po4 0.2.0

Example

• Your want a tool to better manage your plasmids, but you don’t want to spend a bunch of time re-entering the information for your existing plasmids, which is stored in a big Excel spreadsheet. After installing PO₄, you quickly configure it to recognize the column headers in your spreadsheet. Nothing about your existing spreadsheet needs to change.

• You just designed primers to clone a new plasmid. You enter both the primers and the plasmid into your spreadsheet. While it’s fresh in your head, you also describe how the plasmid will be made in a column labeled “Construction”. The description might look like:

  INV: template=p2 primers=o2,o3

  This syntax indicates that the plasmid is constructed by inverse PCR (“INV”) using the plasmid “p2” as a template and the oligos “o1” and “o2” as primers.

• You just received the primers you designed last week, but you don’t remember exactly how you were planning to use them. You use PO₄—in conjunction with stepwise—to generate an protocol specifying all the relevant details: volumes, concentrations, construct names, etc. If you are making several constructs, the protocol will even group similar constructs and create master mixes of common reagents when possible:

  $ stepwise make p3
  March 27, 2020
  
  $ stepwise make p3
  
  1. Prepare 10x primer mix [1]:
  
     Reagent   Stock    Volume
     ─────────────────────────
     o2       100 µM   0.50 µL
     o3       100 µM   0.50 µL
     water             9.00 µL
     ─────────────────────────
                      10.00 µL
  
  2. Setup 1 PCR reaction and 1 negative control [2]:
  
     Reagent           Stock    Volume
     ─────────────────────────────────
     water                     3.00 µL
     p2             20 pg/µL   1.00 µL
     primer mix          10x   1.00 µL
     Q5 master mix        2x   5.00 µL
     ─────────────────────────────────
                              10.00 µL
  
  3. Run the following thermocycler protocol:
  
     - 98°C for 30s
     - Repeat 35x:
       - 98°C for 10s
       - 60°C for 20s
       - 72°C for 3 min
     - 72°C for 2 min
  
  4. Run 1 ligation reaction:
  
     Reagent              Stock    Volume
     ────────────────────────────────────
     water                        6.75 µL
     T4 ligase buffer       10x   1.00 µL
     T4 PNK             10 U/µL   0.25 µL
     T4 DNA ligase     400 U/µL   0.25 µL
     DpnI               20 U/µL   0.25 µL
     PCR product       50 ng/µL   1.50 µL
     ────────────────────────────────────
                                 10.00 µL
  
     - Incubate at room temperature for 1h.
  
  Notes:
  [1] For resuspending lyophilized primers:
      100 µM = 10 µL/nmol
  
  [2] For diluting template DNA to 20 pg/µL:
      Dilute 1 µL twice into 7*sqrt(DNA) µL

Installation

Install PO₄ using pip:

$ pip install po4

Usage

More details coming soon…