Tweet
Phred+64 refers to old format illumina (v.1.3+) quality calls. If you have recent data then it is going to be in Phred+33 (Sanger) format. See: http://en.wikipedia.org/wiki/FASTQ_format
-
-
thanks. So if you have RNAseq data from an Hiseq 2500 then you should specify Phred+33 in tophat?Comment
-
That is the default.Comment
-
great, thanks again!
gComment
-
I exceuted the tophat to align against the reference genome. I found the following files:
1. accepted_hits.bam
2. unmapped.bam
3.junction.bed
4. align.txt
here accepted_hits.bam, size is 1 KB while unmapped.bam has 200MB. Align.txt shows :
Reads:
Input : 528840
Mapped : 0 ( 0.0% of input)
0.0% overall read mapping rate.
Please help me to define the parameters to align and mapped against the heterologous genomes.Comment
-
I download the data from Ensembl. Is it any problem in it.
I exceuted the tophat to align against the reference genome. I found the following files:
1. accepted_hits.bam
2. unmapped.bam
3.junction.bed
4. align.txt
here accepted_hits.bam, size is 1 KB while unmapped.bam has 200MB. Align.txt shows :
Reads:
Input : 528840
Mapped : 0 ( 0.0% of input)
0.0% overall read mapping rate.
Please help me to define the parameters to align and mapped against the heterologous genomes.Comment
-
I’m not a sequencing expert. I’m a purification scientist who uses NGS to evaluate workflows my group develops. With this perspective, we think about the sample first and the NGS workflow second. The sequencer is an exceptionally honest reporter, but it can only report on what you give it, so whether you get clean, interpretable data from an NGS workflow is largely determined before you begin.
Here are nine questions we think about, in roughly the order they matter, before...Yesterday, 07:11 AM -
Data variability is still an issue in sequencing technologies despite the advances in reproducibility and accuracy of these platforms. But the problem does not originate in the sequencing itself, but in the previous steps, before the sample reaches the sequencer.
The first step is collection, followed by preservation and sample preparation for analysis. Most scientists overlook those steps, but not being careful might just be skewing the experiment’s results.
...06-02-2026, 10:05 AM -
With the launch of new single-cell sequencing platforms in 2026, the field stands at an exciting inflection point. This article surveys the most impactful advances in the field and discusses how they’re reshaping research in cancer, immunology, and beyond.
Introduction
Single-cell sequencing technologies have undergone remarkable advances over the past decade, transitioning from low-throughput experimental approaches to highly scalable platforms capable of...05-22-2026, 06:42 AM
Comment