The Bases
sgRNA is an abbreviation for...
Single guide RNA
In what order is the OP turned on? off?
ON = Pressure, Machine, Computer
OFF = Computer, Machine, Pressure
What is being filtered out during the first TFF step?
DMSO
When gowning in, what two things should you preform before putting on a lab coat?
Don booties, and wash hands
When a number or letter is changed rather than crossed out and initialed
What is a write over
List the mod. codes used to denote RNA bases.
What are rA, rG, rC, and rU
When changing amidite bottles on the OP100 what PPE should always be worn and why?
A face shield, because the system is pressurized.
sgRNA is dual purified. What are the two purifications used, and in what order?
RP (Reverse Phase) and IE (Ion Exchange)
What direction should the lab and equipment be cleaned?
An additional comment used to explain a cross out
What is a footnote
In what direction is an oligo synthesized?
3'prime to 5' prime
How are modified bases represented in Unicorn?
With a line above the base
What would happen if RP buffer was not properly flushed from the AKTA Pure before attaching IE buffer?
Precipitate would form in the lines and clog the machine.
When cleaning, what is the contact time for steri-perox?
30 minutes
Give an example of something that might be attached to a BR as an attachment...
TFF cassette COA, RP column COA, any material certificates
What two modifications are typically used when synthesizing an sgRNA? Where do they occur in the sequence?
O-Methyl and Thiolation. Typically at the beginning and end of the sequence.
What two steps should be added to the last synthesis part?
A final detrit for oligos without a ndtag and a DEA wash.
What size filter is used for the final filter (pre lyo)?
0.22 micron filter
A face shield is required when cleaning this part of the lab.
What is the ceiling.
These two things should be included when a cross out occurs.
Initial and date
When synthesizing what is different about the first five bases?
They are double coupled
At what point(s) of the synth method is it safe to pause a run?
During any wash cycle, when there is no color left in the column.
When doing final fill and lyo what technique is used when handling the final product?
Aseptic - a way that minimizes the risk of contaminating an invasive device
How often should lab coat, booties, and scrubs be changed?
Minimum of once daily.
For unused cells what is the proper documentation practice?
Single line through cells, with NA, Date and initials.