Forum: Illumina/Solexa
04-24-2020, 06:05 AM
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Replies: 9
Views: 2,240
I agree that would be best to discuss these...
I agree that would be best to discuss these issues with your sequencing provider.
If you do choose to use staggered bases I recommend making an alignment to check for base diversity in the first...
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Forum: Illumina/Solexa
04-23-2020, 05:53 AM
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Replies: 9
Views: 2,240
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Forum: Sample Prep / Library Generation
04-01-2020, 06:47 AM
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Replies: 12
Views: 3,633
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Forum: Sample Prep / Library Generation
04-01-2020, 05:50 AM
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Replies: 12
Views: 3,633
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Forum: Bioinformatics
02-12-2020, 12:16 PM
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Replies: 2
Views: 1,163
The concatenate function of Seqkit...
The concatenate function of Seqkit [https://bioinf.shenwei.me/seqkit/usage/#concat] will concatenate sequences with matching ID's from two fasta files.
I converted your inputs into fasta format...
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Forum: Metagenomics
11-26-2019, 12:14 PM
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Replies: 2
Views: 3,884
Have you tried or considered using a blocking...
Have you tried or considered using a blocking oligo (such as one with a C3 spacer) designed to reduce amplification of the "host" plant tissue? A quick look at an alignment of nucleotide data might...
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Forum: Illumina/Solexa
11-26-2019, 12:07 PM
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Replies: 3
Views: 1,370
My guess is that you cannot use the "default"...
My guess is that you cannot use the "default" sequencing primers with your scheme.
You appear to have directly attached your gene-specific primer sequence directly to the i5 and i7 illumina...
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Forum: Oxford Nanopore
10-23-2019, 09:28 AM
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Replies: 2
Views: 5,894
I have done a few large volume CTAB-Chloroform...
I have done a few large volume CTAB-Chloroform DNA extractions in polypropylene without any noticeable issues with the nucleic acids. I was told by someone with experience in a forensics lab that...
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Forum: General
06-05-2019, 09:34 AM
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Replies: 3
Views: 2,718
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Forum: Sample Prep / Library Generation
05-24-2019, 09:32 AM
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Replies: 6
Views: 1,619
I tried to use limiting amount of SPRI beads...
I tried to use limiting amount of SPRI beads (Sera-Mag speedbeads and Commercial preparations diluted in their own buffer) for the normalization of amplicons prior to pooling without any success. I...
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Forum: Illumina/Solexa
04-23-2019, 05:42 AM
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Replies: 2
Views: 1,757
A few questions that might help: What DNA...
A few questions that might help: What DNA polymerase are you currently using for PCR? Are your 16S primers tailed for high-throughput sequencing?
If inhibition is the problem, dilution will often...
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Forum: Bioinformatics
04-02-2019, 06:07 AM
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Replies: 7
Views: 1,150
I will just echo cmbetts in saying that you...
I will just echo cmbetts in saying that you should always remove your oligo/primer sequences from Sanger and High-throughput sequencing data since they are artificial. You can sometimes find...
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Forum: Illumina/Solexa
03-18-2019, 06:03 AM
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Replies: 2
Views: 1,811
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Forum: Illumina/Solexa
03-13-2019, 07:24 AM
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Replies: 2
Views: 1,811
NovaSeq error rate?
I was wondering if anybody can quote the error rate for a NovaSeq? Even an idea of how it compares to MiSeq and HiSeq would be great as I can not seem to find any comparisons or hard numbers...
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Forum: Sample Prep / Library Generation
11-22-2018, 11:11 AM
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Replies: 4
Views: 1,303
Since a lot of dsDNA fluorescent dyes have an...
Since a lot of dsDNA fluorescent dyes have an excitation maximum of about 490 nm (blue light) and and emission at about 520 nm (green light) You can make your own blue light epi-illuminator to excise...
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Forum: Sample Prep / Library Generation
06-15-2018, 08:42 AM
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Replies: 5
Views: 2,148
If agarose gel electrophoresis is your only...
If agarose gel electrophoresis is your only option for fragment size distribution, GelStar is also very sensitive and might be more suitable than Sybr Gold for using in the gel itself.
Gelstar use...
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Forum: Sample Prep / Library Generation
06-04-2018, 12:34 PM
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Replies: 3
Views: 1,251
Hi again Sean,
Sorry for making that...
Hi again Sean,
Sorry for making that guess/assumption. I forgot that eukaryotes have 16S RNA as well.
So it does appear that you have very low base diversity, you have rapidly declining q...
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Forum: Sample Prep / Library Generation
06-01-2018, 05:56 AM
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Replies: 3
Views: 1,251
Hi Sean,
Aside from PhiX, are you taking any...
Hi Sean,
Aside from PhiX, are you taking any measures to increase diversity in your amplicon libraries such as staggering/offsetting and/or adding other loci?
At a glance, your cluster...
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Forum: General
05-31-2018, 05:28 AM
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Replies: 6
Views: 2,431
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Forum: General
05-30-2018, 06:40 AM
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Replies: 6
Views: 2,431
I find it can be hard to get more than 80-90%...
I find it can be hard to get more than 80-90% recovery and I sometimes get as low as 50%.
Here are a few suggestions you can try for improving recovery:
-Do not dry the beads too long. If you...
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Forum: Illumina/Solexa
09-06-2017, 07:12 AM
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Replies: 7
Views: 2,733
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Forum: Illumina/Solexa
09-06-2017, 06:19 AM
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Replies: 7
Views: 2,733
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Forum: Sample Prep / Library Generation
07-04-2017, 12:52 PM
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Replies: 9
Views: 2,610
Hi SNPsaurus,
I think that they are just...
Hi SNPsaurus,
I think that they are just referring to the typical single-digest RAD method where restriction is followed by sonication or another shearing method to generate fragment size...
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Forum: Sample Prep / Library Generation
07-04-2017, 06:13 AM
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Replies: 3
Views: 1,992
I have always thought of the NaCl (or another...
I have always thought of the NaCl (or another source of Na+ like Sodium acetate) as a co-factor required for an ethanol, isopropanol or PEG-base nucleic acid precipitation. NaCl might precipitate...
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Forum: Sample Prep / Library Generation
06-29-2017, 05:37 AM
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Replies: 9
Views: 2,610
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