bacteria and need to purify plasmid DNA from it, you will want to isolate an individual...will minimize the chance of having a mixture of plasmids in your purified DNA. This protocol explains how...bacterial colony by streaking it onto an LB agar plate. Last Update: Feb. 28, 2017 Protocol Video Watch the protocol...antibiotic. Label the bottom of the plate with the plasmid name and the date. It is also a good idea to add...through streak #2 and spread the bacteria over the last section of the plate, to create streak #3. Incubate...overnight (12-18 hours) at 37 °C. Pro-Tip Some plasmids or bacteria need to be grown at 30 °C instead ...

...also like... Plasmids 101 Blog Posts Mol Bio Protocols Cloning Protocols Introduction Plasmids can carry ...resistance gene on a plasmid allows researchers to easily isolate bacteria containing that plasmid from bacteria...resistance to that antibiotic - usually conferred by a plasmid carrying the antibiotic resistance gene. The following...LB/agar plates with your antibiotic of interest. Last Upload: September 14, 2016 Protocol Video Before...which can hold 10 - 15 mL of agar. Autoclavable flasks Sterile pipettes Ice bucket to hold antibiotic ...effective at selecting only bacteria containing the plasmids of interest (for example, fewer satellite colonies...at the right temperature, we recommend using a laser thermometer. Light the flame at the plate pouring...

...Design your plasmid and order primers (see figure to the right). When designing your plasmid, think about...correct plasmid product by restriction digest . Sequence the important regions of your final plasmid, particularly...efficient and effective method for the assembly of plasmids, and molecular biologists now use this method ...segments you will need to join to create your final plasmid. Adjacent segments should have identical sequences... background and enrich for correctly assembled plasmids by designing primers to split an antibiotic resistance...reaction if you're only making a small change in a plasmid (such as point mutations). Generate DNA segments...

...transduced with lentiCas9-Blast 1 and then selected with 1 μg/mL blasticidin for 9 days. Single cells ...lentiCas9-Blast 1 . A549 cells were transduced (MOI = 37) and selected with 1 μg/mL blasticidin for 9 days...loading control. 1 lentiCas9-Blast was a gift from Feng Zhang (Addgene plasmid #52962 ) and is described ...and expand monoclonal lines of interest Equipment Class II, Type A2 Biological Safety Cabinet 0.5–10 µL ...then expanded for an additional 3 weeks under blasticidin selection. Anti-Cas9 Western blotting was performed...2014 Aug;11(8):783-4. (Link opens in a new window) Last reviewed on: September 9, 2023...

...follows: Remove the gel from the plastic packaging. Remove tape and plastic comb. Rinse the wells with deionized...Blot Western Blot You may also like... Antibody Plasmid Collection Antibody Blog Posts Antibody Guide Immunocytochemistry...adapted for wet and semi-dry transfer approaches. Last Update: January 24, 2022 Workflow Timeline Day 1...well microtiter plate Chemiluminescence substrate Plastic wrap Primary antibody Secondary antibody Deionized... white separator. Keep the Bottom Stack in the plastic tray. Place the Bottom Stack on the blotting surface...for 5 min at RT . Cover the membrane with clear plastic and use a gel imager with chemiluminescence detection...

...Agar Plates Bacterial Transformation Recovering Plasmid DNA from Bacterial Culture Introduction This protocol... plate and select transformed cells containing plasmids differing in their resistance genes, as one does...curve to determine optimal antibiotic concentration. Last Update: Oct. 27, 2017 Protocol Video Watch the protocol...or other antibiotic) E. coli transformed with a plasmid containing the carbenicillin (amp) resistance gene...effective at selecting only bacteria containing the plasmids of interest (for example, fewer satellite colonies...transform DH5α E. coli cells by heatshock with the plasmid of interest. See our transformation page for a ...

...Storing and Handling Addgene Plasmids What to do after you receive your plasmids from Addgene Instructions...Protocol Diagnostic Restriction Digest Verify a plasmid using restriction enzymes Diagnostic Restriction...Instructions for Storing and Handling Plasmids Genomic Deletions with CRISPR Generate genomic deletions in mammalian...Transformations We walk you through a standard plasmid transformation protocol and offer some tips and...Center How to Deposit Your guide to depositing plasmids with Addgene Depositing Page Career Videos Get...

