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Old 09-09-2019, 11:06 PM   #1
lineras
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Location: Norway

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Question ATACseq library prep without commercial bought kit

Hi

I have previously done ATACseq with the use of the OmniATAC protocol (Corces et al 2017) and the Nextera DNA prep kit from Illumina.

The old Nextera DNA prep kit, as many of you know, is no longer on the market. However, Illumina sells the transposase enzyme separate from any library prep kit. It is also possible to buy the Nextera indexes separately.

From what I understand after reading the ATACseq papers by Buenrostro (2013 & 2015) and Corces (2017), in addition to the transposase and the Nextera indexes, I need a PCR mastermix to complete the list of components I need to make sequencing libraries.

Does anyone know if there are any details about the Illumina transposase adapters or the Nextera indexes that calls for a specific type of polymerase or a specific type of PCR master mix?

I have been thinking of trying out the NEBnext PCR Master mix. Or the new one from NEB: NEBNext Ultra II Q5*Master Mix (M0544S)


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Old 10-18-2019, 01:32 PM   #2
JackieM
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I work with trypanosomes, so I used the Muller 2018 protocol for AtacSeq, but I used the Q5 master mix (we prep a lot of standard NGS libraries, so it's what I had lying around) and had great results.
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Old 10-20-2019, 09:47 PM   #3
Meyana
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I always use the NEBNext High-Fidelity 2 PCR Master Mix (New England Biolabs, cat. no. M0541) for PCR when making ATAC libraries, I find it much more reliable than the one in the Nextera prep kit.

This is also the one suggested on the original ATAC papers.
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