We have one paired-end (2x 101 bp)libray Whole gnome short sequence (WGS) sequenced through illumina hiseq 2000. We have assembled genome assembly with Abyss software. The assembly has lower N50 value around 1Kbp and does not improve in scaffolding because we have only one paired-end libarary. The estimated insert size is around 240 bp and coverage is arond 5x. Our sequenced plant is a polyploid, we lack data in getting good draft assembly and as we have budget constraint, we cannot go for additional illumina paired-end sequencing. But we some budget for 1x pacBio sequencing. My question does 1x pacBio sequencing helps in improving assembly atleast to get organelle genomes.
Seqanswers Leaderboard Ad
Collapse
Announcement
Collapse
No announcement yet.
X
-
-
Originally posted by bioman1 View PostWe have one paired-end (2x 101 bp)libray Whole gnome short sequence (WGS) sequenced through illumina hiseq 2000. We have assembled genome assembly with Abyss software. The assembly has lower N50 value around 1Kbp and does not improve in scaffolding because we have only one paired-end libarary. The estimated insert size is around 240 bp and coverage is arond 5x. Our sequenced plant is a polyploid, we lack data in getting good draft assembly and as we have budget constraint, we cannot go for additional illumina paired-end sequencing. But we some budget for 1x pacBio sequencing. My question does 1x pacBio sequencing helps in improving assembly atleast to get organelle genomes.
If organellar genome is all you are interested in, you would be better off harvesting and concentrating the plastids using a cell sorter and subsequently sequencing the DNA extracted from the sorted organelles with PacBio
-
1x of PacBio will do absolutely nothing to improve the assembly, the minimum coverage would be ~10x for assembly improvement (gap filling / scaffolding). Even then, given that your illumina assembly is essentially not assembling (1kb N50 is significantly lower than a pacbio read length N50) it is highly unlikely that anything would be improved.
For a draft plant assembly there are no budget options, do you already have transcript data? it is generally much more cost effective for large genomes. Or as the previous poster points out, if you can reduce the problem (isolate organelles) and get higher coverage either for illumina or PacBio then an assembly will be much more successful.
Comment
-
Well given the length of PacBio reads it would be hard not to improve the assembly even with 1x coverage since many of the small contigs will be attracted by single long reads. If your budget was calculated some time ago it might be that you can get substantially higher coverage given that it nov can give ~1 Gb per cell. Not saying that low coverage illumina + low coverage pacbio will give you a good assembly though.
Comment
-
The problem is that there is a low, but nonzero chance of a single read being chimeric (blunt end ligation of adapters can also result in blunt end ligation of fragments), therefore without multiple observations it is impossible to validate the long range information in a single read.
Comment
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...-
Channel: Articles
04-22-2024, 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, 04-25-2024, 11:49 AM
|
0 responses
19 views
0 likes
|
Last Post
by seqadmin
04-25-2024, 11:49 AM
|
||
Started by seqadmin, 04-24-2024, 08:47 AM
|
0 responses
19 views
0 likes
|
Last Post
by seqadmin
04-24-2024, 08:47 AM
|
||
Started by seqadmin, 04-11-2024, 12:08 PM
|
0 responses
62 views
0 likes
|
Last Post
by seqadmin
04-11-2024, 12:08 PM
|
||
Started by seqadmin, 04-10-2024, 10:19 PM
|
0 responses
60 views
0 likes
|
Last Post
by seqadmin
04-10-2024, 10:19 PM
|
Comment