Sequencing reagent stability

AciBASE · 10-25-2010, 07:15 AM

Has anyone had any problems with the stability of the sequencing reagents? In particular a reduction in read length due to storage of the reagents for longer than 16 hours (at 4 °C).
In the manual, the reagents are said to be stable for up to 60 h at 2 - 8 °C but we’ve found that isn’t the case. Anybody suffer from something similar?

Soulbee · 03-01-2011, 07:02 AM

I always thaw out my sequencing reagents at 4 degrees overnight (from -20) and never have failed or saw any decrease in read length... could anything else be an issue?

MissDNA · 03-01-2011, 07:15 AM

We always thaw our reagents by putting them in water right when we start to set up the run. Yet we have been suffering from short reads since we started using the kits with the new control beads.
However there is a possibility that our reagents are not performing well cause something may have happened during transportation. Our case is under investigation.
I would like to know if people are still getting 400+ reads on regular basis since this change. Because with the old kits most of our runs were 400+.

ashchin · 03-01-2011, 11:55 AM

Hi ,

Is the % of formamide in hiseq 2000 waste around 0.1.
Please correct me if I am wrong. How do you guys dispose the waste?

Thanks!

Santosh · 03-01-2011, 11:39 PM

We have a very strange problem with the GA2X machine here. We spotted few bubbles while doing the leak check with the actual reagents after placing the flow cell. The bubbles were found in the inlet pipe pumping reagents into the Flow cell, within the flow cell and also in the syringes that pump the fluids out of the machine. We do not know how the buubles found their way into the pipes and syringes. We always make sure that the reagent bottles are kept filled all the time. An engineer had visited the lab recently and could not conclude on how the bubbles were introduced into the system and neither could he solve our bubble problem. Instead, he suggested us to start wth the run. After the run started, we found that in each lane, there were few tiles which were black, instead of being either blue, green or yelow. I could make out quite clearly that it was a bubble issue. Does anyone here wrking on a same kind of machine, ever had a similar problem? If ye, I would be very grateful to that person if he or she could suggest me a solution.
Santosh

RCJK · 03-02-2011, 12:08 AM

Quote:

Originally Posted by ashchin

Hi ,

Is the % of formamide in hiseq 2000 waste around 0.1.
Please correct me if I am wrong. How do you guys dispose the waste?

Thanks!

Hi there, I'd suggest posting this in the Illumina forum, not the 454 forum, so that you can improve your chance of getting a response.

Santosh · 03-02-2011, 03:27 AM

Hey there,
I'm new to this forum. I didn't know it.Thank u very much for the advise

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10-25-2010, 07:15 AM	#1
AciBASE Junior Member Location: Birmingham Join Date: Oct 2010 Posts: 1	Sequencing reagent stability Has anyone had any problems with the stability of the sequencing reagents? In particular a reduction in read length due to storage of the reagents for longer than 16 hours (at 4 °C). In the manual, the reagents are said to be stable for up to 60 h at 2 - 8 °C but we’ve found that isn’t the case. Anybody suffer from something similar?

03-01-2011, 11:39 PM	#5
Santosh Member Location: India Join Date: Feb 2011 Posts: 11	GA2X Bubble Problem We have a very strange problem with the GA2X machine here. We spotted few bubbles while doing the leak check with the actual reagents after placing the flow cell. The bubbles were found in the inlet pipe pumping reagents into the Flow cell, within the flow cell and also in the syringes that pump the fluids out of the machine. We do not know how the buubles found their way into the pipes and syringes. We always make sure that the reagent bottles are kept filled all the time. An engineer had visited the lab recently and could not conclude on how the bubbles were introduced into the system and neither could he solve our bubble problem. Instead, he suggested us to start wth the run. After the run started, we found that in each lane, there were few tiles which were black, instead of being either blue, green or yelow. I could make out quite clearly that it was a bubble issue. Does anyone here wrking on a same kind of machine, ever had a similar problem? If ye, I would be very grateful to that person if he or she could suggest me a solution. Santosh

03-01-2011, 07:02 AM	#2
Soulbee Member Location: Toronto, Canada Join Date: Jun 2010 Posts: 25	I always thaw out my sequencing reagents at 4 degrees overnight (from -20) and never have failed or saw any decrease in read length... could anything else be an issue?

03-01-2011, 07:15 AM	#3
MissDNA Senior Member Location: Brazil Join Date: Nov 2010 Posts: 146	We always thaw our reagents by putting them in water right when we start to set up the run. Yet we have been suffering from short reads since we started using the kits with the new control beads. However there is a possibility that our reagents are not performing well cause something may have happened during transportation. Our case is under investigation. I would like to know if people are still getting 400+ reads on regular basis since this change. Because with the old kits most of our runs were 400+.

03-01-2011, 11:55 AM	#4
ashchin Member Location: USA Join Date: Dec 2010 Posts: 19	Hi , Is the % of formamide in hiseq 2000 waste around 0.1. Please correct me if I am wrong. How do you guys dispose the waste? Thanks!

03-02-2011, 03:27 AM	#7
Santosh Member Location: India Join Date: Feb 2011 Posts: 11	Hey there, I'm new to this forum. I didn't know it.Thank u very much for the advise