Tag Archives: MICROBExpress Kit

RNA Gel – V. tubiashii mRNA samples (from 20081224)

external image 20081229.png

Lane 1 – Empty
Lane 2 – Total RNA, Control
Lane 3 – mRNA, Control
Lane 4 – Total RNA, Vibrio+gigas
Lane 5 – mRNA, Vibrio+gigas

Results:

rRNA removal seems to have worked relatively well. Still some residual rRNA present in the mRNA samples. Will submit 200ng of the Vibrio+gigas mRNA to HTGU for Illumina sequencing.

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rRNA Removal – V. tubiashii total RNA from yesterday

rRNA removal was continued from O/N precipitation. Processed the samples according to the Ambion MICROBExpress Kit protocol and resuspended final pellets in 25uL of The RNA Storage Solution. Samples were spec’d on the NanoDrop.

external image 20081224%20RNA%20SJW.PNG

Samples were stored @ -80C in Sam’s RNA Box #1.

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RNA Isolation – V. tubiashii from challenge (see 20081216)

Added 10mL TriReagent to 2 x 50mL pellets (5.63 x 10^11 total bacteria; see calcs on 20081219) from the control samples collected on 20081219. Added 10mL TriReagent to 1 x 50mL pellet (1.835 x 10^12 total bacteria; see calcs on 20081219) from the V.tubi + oyster samples collected on 20081219. Scaled rest of RNA protocol to match. Resuspended pellets in 100uL 0.1%DEPC-H2O. Samples were NanoDropped.

external image 20081222%20RNA%20SJW.PNG

The V. tubi + gigas sample was eventually diluted to contain 400uL (see final reading for that sample above).

RNA samples were precipitated O/N @ -20C according to Ambion protocol.

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