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Hello everyone,
I am trying to do a library of FACS sorted T-cells using the new Nextera DNA Flex Library Prep kit to perform ATAC-seq.
We are using adapters ordered from Sigma that have the same sequence as the Illumina ones:
Ad1 - AATGATACGGCGACCACCGAGATCTACACTCGTCGGCAGCGTCAGATGTG
Ad 2.1 - CAAGCAGAAGACGGCATACGAGATTCGCCTTAGTCTCGTGGGCTCGGAGATGT
Ad 2.2 - CAAGCAGAAGACGGCATACGAGATCTAGTACGGTCTCGTGGGCTCGGAGATGT
I've already tested the following parameters:
- Tested different amounts of cells - 25k, 50k, 75k;
- Removed the final bead purification step and changed it by a MinElute column purification - when I performed this, my Bioanalyzer profile showed an enormous amount of adapter dimers and no library amplification at all;
Finally, Illumina had the following information in their page for the discontinued Nextera DNA Library Prep Kit:
However, they've since made that page redirect to the new kit and now that information is no longer available. Has anyone been able to make the kit work?
I am trying to do a library of FACS sorted T-cells using the new Nextera DNA Flex Library Prep kit to perform ATAC-seq.
We are using adapters ordered from Sigma that have the same sequence as the Illumina ones:
Ad1 - AATGATACGGCGACCACCGAGATCTACACTCGTCGGCAGCGTCAGATGTG
Ad 2.1 - CAAGCAGAAGACGGCATACGAGATTCGCCTTAGTCTCGTGGGCTCGGAGATGT
Ad 2.2 - CAAGCAGAAGACGGCATACGAGATCTAGTACGGTCTCGTGGGCTCGGAGATGT
I've already tested the following parameters:
- Tested different amounts of cells - 25k, 50k, 75k;
- Removed the final bead purification step and changed it by a MinElute column purification - when I performed this, my Bioanalyzer profile showed an enormous amount of adapter dimers and no library amplification at all;
Finally, Illumina had the following information in their page for the discontinued Nextera DNA Library Prep Kit:
The Nextera DNA Library Prep Kit has been obsolesced. For whole-genome sequencing, Nextera DNA Flex Library Prep Kit is the recommended replacement product. For ATAC-seq and other custom applications, contact your local Illumina Sales representative to order stand-alone components TDE1 Tagment DNA Enzyme (Catalog No. 15027865) and TD Tagment DNA Buffer (Catalog No. 15027866). Illumina remains committed to providing you with high-quality support and service.
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