SEQanswers

Go Back   SEQanswers > Sequencing Technologies/Companies > Illumina/Solexa



Similar Threads
Thread Thread Starter Forum Replies Last Post
Nextera exome kit cnicolet Illumina/Solexa 4 01-10-2017 03:12 AM
adaptors and primers Nextera kit: modified or not? Simone78 Sample Prep / Library Generation 7 07-29-2016 12:03 AM
Changing the Nextera XT kit protocol and other questions dfhdfh Illumina/Solexa 11 11-26-2013 01:41 AM
TruSeq or Nextera kit for library prep? rooty Sample Prep / Library Generation 12 10-29-2013 07:02 AM
Nextera Exome Enrichment kit dummyseq Illumina/Solexa 8 10-03-2013 06:46 AM

Reply
 
Thread Tools
Old 05-12-2015, 04:16 PM   #1
Angus's Microbiota
Junior Member
 
Location: San Francisco

Join Date: Dec 2013
Posts: 1
Default Using Nextera Indexes with Nextera XT Kit

Hello All,

I am planning to use a Nextera XT library prep kit (FC-131-1024) for my current project. The lab I work in already uses the other Nextera Kit (FC-121-1030) and therefore has the indices for that (non-XT) kit (FC-121-1011).

Does anyone know if the non-XT indices work for the XT kit? Illumina Technical Support said it isn't recommended, but they have heard of individuals "accidentally" using indices for the other kit and it still working. They couldn't recommended this option, but it "might" work.

I asked if the two kits (FC-121-1011 and FC-131-1001) were different and, of course, that information is proprietary.

Any help would be appreciated,

Angus
Angus's Microbiota is offline   Reply With Quote
Old 05-12-2015, 11:46 PM   #2
nucacidhunter
Senior Member
 
Location: Iran

Join Date: Jan 2013
Posts: 889
Default

I have used both of them interchangeably with customised Nextera using third party polymerase for PCR and results were similar. Sequence wise they are the same. I think the difference is that S5XX primers (for XT) have been resuspended in different buffer from N5XX (Nextera). This could be to compensate for different composition of tagged DNA resulting from two kits where Nextera tagged DNA is resuspended in RBS but XT tagmented DNA is not purified and contains salts from tagmentation and naturalisation buffer.
nucacidhunter is offline   Reply With Quote
Old 05-13-2015, 04:17 AM   #3
exo
Member
 
Location: Germany

Join Date: Dec 2012
Posts: 26
Default

Quote:
Originally Posted by nucacidhunter View Post
I have used both of them interchangeably with customised Nextera using third party polymerase for PCR and results were similar. Sequence wise they are the same. I think the difference is that S5XX primers (for XT) have been resuspended in different buffer from N5XX (Nextera). This could be to compensate for different composition of tagged DNA resulting from two kits where Nextera tagged DNA is resuspended in RBS but XT tagmented DNA is not purified and contains salts from tagmentation and naturalisation buffer.
Have you seen better results when you changed the polymerase?
exo is offline   Reply With Quote
Old 05-13-2015, 04:44 AM   #4
nucacidhunter
Senior Member
 
Location: Iran

Join Date: Jan 2013
Posts: 889
Default

I have worked with species that have 50% GC and performance was equal to Nextera. That polymerase is claimed to be more robust in amplifying low or high GC regions but I have not used it for amplifying tagmented DNA from organisms with extreme GC content.
nucacidhunter is offline   Reply With Quote
Old 11-11-2016, 01:39 PM   #5
JVGen
Member
 
Location: East Coast

Join Date: Jul 2016
Posts: 29
Default

Quote:
Originally Posted by nucacidhunter View Post
I have used both of them interchangeably with customised Nextera using third party polymerase for PCR and results were similar. Sequence wise they are the same. I think the difference is that S5XX primers (for XT) have been resuspended in different buffer from N5XX (Nextera). This could be to compensate for different composition of tagged DNA resulting from two kits where Nextera tagged DNA is resuspended in RBS but XT tagmented DNA is not purified and contains salts from tagmentation and naturalisation buffer.
Nuc, can you provide anymore information on using the Nextera XT indexes with the Nextera protocol? We're about to purchase a MiniSeq with the intention of running the reduced-volume reactions for Nextera library prep that I found published in S. Kryazhimskiy et al. I'd like to add more indexes per flow cell - I could run 384 and still have a surplus of coverage (I'm sequencing PCR-amplified viral genomes of 10 kb). However, the standard Nextera kit only has 96-indexes available, so I'm trying to figure out if I can use the Nextera XT indexes easily....

Thanks!
JVGen is offline   Reply With Quote
Old 11-11-2016, 03:26 PM   #6
SNPsaurus
Registered Vendor
 
Location: Eugene, OR

Join Date: May 2013
Posts: 390
Default

We use Nextera and Nextera XT indexes on Nextera libraries. Like you, we want to multiplex more than 96 samples in a lane. We were told by Illumina to do this, and we see no issues.
__________________
Providing nextRAD genotyping services. http://snpsaurus.com
SNPsaurus is offline   Reply With Quote
Old 11-11-2016, 03:37 PM   #7
JVGen
Member
 
Location: East Coast

Join Date: Jul 2016
Posts: 29
Default

Thanks SNP. You use the indexes interchangeably? No alterations? Just treat the XT indexes as the standard Nextera indexes?

Thanks much!
JVGen is offline   Reply With Quote
Old 11-12-2016, 12:06 AM   #8
SNPsaurus
Registered Vendor
 
Location: Eugene, OR

Join Date: May 2013
Posts: 390
Default

That's right. We prep a bunch of samples and just move from one set to the other with everything else the same.
__________________
Providing nextRAD genotyping services. http://snpsaurus.com
SNPsaurus is offline   Reply With Quote
Reply

Tags
illumina, nextera indices

Thread Tools

Posting Rules
You may not post new threads
You may not post replies
You may not post attachments
You may not edit your posts

BB code is On
Smilies are On
[IMG] code is On
HTML code is Off




All times are GMT -8. The time now is 11:10 AM.


Powered by vBulletin® Version 3.8.9
Copyright ©2000 - 2017, vBulletin Solutions, Inc.
Single Sign On provided by vBSSO