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We narrowed to 18 results for: total dna

Showing: 1 - 10 of 18 results

  1. Molecular Biology Protocol - Restriction Digest of Plasmid DNA

    Type
    Protocol
    ...Recovering Plasmid DNA from Bacterial Culture Purifying DNA from an Agarose Gel DNA Ligation Introduction...involves ∼500 ng of DNA, while molecular cloning often requires 1 µg of DNA. The total reaction volume usually... Protocols Restriction Digest of Plasmid DNA Restriction Digest of Plasmid DNA You may also...following: DNA Restriction Enzyme(s) Buffer BSA (if recommended by manufacturer) dH 2 O up to total volume...volume of DNA to be cut. A typical restriction digestion reaction could look like this: 1 µg DNA 1 µL of...the amount of DNA you want to cut. By definition: one unit of enzyme will cut 1 µg of DNA in a 50 µL reaction...purifying the DNA via a purification kit, such as a (Link opens in a new window) QIAGEN DNA cleanup kit ...

  2. Lentivirus Production

    Type
    Protocol
    ... using μg total DNA to μg PEI ratios of 1:1, 1:2, 1:3 and 1:6. The 1:2 and 1:3 total DNA:PEI μg ratios...1000 μL total per 10 cm dish). Using transfer plasmid pHAGE TRE dCas9-KRAB (total μg of plasmid DNA 27.8 ...plasmid DNA purification should include an endotoxin removal step. For high quality plasmid DNA, the plasmids...the diluted DNA mixture. Add the diluted PEI dropwise while gently flicking the diluted DNA tube. Incubate...mixture of a total of 500 μL PEI-OptiPro SFM with enough PEI such that the ratio of μg DNA:μg PEI is 1:3...possible range of ratios to test: Ratio of DNA:PEI μg of DNA μL of 1 mg/mL PEI 1:1 18.9 18.9 1:2 18.9 37.8...polyethersulfone (PES) filter Transfection grade DNAs: Lentiviral Packaging plasmid(s), Envelope plasmid...

  3. CRISPR Library Amplification

    Type
    Protocol
    ...pooled library DNA (100 ng plasmid DNA per 25 µL electrocompetent cells) Less library DNA can be used but...Each different DNA measurement type can report slightly different concentrations of DNA. In our hands ...contain sufficient DNA for use, pool samples, and continue with analysis. “Sufficient” DNA will vary upon ...several hundred micrograms to a milligram of total plasmid DNA. Tips and Troubleshooting What do I do if ...pooled plasmids differ by only a short region of DNA: guide RNAs targeting different genes in an organism's...recombinant band makes up a significant proportion of the DNA pool. Last Update: August 17, 2023 Workflow Timeline...times are limited. Day 2: Harvest cells and purify DNA (Estimated time 3-4 hours) Cells should be harvested...

  4. AAV Production in HEK293 Cells

    Type
    Protocol
    ...with 2 mg total DNA per CS5 Plasmid Plasmid Size (bp) DNA Concentration (μg/μL) Volume of DNA (μL) RepCap...1.00 584.2 Total bp 24,961 In total, we would like 2 mg of DNA or 2000 μg. Using the total number of base...each plasmid DNA into the bottle containing the Opti-MEM. Mix well. Add 4 mL of PEI (1:2 μg DNA to μg PEI... million cells from the CS2 into one CS5 with a total volume of 850 mL. Return to the incubator for 24...pairs from all three plasmids, we can determine the total μg/bp we need to achieve a 1:1:1 molar ratio of ...reduce transfection efficiency. Add the OptiMEM + DNA + PEI solution to the bottle containing 774 mL of...into a waste container. Carefully add the OptiMEM + DNA + PEI solution to the CS5. Make sure that all five...

  5. What is Polymerase Chain Reaction (PCR)

    Type
    Protocol
    ...stranded DNA template strands. The DNA polymerase is also stable enough to now bind to the primer DNA sequence...Plasmid Cloning by PCR Restriction Digest of Plasmid DNA DNA Ligation Background Information A standard Polymerase...which would denature DNA polymerases from other species. In addition to the template DNA and the Taq polymerase...several micrograms of target DNA from just a few nanograms of template DNA through several cycles of denaturation...this step enables the DNA polymerase to synthesize and elongate the new target DNA strand accurately and... are using the same template DNA for all your reactions, the Template DNA can be added to the master mix...reactions = 35μl total + 10% (3.5μl) = 38.5μl dNTP mix (10 mM each nt): 1µl x 7 reactions = 7µl total + 10% (0.7µl...

