What to do when a plasmid comes for you: agar stab to plate to glycerol stock, miniprep & sequencing

I ordered a bunch of plasmids, and many of them come as agar stabs, where they've taken bacteria with the plasmid and stabbed it into a tube of agar. So it's like a Petri dish in a tube. My gameplan:
- streak a plate
- isolate a colony
- grow a 5mL overnight culture (LB + appropriate antibiotic)
- make a glycerol stock
-- take 500μL of that culture & add to 500μL sterile glycerol; invert to mix & store in -80°C freezer - should be good for years if you avoid freeze-thaws
- miniprep the rest of the overnight culture to isolate the plasmid
- send the plasmid for sequencing
- if the sequence checks out, do whatever the heck you got it for! (subclone into a different vector, transform into expression cells, etc.)

blog form: https://bit.ly/plasmid_prepping

more on cloning cells: blog form: https://bit.ly/dh5alpha ; YouTube:    • DH5α strains and a PSA to sequence yo...    

more on expression cells: https://bit.ly/bl21_rare_codons ; YouTube:    • Bacterial expression strains - esp BL...  

more on molecular cloning: http://bit.ly/molecularcloningguide ; YouTube:    • Molecular cloning overview - techniqu...  

more on transformation: http://bit.ly/transformheatshock ; YouTube:    • Bacterial transformation - chemically...  

more on T7 RNAP: http://bit.ly/t7rnap

more on mini preps: http://bit.ly/minipreps ; YouTube:    • Miniprep science - what goes on in ea...    

more on glycerol solution-making:    • Making a 80% glycerol stock solution ...  

some tips for pipetting & working with viscous solutions: blog: https://bit.ly/pipetting_problems ; YouTube:    • Pipetting viscous liquids - reverse p...  

random practical lab tips & tricks: https://bit.ly/lab_tricks_page 

link to the plasmid I was talking about: https://www.addgene.org/174866/

helpful guide from addgene: https://www.addgene.org/recipient-ins...

more about all sorts of things: #365DaysOfScience All (with topics listed) 👉 http://bit.ly/2OllAB0 or search blog: http://thebumblingbiochemist.com                             
                               
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