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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
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thanks. So if you have RNAseq data from an Hiseq 2500 then you should specify Phred+33 in tophat?Comment
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That is the default.Comment
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great, thanks again!
gComment
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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
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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
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Love this - good data definitely starts from good input, and poor input can only give relatively poor data. I particularly like the mention of Nanodrop/absorbance based methods for quantification. It's such a toss up if you'll get an accurate reading or what amounts to a randomly generated number, and a lot of library/sequencing related issues can be traced back to poor quant.07-01-2026, 11:43 AM
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