  6. Protocol - How to Ligate Plasmid DNA

    Type
    Protocol
    ...and insert DNA to use. Combine the following in a PCR or Eppendorf tube: Vector DNA Insert DNA Ligase Buffer...for 5X buffer) 0.5-1μL T4 DNA Ligase H 2 O to a total of 10μL Notes: If the DNA concentrations are low such... Protocols DNA Ligation DNA Ligation You may also like... Plasmid...backbone of the two DNA fragments. This reaction, called ligation, is performed by the T4 DNA ligase enzyme....the ligation reaction. The volume of vector DNA and insert DNA used in the ligation will vary depending ... work just fine. We recommend around 100ng of total DNA in a standard ligation reaction. Use a (Link opens... of DNA ends available for ligation. Simply put, there are only two ends on any given piece of DNA no ...

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

    Type
    Protocol
    ...may also like... Restriction Digest of Plasmid DNA DNA Ligation Bacterial Transformation Summary The following... your digested DNA on an agarose gel and conduct a gel purification to isolate the DNA. When running a...concentration of recovered DNA . Ligate your insert into your vector Conduct a DNA Ligation to fuse your insert...recipient plasmid. We recommend around 100ng of total DNA in a standard ligation reaction. You ideally ...such as DH5alpha or TOP10. If using much less total DNA (<1ng) or if you are having trouble getting colonies...and grow overnight cultures for DNA purification. After purifying the DNA , conduct a diagnostic restriction...technique can be used to easily move any piece of DNA from one vector to another as long as it is already...

  8. Plasmid Cloning by PCR (with Protocols)

    Type
    Protocol
    ... are amplifying from plasmid DNA (rather than from genomic DNA or a cDNA library), roughly 18-21bp is ...may also like... Restriction Digest of Plasmid DNA DNA Ligation Bacterial Transformation Summary PCR based... piece of DNA and at the same time adding restriction sites to the ends of that piece of DNA so that it...Run your digest DNA on an agarose gel and conduct a gel purification to isolate the DNA. When running a...concentration of recovered DNA . Ligate Your Insert Into Your Vector Conduct a DNA Ligation to fuse your insert...recipient plasmid. We recommend around 100ng of total DNA in a standard ligation reaction. You ideally want...such as DH5alpha or TOP10. If using much less total DNA (<1ng) or if you are having trouble getting colonies...

  9. Protocol - How to Purify DNA from an Agarose Gel

    Type
    Protocol
    ... Protocols Purifying DNA from an Agarose Gel Purifying DNA from an Agarose Gel You may also..... Restriction Digest of Plasmid DNA Agarose Gel Electrophoresis DNA Ligation Background Information Gel...which separates DNA by their length in base pairs. Following electrophoresis, you can cut DNA bands out of... minimize the risk of DNA damage, it is best to limit the UV exposure of the DNA. Therefore, it is a bad...important during the DNA purification step, as many kits cannot handle more than a certain total volume of gel...buffer to add during the DNA isolation step. Finally, you will want to isolate the DNA from the gel. This is...intended step with the now gel purified DNA. Find more information about DNA quantification here . Tips and FAQ...

  10. Protocol - Bacterial Transformation

    Type
    Protocol
    ...of Plasmid DNA DNA Ligation Introduction Transformation is the process by which foreign DNA is introduced...with less DNA, especially when using highly competent cells. If you used 100-1000 ng of total DNA in a ligation... cells to take up the DNA, an electro-magnetic field is applied to the cell/DNA mixture to induce membrane...the plasmid without rearrangement of the plasmid DNA. Additionally, specific treatments have been discovered...least expensive) are fine for transforming plasmid DNA for the purposes of storage and amplification. Higher... will be transforming with very small amounts of DNA or if you're multiple plasmids at once. To save money...appropriate antibiotic) LB or SOC media Competent cells DNA you'd like to transform Procedure Take competent ...
Showing: 1 - 10 of 18 results