16 Questions 11 Answers 0 Followers
Questions related from Gary Sur
I'm using the Qubit fluorometer to quantify dsDNA Broad Range. According to the manual, it should not differ if I using "1-ul sample + 199-ul Working solution (Buffer & dye)" vs. "5-ul sample...
07 March 2023 3,788 3 View
I am using the Tecan MagicPrep automated platform to prepare NGS libraries of PCR products from DNA extractions. https://lifesciences.tecan.com/magicprep-ngs?p=tab--7 Despite its fairly recent...
30 January 2023 276 1 View
Hello, Because my data has non-normal distributions, I understand I need to run non-parametric tests. My data includes multiple samples from several distant collection sites. Under what...
23 January 2019 5,668 5 View
Hello, I'm using the command line to NCBI BLAST fungal DNA sequences (Table-C1, https://www.ncbi.nlm.nih.gov/books/NBK279684/). The default evalue is 10.0, but one paper I'm following used...
18 November 2018 1,166 3 View
Dear colleagues, I pooled together dozens of samples into one library, and sequenced them together. This yielded millions of reads, with the sequencer assigning each unique read-IDs. While the...
09 October 2018 5,649 4 View
Hello, I'd like to know your opinions on identifying fungal DNA sequences using the BLASTn with regards to Max Score, Total Score, Query Cover, E-value, and Identity match %? What are common...
07 July 2018 10,043 1 View
Hello, I understand there's disagreement regarding the similarity threshold for fungal OTUs, be it 95%, 97%, or 90-99%. In the literature, anyone of these threshold may be cast as adequate or too...
02 June 2018 3,529 4 View
Hello, I'm following Thermofisher's "Ion 16S Metagenomics Kit User Guide", and I want to ligate Ion P1 Adaptors, and an Ion Xpress Barcode onto dsDNA amplicons for ion torrent sequencing (p. 16,...
30 March 2018 2,123 2 View
Hello all, Previously after completing my PCRs, their electrophoresis gels indicated all PCRs had primer dimers. I am having difficulty ridding my PCRs of dimers as well as anything 100...
15 December 2017 3,000 4 View
Background: First, I want to sequence fungal DNA collected from samples of over 100 individual trees. Each PCR sample was amplified with an unique barcoded combination of forward and reverse...
13 December 2017 5,134 2 View
Hello all, I am trying to optimize my PCRs to amplify fungal DNA isolated from extracts of mostly plant DNA. I'm using ITS-1F and ITS-2R for my forward and reverse primers, respectively. Past...
30 October 2017 5,362 6 View
Hi all, I need help calculating the melting temperature (Tm) of barcoded primers for PCRs. I am using two barcoded primers to separate dozens of samples' DNA sequences. Using the equation...
07 September 2017 2,698 3 View
Hello all, I am using the Ion 16S Metagenomic Kit (Thermofisher Scientific) on an Ion Torrent PGM platform. However, instead of 16S primer, I am using ITS primers. I want to isolate and sequence...
14 June 2017 3,610 2 View
I'm assisting in a light microscopy study where I count the number of pollen grains in a sample, separating normal grains from malformed grains. The pollen are suspended and stained in cotton...
08 October 2016 7,759 3 View
Hi, I'm studying fungal endophytes isolated from frozen leaves, and I am following a protocol that submerges the leaves in a 0.5% bleach solution for two minutes for surface-sterilization....
20 April 2016 6,313 2 View
I'm extracting fungal DNA from plant leaves, and I'm using ITS-1 and ITS-2 as my forward and backward primers, respectively, to barcode fungal operational taxonomic units. I want to know the...
14 April 2016 7,262 2 View
