reset reagent

eoh001 · 07-06-2011, 10:37 PM

Hello
I observed a strange sequence result for two times.
The run seemed OK until a certain point, and sequnce quality collapse and the low sequence quality retained until the end of the run.
Two Times !!!
I found that the sequnce quality drop down right after resetting new primer. and I found that there was a reset process right before applying new primer from the log file.
I checked the reset reagent, and the reset reagent bottle was almost empty!!!
I am not sure that the low sequence quality mainly from the shortage of the reset reagent, but I am kind of positive.

Anyone has an experience of running out of the reset reagent?
Any idea about the low sequence quality observed from the first cycle of new primer?

bioits · 07-07-2011, 08:50 AM

Since there is no cleave step after last cycle of each primer, if you run out of reset buffer, any further ligation won'thappen and most likely the instrument will just repeat to scan the last cycle of the primer before reset. What you can do is to get a new bottle of reset buffer and rerun the primer with low quality as far as you haven't clear the run and slide is still sitting on the instrument.

GenoMax · 07-07-2011, 10:18 AM

Quote:

Originally Posted by eoh001

Hello
I observed a strange sequence result for two times.
The run seemed OK until a certain point, and sequnce quality collapse and the low sequence quality retained until the end of the run.
Two Times !!!
I found that the sequnce quality drop down right after resetting new primer. and I found that there was a reset process right before applying new primer from the log file.
I checked the reset reagent, and the reset reagent bottle was almost empty!!!
I am not sure that the low sequence quality mainly from the shortage of the reset reagent, but I am kind of positive.

Anyone has an experience of running out of the reset reagent?
Any idea about the low sequence quality observed from the first cycle of new primer?

Deleting the comment. Did not see that the original comment referred to a SOLiD machine.

07-08-2011, 11:52 AM

There is a list of required volumes for each buffer in the appendix of the instrument manual. You sometimes have to add more reset depending on the type of run.

asaleh · 07-11-2011, 06:20 AM

Quote:

Originally Posted by SeqAA

There is a list of required volumes for each buffer in the appendix of the instrument manual. You sometimes have to add more reset depending on the type of run.

Agreed. I have noticed also that the reset buffer get sucked up a little faster than the manual states on my solid 4. However I think it is always a good idea to pause at a safe pause point and check your buffer levels if you think any of your reagents will come close to running out based on your calculations at the beginning of your run.

sciencegnome · 07-15-2011, 10:22 AM

As a general rule we change the bottle of reset buffer when we change the tags. We have a huge surplus of that particular buffer so "when in doubt, change it out".

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07-06-2011, 10:37 PM	#1
eoh001 Member Location: seoul Korea Join Date: Mar 2011 Posts: 19	reset reagent Hello I observed a strange sequence result for two times. The run seemed OK until a certain point, and sequnce quality collapse and the low sequence quality retained until the end of the run. Two Times !!! I found that the sequnce quality drop down right after resetting new primer. and I found that there was a reset process right before applying new primer from the log file. I checked the reset reagent, and the reset reagent bottle was almost empty!!! I am not sure that the low sequence quality mainly from the shortage of the reset reagent, but I am kind of positive. Anyone has an experience of running out of the reset reagent? Any idea about the low sequence quality observed from the first cycle of new primer?

07-07-2011, 08:50 AM	#2
bioits Member Location: USA Join Date: Aug 2010 Posts: 23	Since there is no cleave step after last cycle of each primer, if you run out of reset buffer, any further ligation won'thappen and most likely the instrument will just repeat to scan the last cycle of the primer before reset. What you can do is to get a new bottle of reset buffer and rerun the primer with low quality as far as you haven't clear the run and slide is still sitting on the instrument.

07-08-2011, 11:52 AM	#4
SeqAA Guest Posts: n/a	There is a list of required volumes for each buffer in the appendix of the instrument manual. You sometimes have to add more reset depending on the type of run.

07-15-2011, 10:22 AM	#6
sciencegnome Junior Member Location: Philly Join Date: Oct 2010 Posts: 4	As a general rule we change the bottle of reset buffer when we change the tags. We have a huge surplus of that particular buffer so "when in doubt, change it out".