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szy0931 03-12-2019 10:43 PM

very basic questions about fragment and read
 
This is my understanding for fragment and read. Is my understanding correct?

If a fragment is 1000 bp and the read is 150 bp, only the first 150 bp of the fragment is sequenced.

How is the rest 850 sequenced?

My understanding is the initial DNA have many copies and different copies are fragmented at different positions. So, let's say in to a whole genome, the 1st copy is fragmented into 1st bp - the 1000th bp, the 1001st - 2000th bp, et al, while the 2nd copy is fragmented into 59th bp - 1058th bp, the 1059th - 2058th bp et. al, and the 3rd copy is fragmented into 74th bp - 1073rd bp, 1074th bp - 2073rd bp et.al, the forth, the fifth ......

So, it can result in read overlapping and the reads can be assembled based on the overlapping.

Is my understanding correct?

Gopo 03-12-2019 11:33 PM

The "fragment" of 1000bp in your example would be flanked by sequencing adapters in traditional Illumina sequencing, but really, you would call what you are calling the "fragment" as the insert sequence and the insert plus the sequencing adapters would be the entire fragment. I would agree with your understanding of how the rest of the 850bp are sequenced except that you rarely have a fragment (insert + sequencing adapters) size distribution with only fragments of 1000bp (rather it follows some distribution) consisting of smaller and larger fragments. Note that fragments resulting from PCR and then Illumina library prep can follow a tight length distribution.

Bukowski 03-13-2019 04:19 AM

short answer: yes

szy0931 03-13-2019 06:24 AM

Thank you very much!
I simplified the questions!

Quote:

Originally Posted by Gopo (Post 224605)
The "fragment" of 1000bp in your example would be flanked by sequencing adapters in traditional Illumina sequencing, but really, you would call what you are calling the "fragment" as the insert sequence and the insert plus the sequencing adapters would be the entire fragment. I would agree with your understanding of how the rest of the 850bp are sequenced except that you rarely have a fragment (insert + sequencing adapters) size distribution with only fragments of 1000bp (rather it follows some distribution) consisting of smaller and larger fragments. Note that fragments resulting from PCR and then Illumina library prep can follow a tight length distribution.



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