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PPMS Orders & Cherry picks

(Last updated: March 2020)

Accepting DRSC orders: 

1)  Login to PPMS and check “Orders – New” for new orders (https://ppms.us/hms-drsc/login/?pf=2 )

2)  Click the order number, then select “service form" (located next to the description of the service).

3)  See if the user selected “Choose My Own (I have DRSC IDs)” or “Please assist with amplicon selection (e.g. based on gene names).” 

  • If “Please assist with amplicon selection” is selected, see the Cherry pick amplicons protocol for details about how to select appropriate amplicons for a project. 
  • If DRSC IDs are provided, copy those IDs into a new Excel sheet. Name it something like “order ### cherry pick.” 

4)  Look under “Please indicate what common controls you would like” to see which controls should be included. 

  • LacZ and thread are the most common, but there are PCR products of all four  controls.
  • If the user specified no controls, verify with DRSC Director. 

5)  Return to the order page and check the box “Send a notification by email.” Select the user’s email, DRSC Director email, and Bioinformatics Director email. Then click “Accept order.” The order will now be found under “Orders – Accepted” on the main PPMS page. 

Finding cherry pick samples: 

1)  Login to the FlyRNAi Internal Screening Database & LIMS.  
https://www.flyrnai.org/cgi-bin/RNAi_login.pl 

2)  Click  "Find wells for Amplicon and Genes” (under “RNAi Center Internal Functions"). Copy the DRSC IDs from your Excel file and paste them in the box. 

  • Order: Keep original order 
  • Use: PCR/Cherry Pick plates only 
  • Check “Quentinize – remove extra sources”
  • Hit “Find Wells” 

3)  Add another sheet to your Excel file. Copy/paste in the table results (exclude “plate type”). 

  • Copy/paste the original list of DRSC IDs from the first sheet to the second sheet in order to check if all IDs are accounted for. This can be done manually or by using a formula. 

4)  Copy/paste the table from “Amplicon” to “Promotor (T7)” into a new Excel workbook. This workbook is used to make the MultiProbe robot files. 

  • Sort the list by “Plate” so that amplicons in the same plate are next to each other. 

Preparing Plates for Cherry-Picking 

1)  The plates are in the green racks in Freezer #1. All racks are labeled for easy plate location. 

  • The top shelf of Freezer #1 sags down, so all racks on that shelf must be removed to access the second shelf. 
  • These source plates are labeled “rePCR,” meaning that they’re the diluted and normalized PCR amplicons . The original PCR plates are in the other -20 freezer. 

2)  Thaw out the plates, then vortex using the plate vortexer for 30 seconds. Spin down for 1 minute (can use the Eppendorf plate-spinning centrifuge in Bay 1, but the tabletop plate-spinning centrifuge in Bay 2 is also sufficient). 

3)  Move onto Operating the MultiProbe!