I don't think that question's been answered. But, you might want to look at this paper, which compares V4 to full-length:
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Suppose I can the sequence of the full 16S gene at high accuracy, which region(s) can give me the closest discriminating power to the whole gene sequence?
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Primers and barcodes for V1-V2
Hello,
I was wondering whether any of you has sequenced using Illumina primers for V1-V2 region....Does anyone have a list of barcodes I could used? We need to focus on this region, since it is the best approach to identify sp within oral samples. ...
Many thanks
Elisa
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Originally posted by fibar View PostBut what has happened in the field during the last year? Are there improved methods to deal with this problem?
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I want to sequence the V4 region using illuminas 16S metagenomic sequencing library preparation protocol (dual indexing approach using nextera indexes). Anyone have experience doing this?
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We have been using the UPARSE workflow for some time now and are in general very happy with it (http://drive5.com/uparse/).
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Hi everyone,
I go back to Pat Schloss: http://blog.mothur.org/2014/09/11/Wh...nce-matrix%3F/
If you don't de-noise properly (ie. fully overlapped reads), you get too many unique sequences. Hence, you end having a huge matrix downstream.
But what has happened in the field during the last year? Are there improved methods to deal with this problem? MadsAlbertsen, which method do you use?Last edited by fibar; 12-15-2015, 12:39 PM.
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Microbial profiling is new to me but I understand you can now combine variable regions due to the longer read lengths on the MiSeq. Has anyone combined regions for 18S similar to 16S as described by Mads Albertsen?
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I would talk to your service provider (the people doing the actual sequencing) about your problems especially in regards to the phiX.
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Illumina Miseq V4-V5 16S rRNAsequencing
Hi all,
This is Richa. I have sent my 16s V4-V5 amplicons for sequencing with illumina MiSeq. But there is a problem. PCR fragments cluster very poorly. There are very less no. of sequences that we are receiving and 50% are phix.
What could be the reason ? How to troubleshoot. Kindly help...!!!!
Thanks.
Richa
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Originally posted by snetmcom View PostWhy is that the case? Is it just poor primer design?
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Originally posted by rhinoceros View PostTo everyone doing 16S with MiSeq, I very much recommend that you check JGI's related protocols. Your average 454-primers are going to introduce huge bias into 16S MiSeq sequencing, especially so while studying GC-rich communities.
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To everyone doing 16S with MiSeq, I very much recommend that you check JGI's related protocols. Your average 454-primers are going to introduce huge bias into 16S MiSeq sequencing, especially so while studying GC-rich communities.
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Hi,
I know I'm late to this conversation but we had the same issues. I managed to 'borrow' some Indexed primers for V4 and for V5-7 regions and compare just a few on a small set of my samples. The V5-7 gave better results.
I agree it's hard to know what to do and the best case scenario if you have money, sample and time is to try a few different ones! Sadly, that wasnt possible in my case but it worked out well.
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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...-
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Despite advancements in sequencing platforms and related sample preparation technologies, certain sample types continue to present significant challenges that can compromise sequencing results. Pedro Echave, Senior Manager of the Global Business Segment at Revvity, explained that the success of a sequencing experiment ultimately depends on the amount and integrity of the nucleic acid template (RNA or DNA) obtained from a sample. “The better the quality of the nucleic acid isolated...-
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