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Forum: Bioinformatics 08-05-2019, 05:10 PM
Replies: 3
Views: 753
Posted By SDPA_Pet
Illumina output questions

Hello, I am gonna do some Illumina 2X300bp amplicon sequencing. I suppose the platform (MiSeq) will run 600 cycles and time fixed. I just recently notice on the official website
...
Forum: Bioinformatics 07-12-2019, 03:49 PM
Replies: 10
Views: 1,559
Posted By SDPA_Pet
Hmm, interesting. I didn't realize MiSeq has...

Hmm, interesting. I didn't realize MiSeq has 2X150bp runs choice. Again, this is new sequencing center. My previous center for Amplicon sequencing only has 2X300bp or default is this settings. I am...
Forum: Bioinformatics 07-12-2019, 01:06 PM
Replies: 10
Views: 1,559
Posted By SDPA_Pet
"What does that mean?" -- This is new sequencing...

"What does that mean?" -- This is new sequencing center that I recently changed. It's first time that I used. "sloppy work" means they didn't do a good job and something wrong about the sequencing...
Forum: Bioinformatics 07-12-2019, 12:56 PM
Replies: 10
Views: 1,559
Posted By SDPA_Pet
I am sorry. I am not sure if I understand you. ...

I am sorry. I am not sure if I understand you.

These days MiSeq 300bpX2 is popular (default) and long reads also better for downstream analysis such as taxonomic assignment . It doesn't matter...
Forum: Bioinformatics 07-12-2019, 12:54 AM
Replies: 10
Views: 1,559
Posted By SDPA_Pet
Did my sequencing center did a sloppy job?

Hello, I have just got my 16S rRNA amplicon sequencing back. We use typical Earth Microbiome Project primers 515F-806R, which is about 290bp size of amplicon.

We requested MiSeq 300bp X 2 pair-end...
Forum: General 05-27-2019, 07:41 PM
Replies: 6
Views: 1,975
Posted By SDPA_Pet
Hi Thanks for you information. I am using a...

Hi Thanks for you information. I am using a different brand called Geneaid. However, the product comes with very simple protocol. It only says that I can clean up the DNA from 100bp to 10kb. It only...
Forum: General 05-26-2019, 06:58 PM
Replies: 6
Views: 1,975
Posted By SDPA_Pet
Hello AvantS, Thank you very much. Can you...

Hello AvantS,

Thank you very much. Can you give me the details (website link) of AMpure beads 1.2X. I am not sure what is 1.2 X? I heard Ampure beads XP. Are they same thing?

Thanks.
Forum: General 05-23-2019, 03:59 PM
Replies: 6
Views: 1,975
Posted By SDPA_Pet
NGS clean up kit

Hello, I am want to purify some PCR amplicons (16S rRNA) for Illumina sequencing.

I have many samples and I did my PCR in 96-well plates. I was told the best way to do it is using magnetic beads...
Forum: Illumina/Solexa 05-09-2019, 02:22 PM
Replies: 5
Views: 1,354
Posted By SDPA_Pet
Thanks. I just want to make sure if the...

Thanks. I just want to make sure if the sequencing primers for bridge amplification step for Illumina sequencing? Am I right? I just want to know which step the sequencing center would use these...
Forum: Illumina/Solexa 05-09-2019, 01:06 PM
Replies: 5
Views: 1,354
Posted By SDPA_Pet
Thanks. Do you have any idea about the so-called...

Thanks. Do you have any idea about the so-called sequencing primers?
Forum: Illumina/Solexa 05-07-2019, 05:47 PM
Replies: 5
Views: 1,354
Posted By SDPA_Pet
Basic questions about Illumina Amplicon sequecing

Hi, I've just moved to a new lab. We normally do barcoded bacterial 16S rRNA and fungal ITS sequencing. However, this new lab's protocol and primers design are quite different from my previous lab's....
Forum: Illumina/Solexa 04-14-2019, 03:07 PM
Replies: 1
Views: 792
Posted By SDPA_Pet
Illumina MiSeq 16S sequencing EMP protocol questions

Hi, I'm going to do some 16S barcoded PCR following the EMP standard protocol here (http://www.earthmicrobiome.org/protocols-and-standards/16s/).

The PCR potocol seems straight-forward. However, I...
Forum: Illumina/Solexa 02-20-2019, 06:37 PM
Replies: 5
Views: 942
Posted By SDPA_Pet
ôR1 and R2 of the fragment are in oppossite...

