solexa genomic adapter and pictures?

[weizhu]

Seqgirl123,

I don't know of any images from Illumina (I've looked too) but I have attached a schematic of the genomic library prep (for single end sequencing libraries) which I created myself so I could understand how it worked. Hope it helps.

I am a newcomer of this forum. could anyone tell me what I shall do to view the attachment?

Thanks,

Wei

[bmcjc]

I want to sequence my PCR product on Solexa (it's a long story). Will it be feasible to PCR amplify with primers that has the Illumina's long PCR primer sequence extended to the 5'end so I don't have to do adaptor ligation again?

[wzhang25]

It is very useful. Have another quetions about ChIP-Seq or genomic DNA-Seq library construction. Does anyone know the sequence details in adaptor oligo mix and primer 1.1 and 2.1 from the Kit for ChIP-seq or genomic DNA-seq preparation from Illumina.Thanks a lot. wzhang25

[Chen Rui]

It's useful. Thanks a lot!

[Sadia]

Hello
Can anybody tell me if I have a template having internal biotin(The biotin is linked to the internal dT via a 10-atom spacer arm) wether it will be amplified in PCR or polymerase will block at the biotinylated nucleotide

[kaichen]

I wish I saw this first~~~~~

[seqgirl123]

kmcarr,

Thank you very much for sharing your diagram. I am also trying to learn. It's very informative and much better to look at than Illumina's colorful sticks to depict the mechanism!

I have another question if anyone knows. The other day, I had run a gel on PhiX DNA that went through the dA extension and ligation steps (my PhiX comes blunt ended so I skipped end repair). I also ran a negative control alongside where PhiX was ligated (no adapters, just PhiX ligated together using DNA ligase).

The negative control showed high molecular weight product while the other PhiX that went through extension & ligation steps was lower in molecular weight and ran a bit farther down the gel. My question is, why was the negative control in higher molecular weight than the other PhiX? I know it has something to do with the way it ligated, but I don't understand the underlying mechanism, if someone can explain? I would have thought that the dA extension and adaptor ligation would cause high molecular weight?

[kmcarr]

Seqgirl123,

I don't know of any images from Illumina (I've looked too) but I have attached a schematic of the genomic library prep (for single end sequencing libraries) which I created myself so I could understand how it worked. Hope it helps.

[seqgirl123]

I am trying to find slides from Illumina that depict these adapter structures along with mechanism of how they ligate to the genomic DNA after going through the end repair and 3' A extension steps. But Illumina's kept their slides pretty simple, not going into too many pictorials about the underlying mechanism.

Would anyone have illustrations of the exact adapter structure used for preparing genomic libraries? I know they are in some unusual structure (like a two pronged fork). Any pictures depiciting the entire mechanism of library construction would be good too (from end repair to ligation).