Skip to main content
Addgene

Enter a sequence to perform a BLAST-based search of Addgene plasmid sequence databases.

In Category

We narrowed to 46 results for: form

Showing: 11 - 20 of 46 results

  1. Pouring LB Agar Plates

    Type
    Protocol
    ...our case 220 mL) to the same bottle and swirl to form a medium/agar colloid. Cover the opening of the ...lab. The addition of agar to LB results in the formation of a gel that bacteria can grow on, as they are...

  2. Isolating a Monoclonal Cell Population by Limiting Dilution

    Type
    Protocol
    ...and left undisturbed for 13 days. (a, b) Colonies formed in individual wells of a 96-well plate after expansion...monoclonal cell lines. When this protocol was performed at Addgene on the A549 cells described in our ...expression and/or other phenotypes. For example, perform Western blotting to screen for lines with the highest...blasticidin selection. Anti-Cas9 Western blotting was performed on whole-cell extracts from the initial stable...

  3. Protocol - pLKO.1 – TRC Cloning Vector

    Type
    Protocol
    ... is stable at 4°C for up to one year. The powder form of polybrene is stable at 4°C for several years....provided for your convenience. See “warranty information” in appendix. Table of Contents A. pLKO.1-TRC...Digesting pLKO.1 TRC-Cloning Vector C.4 Ligating and transforming into bacteria D. Screening for Inserts D.1 Recommended... TRC-Cloning Vector I.2 Recipes I.3 Warranty information Back to Top A. pLKO.1-TRC Cloning Vector A.1 ...pCMV-VSVG from Robert Weinberg’s lab. For more information, visit Addgene’s Mammalian RNAi Tools page. Several...Measure the DNA concentration. C.4 Ligating and Transforming into Bacteria Use your ligation method of choice...final volume Incubate at 16°C for 4-20 hours. Transform 2 μL of ligation mix into 25 μL competent DH5 ...

  4. CRISPR Library Amplification

    Type
    Protocol
    ... Day 1: Transform, recover, set up overnight growth (Estimated time 2-3 hours) Transformation should be...of 5 mL (3 mL SOC + 2 mL transformed Endura from two separate transformations). Pro-Tip Do not pipette...Follow this protocol to perform amplification of CRISPR pooled plasmid libraries in Escherichia coli ...assumes familiarity with standard bacterial transformation and basic knowledge of the library being amplified...amplified. Please see our protocol on Bacterial Transformation . The generalized pooled library amplification... be performed at the end of the day to ensure that growth times are limited. Day 2: Harvest cells and ...using a new vented falcon tube for every two transformations. Each of the four 14 mL Vented Falcon Tubes...

  5. Immunocytochemistry

    Type
    Protocol
    ... of cold 4% paraformaldehyde in PBS on ice for 15 min . Pro-Tip While 4% Paraformaldehyde fixation works...rocking platform. Permeabilize cells for 10 min at room temperature ( RT ) on a rocking platform in 500...rocking platform. Section 3: Labeling with antibody Block for 20 min at RT on a rocking platform in 500...labeling HeLa cells for a target protein using the formaldehyde fixation method. The protocol may need to be...specific products or equipment. Inclusion of this information is solely for transparency intended to support...Pipette controller Pipette tips and pipettes Rocking platform Tweezers Fluorescent microscope 0.5–10 µL single...coated coverslips HeLa cells 24-well plate 4% Paraformaldehyde 5 mg/mL 4′,6-diamidino-2-phenylindole (DAPI...

  6. Protocol - Over-Agar Antibiotic Plating

    Type
    Protocol
    ...incubation, transform DH5α E. coli cells by heatshock with the plasmid of interest. See our transformation page...for a detailed heatshock transformation protocol. Plate 50 µL of transformed E. coli /rescue media suspension...also like... Making LB Agar Plates Bacterial Transformation Recovering Plasmid DNA from Bacterial Culture...plating antibiotic over-agar for the selection of transformed E. coli . Over-agar spreading of antibiotic makes...investigator to conveniently plate and select transformed cells containing plasmids differing in their ... sterile water (or other antibiotic) E. coli transformed with a plasmid containing the carbenicillin (...for 18 hours. Day 2 Observe plates for colony formation. Shown below are the results from an experiment...

  7. Western Blot

    Type
    Protocol
    ... a shaking platform. Wash the membrane 3x for 5 min in 1X TBST at RT on a shaking platform. Section 6:... a shaking platform. Wash the membrane 3x for 5 min in 1X TBST at RT on a shaking platform. Prepare the...specific products or equipment. Inclusion of this information is solely for transparency intended to support...supply iBlot 2 Gel Transfer Device Roller Spatula Platform shaker Cold room Gel imager -80 °C freezer Reagents...the manufacturer's instructions for additional information specific to your antibody, such as ideal blocking...between antibodies. Refer to the manufacturer's information before running your experiment. Remove the membrane...overnight in primary antibody at 4 °C on a rocking platform. Pro-Tip Primary antibody incubation can be reduced...

  8. Plasmid Cloning by Restriction Enzyme Digest (with Protocols)

    Type
    Protocol
    ...self-ligating recipient plasmid backbone. Transformation Transform your ligation reaction into your bacterial...from your transformation will give you the first indication as to whether your transformation worked. our...Information about plasmid cloning by restriction enzyme digest (subcloning), including design and experimental...Digest of Plasmid DNA DNA Ligation Bacterial Transformation Summary The following technique can be used...in a new window) New England Biolabs for more information about restriction enzyme buffers). If you select...competent cells. For most standard cloning, you can transform 1-2μl of your ligation reaction into competent...conduct a positive control to ensure that your transformation worked. You should also verify that you are...

  9. Protocol - How to Ligate Plasmid DNA

    Type
    Protocol
    ...Digest of Plasmid DNA Bacterial Transformation Background Information The final step in the construction...ligation, is performed by the T4 DNA ligase enzyme. The DNA ligase catalyzes the formation of covalent ...the backbone and the complete plasmid can be transformed into bacterial cells for propagation. The majority...digested vector backbone with a phosphatase before performing the ligation reaction (phosphatase removes the... incubation at 37°C. Proceed with bacterial transformation . Tips and FAQ Do controls When doing ligations...contamination of intact plasmid in ligation or transformation reagents Optimizing the Vector:Insert Ratio...

  10. Coomassie Purity Stain of Recombinant Antibodies

    Type
    Protocol
    ...specific products or equipment. Inclusion of this information is solely for transparency intended to support...Microcentrifuge Electrophoresis chamber Power supply Rocking platform Fume hood Metal spatula Razor blade Plastic tray...water for 5 min with gentle agitation on a rocking platform. Pour off the water in the sink. Add 20 mL of ...incubate for 1 h with gentle agitation on a rocking platform. Pour off the SimplyBlue SafeStain in the sink...incubate for 1 h with gentle agitation on a rocking platform. Pour off the water in the sink. Add 100 mL of...incubate for 1 h with gentle agitation on a rocking platform. Pour off the water in the sink. Take a brightfield... exported CSV file. Select import. Choose file. Format as needed. Select the Insert tab, then Chart . ...
Showing: 11 - 20 of 46 results