...Electrophoresis You may also like... Restriction Digest of Plasmid DNA Purifying DNA from an Agarose Gel DNA Ligation...a collection of DNA fragments of known lengths). Last Update: Feb. 20, 2018 Protocol Video Watch the protocol...agarose powder with 100 mL 1xTAE in a microwavable flask. See TAE Recipe . Pro-Tip TBE can be used instead...people prefer to microwave in pulses, swirling the flask occasionally as the solution heats up.). Caution...boil over. Placing saran wrap over the top of the flask can help with this, but is not necessary if you ... when you can comfortably keep your hand on the flask), about 5 mins. Optional : Add ethidium bromide ...

...station if unwanted or biohazardous materials are splashed onto your face and/or get into your eyes. Wash...your skin, including, but not limited to broken glass or needles. Sharps containers must be thick walled...shaking or mixing, which may produce aerosols or splashes. These procedures should be done underneath a ...Working in a BSL-2 laboratory requires laboratory glasses in addition to coat and gloves. This lab coat should...here has been designed by the Addgene nonprofit plasmid repository and is being shared outside of Addgene...

...Recombinant Antibodies You may also like... Antibody Plasmid Collection Antibody Blog Posts Antibody Guide Transfection... intended to support reproducibility in science. Last Update: June 14, 2023 Workflow Timeline Day 1: Run...Rocking platform Fume hood Metal spatula Razor blade Plastic tray Gel imaging system ImageJ or similar software...in the microcentrifuge. Remove the gel from the plastic wrapper and rinse with deionized water. Gently ...Section 2: Staining the Gel Place the gel in a plastic tray with 100 mL of deionized water. Rinse gel ...

...1) and biosafety level 2 (BSL-2). The BSL-1 classification is for labs working with low-risk microbes ...not be possible depending on the type of sleeve. Lastly, don’t touch your face! Close-toed shoes and long... be used as needed. For BSL-2 work always wear glasses/goggles in addition to the BSL-1 requirements. ...here has been designed by the Addgene nonprofit plasmid repository and is being shared outside of Addgene...

...good idea to place the gel on a glass plate if available. Unlike the plastic tray, this will not significantly...Gel You may also like... Restriction Digest of Plasmid DNA Agarose Gel Electrophoresis DNA Ligation Background...for your eyes!), remove it from any gel tray as plastic will block much of the UV and with a clean, sterile...

...media Day 4–5: Count fluorescent cells Equipment Class II, Type A2 Biological Safety Cabinet 0.5–10 µL ...obtaining accurate titers. Check the cells for mycoplasma (Link opens in a new window) regularly Do not...mCherry expression using fluorescence microscopy. Last reviewed on: September 9, 2023...

...Immunocytochemistry You may also like... Antibody Plasmid Collection Antibody Blog Posts Antibody Guide Western... intended to support reproducibility in science. Last Update: January 20, 2022 Workflow Timeline Day 1... the protein, such as cells transfected with a plasmid to express the protein of interest, and a negative...

...Stain cells and count resistant colonies Equipment Class II, Type A2 Biological Safety Cabinet 0.5–10 µL ...obtaining accurate titers. Check the cells for mycoplasma (Link opens in a new window) regularly Do not...single colonies are visible in the higher dilutions. Last reviewed on: August 31, 2023...

...PCR) You may also like... Plasmid Cloning by PCR Restriction Digest of Plasmid DNA DNA Ligation Background...detailed instructions and check out our protocol on plasmid cloning by PCR to learn how to design primers for...

...Protein A or Protein G You may also like... Antibody Plasmid Collection Antibody Blog Posts Antibody Guide Transfection...following your institution’s hazardous waste procedures. Last Update: February 7, 2022 Protocol Video Watch this...antibody Day 2 or later: Buffer exchange Equipment Class II, Type A2 Biological Safety Cabinet 4 °C Refrigerator...

...compliance with your institution’s biosafety regulations. Last Update: July 7, 2023 Workflow Timeline Day 1: Seed...harvest cells Day 4+: ddPCR and analysis Equipment Class II, Type A2 Biological Safety Cabinet Aspirating...green) and negative (black) droplets Sample ID Plasmid ID Dilution Copies/20 µL RRE (FAM) Copies/20 µL...

...Culture Recovering Plasmid DNA from Bacterial Culture Restriction Digest of Plasmid DNA Background Information...