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Questions related to Next Generation Sequencing
I am preparing some high quality genomic DNA for NGS and some of my sample have high RNA concentrations. I would like to treat them with RNAse, but I've had some previous problems of DNA...
02 June 2017 2,096 1 View
Hi, I would like to use NGS to quantify the levels of NHEJ after Cas9 cutting of a genomic locus. I have gDNA from pools of transfected cells, on which I run a PCR#1 of about 150bp for sequencing...
23 May 2017 7,505 1 View
I want to do DNA mutation analysis by NGS using Ion Torrent PGM sequencing instrument. I will make use of "Ion Ampliseq cancer hotspot panel v2" to amplify cancer mutation hotspots. May I know...
15 May 2017 3,054 3 View
Hello, we're considering perform functional studies of intronic variants in several genes - could anyone update me on the current trends, besides more conventional methods? (e.g. luciferase assay...
15 May 2017 8,531 3 View
How effective and reliable would "Ion AmpliSeq™ Cancer Hotspot Panel v2" be for identifying variants of oncogenes and tumor suppressors genes in xenograft tumor? Expecting comments from users of...
24 April 2017 7,604 2 View
PGM reproseq data analysis reg. Kindly help me to analyse PGM reproseq data analysis.
15 April 2017 9,412 1 View
Hi, I have some BAC sequences, each assembled as a unique scaffold, and corresponding to a region of interest. I also have scaffolds from an inaccurate genome reference that have been retrieved...
12 April 2017 8,757 2 View
Hello all, I'm trying to make GBS libraries for damselflies. I'm using Pstl enzyme and everything looks good, but when I analyze the library in the bioanalyzer, there is a strange pike at ~200pb,...
05 April 2017 226 3 View
We have tried genotyping this line using Jackson's protocol and a protocol found online, but the gels are always blank (primers and ladders visible). No amplification of even the wildtype band...
05 April 2017 9,235 1 View
Hello, I am doing 16S metagenomics using ion torrent PGM. I am trying to pick OTUs using Qiime; but in case of ion torrent, there are two sets of reads; reads with the forward or reverse primer at...
08 March 2017 7,859 8 View
Dear all, I am facing Subit issues, for mosquito genomic DNA measurements (for gDNA library preparation for NGS). The same sample was measured by 3 different people (in december 2015, august 2016,...
27 February 2017 2,867 5 View
Hi all, Can you suggest me a method to create a heat map based on actual data and not the relative percentages? I need it because I have different treatments with increasing the overall...
17 February 2017 1,034 3 View
Hi If I have 200-1000 SNPs I need to test on regular basis with clinically validated way ( direct-to customer service) which way I may choose e.g. custom made Microarray or Taqman probes ( I need...
14 February 2017 2,772 2 View
HCV genotyping using conventional methods
14 February 2017 3,415 10 View
I am planning to conduct p16Ink4a/Cdkn2a immunohistochemstry in cryosections of mouse tissues.Should you empirically know good antibody for this purpose, your suggestions would be much...
27 January 2017 9,454 4 View
Hello everyone, Could some please explain to me what do the data values in a WIG file mean? I have read the docmentaion about this format on the UCSC webpage and also checked for an answer on the...
17 January 2017 3,912 9 View
Using FastQC, I have checked the over all quality of the file. The output file contains much count of over represented sequences. I simply performed blast on these sequences, and surprisingly they...
29 December 2016 4,266 6 View
Dear all, I would like to compare next generation sequencing (NGS) protocols for Ion Torrent, Illumina and Pacific Biosciences. It would much be appreciated if anyone can provide me with their...
15 December 2016 3,161 1 View
I have identified by RNA-seq a set of genes which I want to study in more detail using three different cell lines and conditions. This would be about 100 genes or I could expand to 1500. Which...
14 December 2016 1,536 3 View
I'm interested in amplifying small quantities of genomic DNA (~2 ng/μl) for use in RadSequencing. The specimens include both fresh and 10+ year old sand flies (Lutzomyia) stored in ethanol at...
02 December 2016 5,138 1 View
I am currently optimizing PCR for amplification of the ITS2 region for fungi and V4 region for archaea/bacteria from environmental gDNA. This amplification is being done using degenerate primers...
01 December 2016 9,324 3 View
Input file (collapsed reads) is: >1-1TGCG>2-4ATAT>3-1TGGC>4-1TGAG>5-1TTCA I would like the output file as...
23 November 2016 6,024 7 View
Hi all, I would like to get some suggestions and clarification about dealing with singletons in NGS of PCR amplicons. in the mothur SOP, the QC steps include the following: initial...
23 November 2016 6,615 4 View
Hi, We sequenced genomes of ancestors/unevolved and evolved populations of a bacterial species. We would like to determine genomic variants (SNPs/small INDELs) of the evolved strains in comparison...
15 November 2016 7,535 2 View