How Many Pass-Filter Reads Should I Expect From An Illumina Gaii Run?
3
2
Entering edit mode
11.3 years ago
Weronika ▴ 300

About how many pass-filter reads would you normally expect to get from a single-end 36bp Illumina GA2 run? Ten million, twenty, forty? I know nothing is guaranteed, but I have a feeling the ten million my lab is currently getting is on the low end of the scale, and I'd like to see what other people consider to be a reasonable number.

(Apologies if this question isn't sufficiently bioinformatics-oriented, but I figured BioStar would at least have a lot of people who see this kind of data often!)

next-gen sequencing illumina • 8.3k views
ADD COMMENT
0
Entering edit mode

I think the question is perfectly fine, since the filtering is done using bioinformatics methods. Upvoted :-)

ADD REPLY
0
Entering edit mode

Good to know, thank you! :-)

ADD REPLY
0
Entering edit mode

@avilella Good to know, thank you! :-)

ADD REPLY
4
Entering edit mode
11.3 years ago

You'll probably want to be a bit more critical, here. The number of resulting reads is a function of the number of clusters. If the number of clusters is too low, you get a high percentage of reads passing filters, but still a low number of reads. If the number of clusters is too high, you get a low number of reads, but a low percentage of reads passing filters. In our group, the former is the more likely problem; learning how much DNA to load is tricky, it seems. So, look at the number of clusters and compare that to what Illumina says you should be running.

ADD COMMENT
0
Entering edit mode

Yes, very good point! I was asking about the total number of "good" reads first because, well, if that's as good as most people get there's probably no need to dig. But since it's clearly too low, you're right, I should look into the details. Or get the non-bioinformatics people in my lab to do it. ;)

ADD REPLY
2
Entering edit mode
11.3 years ago
Ma_Ko ▴ 100

Hello.

At first, I am newbie to NGS, and our data is premitive one.[?] I think we can get about PF-cleared 30 - 35 millions clusters from one lane, when you use GA IIx with Truseq reagents.

This is a plot of cluster density by illumina's SAV (our first result). https://skitch.com/mako/ffcyc/null-status

The lane 2 produced 10145091, 11059614 and 11014741 seqeunces (reported by FastQC).[?] The condition of lane 2 were:

  • 3plex with TruSeq reagents and human exome.
  • 100 bp Paired-end

I hope this helps you.

ADD COMMENT
0
Entering edit mode

Good to know, thank you!

ADD REPLY
0
Entering edit mode
11.3 years ago
Weronika ▴ 300

Answer I got from our sequencing facility: "You could definitely get more reads out of a GAIIx run: typical post-filter read counts are 30 million reads or better, topping out a little less than 40 million."

ADD COMMENT

Login before adding your answer.

Traffic: 2469 users visited in the last hour
Help About
FAQ
Access RSS
API
Stats

Use of this site constitutes acceptance of our User Agreement and Privacy Policy.

Powered by the version 2.3.6