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Forum: 454 Pyrosequencing 07-24-2009, 10:12 AM
Replies: 10
Views: 9,592
Posted By hlu
The paper is quite outdated. For one thing, the...

The paper is quite outdated. For one thing, the paper is about GS-20, which is not compatible with current FLX and Titanium platform.

Titanium and FLX have different error profile than GS20,...
Forum: 454 Pyrosequencing 07-24-2009, 10:05 AM
Replies: 4
Views: 2,951
Posted By hlu
A combination of pooled BACs + 454 Titanium is...

A combination of pooled BACs + 454 Titanium is the way to go for draft assembly. Assembly is very challenging for big plants genome without BACs.

Plants have lots of repeat. Here long reads is...
Forum: 454 Pyrosequencing 07-24-2009, 09:38 AM
Replies: 4
Views: 2,951
Posted By hlu
University of Arizona recently published a paper...

University of Arizona recently published a paper using 454 to sequence rice. Their method is 454 Titanium + pooled BAC strategy. Some new school and some old school stuff. rice genome size is...
Forum: Bioinformatics 07-23-2009, 10:38 AM
Replies: 3
Views: 3,460
Posted By hlu
I suppose that you are trying to do assembly. ...

I suppose that you are trying to do assembly.

The 454 GS Assembler (newbler) software support hybrid assembly with Sanger + 454 data. You could do one big assembly including all the Sanger and...
Forum: 454 Pyrosequencing 06-19-2009, 12:57 PM
Replies: 5
Views: 3,796
Posted By hlu
Might want to contact software support on this...

Might want to contact software support on this issue? This sounds like a mis-behavior for sffinfo software.
Forum: 454 Pyrosequencing 06-19-2009, 12:54 PM
Replies: 14
Views: 4,284
Posted By hlu
Sometimes, the content of reads were very short,...

Sometimes, the content of reads were very short, 40bp, mostly formed from a pair of primer sequences.

This happened in the past in GS-FLX, and GS-20 times.
Forum: 454 Pyrosequencing 04-28-2009, 08:16 AM
Replies: 12
Views: 5,002
Posted By hlu
Yes, it is undocumented. But most of quality...

Yes, it is undocumented. But most of quality filter metrics can be modified with runAnalysisFilter.

DOT parameter can be put inside quality filter block anywhere.

Just have to remember that I...
Forum: 454 Pyrosequencing 04-27-2009, 07:47 AM
Replies: 12
Views: 5,002
Posted By hlu
The current pipeline does not produce all the...

The current pipeline does not produce all the reads before the filter (including dot). This is inconvenient in my opinion as well.


One way to get around this issue that current...
Forum: 454 Pyrosequencing 04-21-2009, 07:34 AM
Replies: 7
Views: 3,598
Posted By hlu
De Novo Assembly into large genome 50 Mb to 100...

De Novo Assembly into large genome 50 Mb to 100 MB, that is into insect range, beyond fungal genomes.

It requires lots of memory. 8Mb memory machine is not enough.

We are using 4 core, 32 MB...
Forum: 454 Pyrosequencing 03-27-2009, 02:44 PM
Replies: 10
Views: 3,388
Posted By hlu
By the way, I do believe in highly repetitive...

By the way, I do believe in highly repetitive regions, particularly short tandem repeat regions, the newbler would either not assemble them, label them as repeat reads, or put them into tons of...
Forum: 454 Pyrosequencing 03-27-2009, 01:50 PM
Replies: 10
Views: 3,388
Posted By hlu
Small contigs are useful for blast QC. But...

Small contigs are useful for blast QC.

But let's say even if the small contigs are not contaminations, they are less trust worthy, of poor quality.

We do QC on small contigs, But at end, we...
Forum: 454 Pyrosequencing 03-27-2009, 01:21 PM
Replies: 14
Views: 4,284
Posted By hlu
This looks like a shape of poor Titanium run. ...

This looks like a shape of poor Titanium run.

For good Titanium shotgun runs, the peak length usually at 500bp, and average length is around 400bp. Short length regions may have a hump, but...
Forum: 454 Pyrosequencing 03-24-2009, 12:56 PM
Replies: 5
Views: 3,606
Posted By hlu
All fasta mapping algorithm are heuristic as my...

