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...opens in a new window) NEB , (Link opens in a new window) Qiagen , (Link opens in a new window) Invitrogen.... For details on how to streak a plate to get individual colonies and to generate liquid bacterial cultures...Invitrogen , and (Link opens in a new window) Promega sell kits for isolating plasmid DNA in quantities as low...plasmid purification without using a kit, you can find a protocol for kit-free plasmid mini-prep at the... DNA or apply the solution to a column that will bind the now precipitated DNA. Wash the pellet or column...resuspension buffer of most kits. Reference Page Top Index...

...sticky ends", are what allow the vector and insert to bind to each other. When the sticky ends are compatible...standard ligation reaction. Use a (Link opens in a new window) ligation calculator to easily quantify how much...take a fresh tube, thaw it one time and aliquot individual tubes of 5, 10 or 20μL for storage at -20°C. ...troubleshooting failed ligations. The following table indicates the various controls: Control Ligase Interpretation... treated vector Insert or water + Any colonies indicate contamination of intact plasmid in ligation or...every available vector end. (Link opens in a new window) Ligation calculators are easily found on the web...exactly how much of each to use. Reference Page Top Index...

...that the only version of full-length cDNA you can find for YGOI is in a bacterial expression vector. Using...are creating a fusion protein) Ideally, you will find two different restriction enzymes for your subcloning...cloned in the correct orientation. If you cannot find enzymes that meet these criteria, do not fear. You...restriction enzyme buffer (see (Link opens in a new window) New England Biolabs for more information about...from your transformation will give you the first indication as to whether your transformation worked. our...Finished Plasmid Finally, you will need to pick individual bacterial colonies and check them for successful... and can begin your studies. Reference Page Top Index...

...recombination can all impact the representation of individual plasmids in the pooled library during amplification...designed to be as general as possible but note that individual libraries may require modifications dictated ...satisfactory results for libraries up to 200,000 individual plasmids and can be scaled according to the actual...cells is sufficient for libraries up to 200,000 individual plasmids. For larger libraries scale the number...but this dramatically increases the chances of individual plasmids being lost from the pool and/or increasing...like Glycerol or DMSO solutions. Quantify the individual Maxipreps by Nanodrop, Picogreen, or Qubit. Each...or reagents are significantly overloaded. If you find that your pellet outstrips the capacity listed in...

... Test a variety of brands and FBS lot numbers to find one suitable with your protocols. FBS can be purchased...Check the cells for mycoplasma (Link opens in a new window) regularly Do not over or under-grow your cells...of resistant cells start proliferating and the individual wells become confluent, expand into larger vessels...Generation of monoclonal cell lines from expansion of individual A549 cells stably expressing Cas9. A549 cells...for 9 days. Single cells were then plated in individual wells of a 96-well plate and left undisturbed...Methods. 2014 Aug;11(8):783-4 (Link opens in a new window) . Last reviewed on: September 9, 2023...

...can hold 5-10 mL of agar and on which you can individually distinguish a maximum of ~100 colonies Pro-Tip...Tetracycline 10 mg/mL 10 µg/mL Notes: Unless otherwise indicated, the antibiotic powder can be dissolved in dH ..., you should prepare your plate pouring station: Find an empty section of lab bench with a working flame... looks like in your particular plate. For the remainder of the plates, pour directly from the bottle. ...test results. Sample Data In all cases below (-) indicates that the tested strain is not supposed to be resistant... resistant to the antibiotic, (+) indicates that the tested strain is supposed to be resistant to the ...

... proteins.    If this video is helpful, you can find our complete collection of protocol videos on the...

...out all of our protocols on our Protocol page, and find our protocol videos on our YouTube page.     ...

...depositing labs (currently at 4,651 as of April 8, 2021) Find these stats and more on the addgene.org homepage...

...Bench Layout,” is below.      And you’ll be able to find all published videos in our Quick Tips playlist:...

...Hmmm, they need a name. And we need your help in finding a good name for Blugene’s virus friend.  During...

...new CRISPR base editors, DIY DNA ladders. You can find additional hot plasmids in our newsletter or on ...

..., and vectors for zebrafish engineering. You can find additional hot plasmids in our newsletter or on ...

...between some meaningful names…. Watch the video to find out the final four! (Note: For cases where the same...

... in 2021 and useful resources, fear not! You can find all of them here.  On the first day of CRISPR, ...

...interested in engaging with us on social media, you can find us on Bluesky: @addgene.bsky.social; or @addgene...

...transfection. Test a variety of brands and lots of FBS to find one suitable with your protocols. FBS can be purchased...Check the cells for mycoplasma (Link opens in a new window) regularly Do not over or under-grow your cells... the minimum concentration (Link opens in a new window) required to kill all of the cells. Use this dose...Figure 1: A549 cells were transduced with the indicated serial dilutions of the lentiviral vector pRosetta...Figure 2: A549 cells were transduced with the indicated serial dilutions of the lentiviral vector pHAGE...

...contributions have helped researchers around the globe find and share knowledge, resources, and new ideas - ...

...Browse our Video Library Check out Our Protocol Pages Find Molecular Biology Resources ...

...control exceptions were made by the US Bureau of Industry and Security, pursuant to the international controls...