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Old 10-09-2017, 09:59 AM   #1
Junior Member
Location: Pennsylvania

Join Date: Sep 2017
Posts: 6
Default Combining/multiplexing Nextera and XT indexes

I am currently using Nextera DNA kits (96 samples, FC-121-1031) to prepare my genomic DNA libraries. I want to be able to multiplex more than 96 individual samples, however.

To do this, I plan to purchase the Nextera XT Index kit v2 'set D' (FC-131-2004), which includes adapter oligos (96 possible combinations for dual indexing) that are unique to the adapters that come with my standard Nextera DNA kit. Combining the two index sets would give me 40 total unique oligos (16 i5 adapters, and 24 i7 adapters, for total of 384 unique combinations) between the two kits.

Does anyone have experience combining standard Nextera DNA indexes with Nextera XT indexes? According to Illumina, they can't validate using XT indexes with the Nextera kit, though they say it generally works fine. What I want to know is, can I successfully combine the indexes from the different kits (i.e. have individual samples with a Nextera i5 index on one end, and an XT i7 index on the other), in order to multiplex 384 samples? I worry that because the XT kits deal with generally lower concentrations/volumes, that the adapter concentrations between the two kits may not match, although there are people on here who say the indexes from the two kits are interchangeable.

Any advice is appreciated!
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