How would I verify whether custom adapters have annealed properly? I get odd results when trying to quantify using QuBit, PicoGreen or TapeStation. Since the majority of the adapter is still single stranded, is there even a way to tell?
Seqanswers Leaderboard Ad
Collapse
Announcement
Collapse
No announcement yet.
X
-
As you have done, Qubit or PicoGreen is the easiest and very informative. With increasing annealing efficiency you should see increasing value in concentration of dsDNA. I regularly use this method as a QC for adapters with full length or partial length complimentary regions to ensure batch to batch consistency. Obviously, adapters with short complimentary region will have lower florescence, but it is sensitive enough to discriminate between annealed and non-annealed oligos. One also can calculate theoretical expected concentration for partially or completely complimentary oligos for any given annealing efficiency.
-
Originally posted by nucacidhunter View PostAs you have done, Qubit or PicoGreen is the easiest and very informative. With increasing annealing efficiency you should see increasing value in concentration of dsDNA. I regularly use this method as a QC for adapters with full length or partial length complimentary regions to ensure batch to batch consistency. Obviously, adapters with short complimentary region will have lower florescence, but it is sensitive enough to discriminate between annealed and non-annealed oligos. One also can calculate theoretical expected concentration for partially or completely complimentary oligos for any given annealing efficiency.
Originally posted by jwfoley View PostYou could also try denaturing them to see how much that reduces their fluorescence with a dsDNA-specific dye.
Comment
-
What kind of values have you seen with say a full TruSeq adapter?
Comment
Latest Articles
Collapse
-
by seqadmin
Technological advances have led to drastic improvements in the field of precision medicine, enabling more personalized approaches to treatment. This article explores four leading groups that are overcoming many of the challenges of genomic profiling and precision medicine through their innovative platforms and technologies.
Somatic Genomics
“We have such a tremendous amount of genetic diversity that exists within each of us, and not just between us as individuals,”...-
Channel: Articles
05-24-2024, 01:16 PM -
-
by seqadmin
The sequencing world is rapidly changing due to declining costs, enhanced accuracies, and the advent of newer, cutting-edge instruments. Equally important to these developments are improvements in sequencing analysis, a process that converts vast amounts of raw data into a comprehensible and meaningful form. This complex task requires expertise and the right analysis tools. In this article, we highlight the progress and innovation in sequencing analysis by reviewing several of the...-
Channel: Articles
05-06-2024, 07:48 AM -
ad_right_rmr
Collapse
News
Collapse
Topics | Statistics | Last Post | ||
---|---|---|---|---|
Started by seqadmin, Yesterday, 06:55 AM
|
0 responses
12 views
0 likes
|
Last Post
by seqadmin
Yesterday, 06:55 AM
|
||
Started by seqadmin, 05-30-2024, 03:16 PM
|
0 responses
24 views
0 likes
|
Last Post
by seqadmin
05-30-2024, 03:16 PM
|
||
Comprehensive Sequencing of Great Ape Sex Chromosomes Yields Insights into Evolution and Genetic Variability
by seqadmin
Started by seqadmin, 05-29-2024, 01:32 PM
|
0 responses
28 views
0 likes
|
Last Post
by seqadmin
05-29-2024, 01:32 PM
|
||
Started by seqadmin, 05-24-2024, 07:15 AM
|
0 responses
215 views
0 likes
|
Last Post
by seqadmin
05-24-2024, 07:15 AM
|
Comment