All fasta mapping algorithm are heuristic as my understanding. blat is super-fast algorithm similar to blast, but less sensitive. It works at close related sequences with small SNP and difference. ...
Forum: 454 Pyrosequencing 02-20-2009, 11:56 AM
Replies: 11
Views: 6,473
Posted By hlu
I see. There is not a lot of public information...

I see. There is not a lot of public information on the algorithm in the past.

I have attended presentation by Newbler bioinformatics developers in the past. My understanding of this algorithm...
Forum: 454 Pyrosequencing 02-20-2009, 09:51 AM
Replies: 11
Views: 6,473
Posted By hlu
Things are moving fast. This paper obviously is...

Things are moving fast. This paper obviously is outdated.

Titanium shotgun is 400bp average length per read, 500 MB per run.

Titanium paired end 3kb, 8kb 20kb is going to be out soon. ...
Forum: 454 Pyrosequencing 02-20-2009, 09:14 AM
Replies: 11
Views: 6,473
Posted By hlu
I am not aware of any publication from 454/Roche...

I am not aware of any publication from 454/Roche on Newbler software.

However, Newbler (or officially called "GS De Novo Assembler") has very thick manual, which gives lots of detailed...
Forum: 454 Pyrosequencing 02-19-2009, 08:15 AM
Replies: 5
Views: 2,923
Posted By hlu
mingkunli 454 Titanium and most recent FLX...

mingkunli

454 Titanium and most recent FLX uses quality score algorithm are based on Broad Institute paper. 454 offline toolkit also has a script called "sffrescore" to allow you to rescore the...
Forum: 454 Pyrosequencing 02-18-2009, 10:16 AM
Replies: 0
Views: 2,395
Posted By hlu
Recommendation from 454 for Titanium PE/Shotgun

Genomeweb published latest recommendation from 454 for Titanium De Novo Assembly of shotgun/paired end reads.

This information was given by Jim Knight, 454's director of bioinformatics...
Forum: Bioinformatics 02-17-2009, 11:31 AM
Replies: 2
Views: 3,042
Posted By hlu
Have you looked at the newbler outout file...

Have you looked at the newbler outout file "454PairStatus.txt"?
This file prints all the pair reads status and coordinates for the assembly. It does not have all the nice pair info on ace file,...
Forum: 454 Pyrosequencing 02-17-2009, 11:11 AM
Replies: 5
Views: 3,606
Posted By hlu
I believe the latest version of GSMapper (2.0...

I believe the latest version of GSMapper (2.0 software for Titanium platform) support nimblegen Seq Cap out of box. I have seen presentation, but I have never used it myself.

I have used...
Forum: 454 Pyrosequencing 02-05-2009, 01:18 PM
Replies: 4
Views: 4,414
Posted By hlu
I am using early access 454 Titanium pairs end...

I am using early access 454 Titanium pairs end kits.

There are not out there yet public. I heard from 454 sales that they will be out possibly within a month. So it will be there soon.
Forum: 454 Pyrosequencing 02-05-2009, 09:20 AM
Replies: 4
Views: 4,414
Posted By hlu
We usually do 20x de novo assembly. This works...

We usually do 20x de novo assembly. This works for either FLX or Titanium.

Sounds that you are still doing FLX.

454 Titanium paired end data improved a lot compared to FLX paired end. Now...
Forum: Bioinformatics 01-21-2009, 10:26 AM
Replies: 8
Views: 9,070
Posted By hlu
I have used 32 bits for 454 data assembly in the...

I have used 32 bits for 454 data assembly in the past in 454 FLX, now I completely moved over to 64 bits linux. 32 bits machine just are too painly fully slow to do any thing right now.
...
Forum: 454 Pyrosequencing 01-13-2009, 02:20 PM
Replies: 6
Views: 5,959
Posted By hlu
I have not modified the default setting in...

I have not modified the default setting in gsMapper running.

gsMapper algorithm is similar to other assembly software (phrap), using the similar concept of "overlap" between reads to obtain...
Forum: Bioinformatics 01-09-2009, 03:02 PM
Replies: 7
Views: 4,297
Posted By hlu
GS De NOVO Assembler (newbler) has its issue with...

GS De NOVO Assembler (newbler) has its issue with repeat rich transcriptome reads.

MIRA claims to use repeat region and support EST for 454 data.

I have never used MIRA and I am looking at...
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