ôR1 and R2 of the fragment are in oppossite direction". -- I know this. However, my question is when they sequence R1 and R2, do they have to use two primers? For example, R1 uses forward primer and...
Forum: Illumina/Solexa 02-20-2019, 06:09 PM
Replies: 5
Views: 942
Posted By SDPA_Pet
I don't quite understand, but I didn't do PCR. I...

I don't quite understand, but I didn't do PCR. I gave them gDNA. They did the PCR and sequencing for me.

They didn't give me raw fastq files such as R1 and R2. They only give me a combined fasta...
Forum: Illumina/Solexa 02-20-2019, 03:09 PM
Replies: 5
Views: 942
Posted By SDPA_Pet
MiSeq amplicon sequencing Primers?

Hi guys,

I sent some soil samples to a commercial sequencing center. I do fungal ITS sequencing (300bpX 2 paired-end) Miseq.

As you know, I use forward and reversed primers when I did PCR. I...
Forum: General 02-20-2019, 02:53 PM
Replies: 5
Views: 1,611
Posted By SDPA_Pet
Thanks. This is what I want.

Thanks. This is what I want.
Forum: General 02-19-2019, 12:31 PM
Replies: 5
Views: 1,611
Posted By SDPA_Pet
Hi GenoMax, I wish I could post a pic. I...

Hi GenoMax,

I wish I could post a pic. I left my previous lab and moved to a new lab. I am setting up a new lab. I want to buy some, but I don't know the name. So, I can't google it. I have to...
Forum: General 02-18-2019, 06:22 PM
Replies: 5
Views: 1,611
Posted By SDPA_Pet
Does anyone know the name of this PCR rack?

Hi, I moved to a new lab. My old lab has some special PCR "clamping" Rack. I want to buy some, but I can't find it online. I don't know the official name, so I can't google for it.

I can describe...
Forum: Bioinformatics 09-06-2018, 07:37 PM
Replies: 0
Views: 756
Posted By SDPA_Pet
Some questions about KO classification

Hello, I have some questions about the KO classification.

I have a KO function abundance table. It will give me the basic level of KO funcitons ( the lowest level). Results like this:

#OTU ID...
Forum: Illumina/Solexa 08-18-2017, 09:01 AM
Replies: 5
Views: 1,894
Posted By SDPA_Pet
Thanks, do you know how much is MiSeq roughly....

Thanks, do you know how much is MiSeq roughly. For individual research lab (not sequencing lab), which is better?
Forum: Illumina/Solexa 08-18-2017, 08:47 AM
Replies: 5
Views: 1,894
Posted By SDPA_Pet
Price of sequencer

Hello, how much is MiSeq (include service. warranty)? I saw Illumina has MiniSeq online. Has anyone used it before? How much is MiniSeq?

Ben
Forum: Bioinformatics 08-16-2017, 10:39 AM
Replies: 1
Views: 1,158
Posted By SDPA_Pet
Should I worry about the direction of my reads

Hello, I recently got my metagenomic illumina sequencing back (hiseq). There is one sample that I would like to do some blast. Since it was sequenced using paired-end sequencing (I have two fastq...
Forum: Bioinformatics 04-12-2017, 12:27 PM
Replies: 1
Views: 776
Posted By SDPA_Pet
What percentage should I use?

Hi, I have some results from BLASTX. If I set % of identity to 60%, I will loss more than 50% of reads (results), compare to identity cutoff to 30%.

Should I use 30%? Is there any rule of thumb?...
Forum: Bioinformatics 04-06-2017, 04:03 PM
Replies: 3
Views: 973
Posted By SDPA_Pet
Can we sequence RNA directly

Hello, couple of years ago, I was in a NGS workshop. I was told some company was trying to develop some kits, so we can do metatranscriptomics without building cDNA libraries. I was wondering if this...
Forum: Bioinformatics 03-30-2017, 10:38 AM
Replies: 13
Views: 1,991
Posted By SDPA_Pet
Hi GenoMax, Thanks. BTW, do you know is...

Hi GenoMax,

Thanks. BTW, do you know is there any good forum to discuss R application. Bioconductor.org focus on molecular bioinformatics. I need some forums such as help people with plots...
Showing results 1 to 25 of 222

 


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