Barcoding.... a lot of questions!!!!

[soleulloa]

Hi all,

I'm Soledad from Chile
I want to do amplicon sequencing, but I don't understand very well the neccesary kits.
My amplicons are 1400 bp length, so I think that I have to shear to make libraries. Do you know the ideal conditions of Bioruptor to obtain suitable fragments (150-250 bp) using 1400 bp DNA?

On the other hand, I have "ION DNA Barcode 1-16 kit" (Cat: 4468654) but I don't know which are the other kits that I need.
Also, searching in web, I found another kit: "ION Xpress Barcode Adapters 1-16 kit" (Cat: 4471250).

What's the difference? Which are the necessary kits to make a complete sequencing of amplicons?

HELP ME, PLEASE!!!!!
Thank you

Soledad
(sorry for my bad english)

[Marzanna]

Hi,
According to the "Ion Xpress™ Plus gDNA and Amplicon Library
Preparation":
To prepare high‐quality libraries from genomic DNA (gDNA) or amplicons for sequencing on the Ion
Personal Genome Machine™ (PGM™) System, use one or more of the following kits.
• Ion Plus Fragment Library Kit (Part no. 4471252). This kit includes reagents for:
– End‐repair of physically fragmented gDNA or unfragmented short (<250 bp) amplicons.
– Library preparation from the end‐repaired DNA.
• Ion Xpress™ Plus Fragment Library Kit (Part no. 4471269): This kit includes:
– Ion Shear™ Plus Reagents, for enzymatic fragmentation of genomic DNA or long
amplicons.
– The Ion Plus Fragment Library Kit (Part no. 4471252), for library preparation from the
enzymatically fragmented DNA.
• Ion Xpress™ Barcode Adapters 1–16 (Part no. 4471250)
– This kit includes P1 adapter and barcoded A adapters that substitute for the non‐barcoded
adapter mix supplied in the Ion Plus Fragment Library Kit.
– Barcoded library preparation is otherwise identical to nonbarcoded library preparation.

The "ION DNA Barcode 1-16 kit" (Cat: 4468654) is the old barcode kit now replaced by the new one.

For the full version of "Ion Xpress™ Plus gDNA and Amplicon Library Preparation" you should look at the Ion Community page. The Ion protocols change very often.

Cheers,
Marzanna