I was just wondering if anyone has used the minIon for plasmid sequencing? I suppose it might be overkill and a waste or reagents for such a small task. But I am asking because I have large plasmids to sequence (in the 18 to 20 kb range), and the plasmids contain several copies (up to as many as 4 copies) of certain sequences. So using sanger sequencing is costly and labor intensive and we end up with reads that could align to more than one spot on the plasmid making it hard to tell if and where a mutation has occurred. I am also exploring Illumina based methods, but am curious about the nanopore technologies. Seems like a 20 kb plasmid could simply be linearized and the entire molecule sequenced in a single read.
Seqanswers Leaderboard Ad
Collapse
Announcement
Collapse
No announcement yet.
X
-
Have a look at methods for Zika virus sequencing project.
This sounds very similar to zika sequencing project, and should work reasonably well (provided it is medium GC% sequence).
Ideally one would like to make partial digest of the plasmid, aiming to cut most of them once or twice, and make 2D library from it.
PS: keep in mind the ~0.1 - 1% systematic error rate remaining after assembly consensus steps. It would really depend on the actual template sequence.
Hopefully there are no high GC hairpins in the sequence, because they would stop the complementary read (which goes from single stranded template).
So you may have some bits that have very low quality hear the hairpins.
Latest Articles
Collapse
-
by seqadmin
The field of epigenetics has traditionally concentrated more on DNA and how changes like methylation and phosphorylation of histones impact gene expression and regulation. However, our increased understanding of RNA modifications and their importance in cellular processes has led to a rise in epitranscriptomics research. “Epitranscriptomics brings together the concepts of epigenetics and gene expression,” explained Adrien Leger, PhD, Principal Research Scientist on Modified Bases...-
Channel: Articles
Yesterday, 07:01 AM -
-
by seqadmin
Proteins are often described as the workhorses of the cell, and identifying their sequences is key to understanding their role in biological processes and disease. Currently, the most common technique used to determine protein sequences is mass spectrometry. While still a valuable tool, mass spectrometry faces several limitations and requires a highly experienced scientist familiar with the equipment to operate it. Additionally, other proteomic methods, like affinity assays, are constrained...-
Channel: Articles
04-04-2024, 04:25 PM -
ad_right_rmr
Collapse
News
Collapse
Topics | Statistics | Last Post | ||
---|---|---|---|---|
Started by seqadmin, 04-11-2024, 12:08 PM
|
0 responses
55 views
0 likes
|
Last Post
by seqadmin
04-11-2024, 12:08 PM
|
||
Started by seqadmin, 04-10-2024, 10:19 PM
|
0 responses
51 views
0 likes
|
Last Post
by seqadmin
04-10-2024, 10:19 PM
|
||
Started by seqadmin, 04-10-2024, 09:21 AM
|
0 responses
45 views
0 likes
|
Last Post
by seqadmin
04-10-2024, 09:21 AM
|
||
Started by seqadmin, 04-04-2024, 09:00 AM
|
0 responses
55 views
0 likes
|
Last Post
by seqadmin
04-04-2024, 09:00 AM
|
Comment