MiSeq custom recipes

[ScottC]

Hi All,

Has anyone ever used a custom recipe on their MiSeq? I've seen a few mentions of them throughout the forums. I'd be interested to see one or to talk to anyone who has made or used them.

Cheers,

Scott.

[kwaraska]

How do you define custom? Since you can put in any parameters you want for the sequencing-what are you customizing that isn't there?

[ScottC]

Hi,

What do you mean by 'put in any parameters you want'... do you mean instrument- and run-configuration using MCS and IEM/sample sheet parameters?

When I say 'custom recipe', I'm talking about recipe files (or folders, or sets of XML files, as seems to be the case with the MiSeq), not instrument configuration through MCS and/or sample sheets (unless there's a way to control the system of which I'm not aware).

You can see the recipes in C:\Illumina\MiSeq Control Software\Recipe\

I'd like to be able to control the runs in a similar fashion to the way custom recipes could be created and run on the GAIIx.

The sorts of instrument behaviour that I'm talking about might include changing the reagent that is pumped at any given given step, changing the volume and dispense rate, setting the temperature of incubations, changing the order or the sequencing reads, changing the bottle/tube from which a reagent is pumped, controlling instrument hardware such as sippers and imaging, etc. (I don't mean that I want to do all of those things, but those are the types of things you might be able to manipulate in a GAIIx recipe for instance).

I've had a look at the standard MiSeq recipes and although I see some commands that would be useful, I don't see commands for all the things I'd like to be able to do.

Illumina tech support says that they're "unable to provide guidance" because they "haven't received training" in that area.

I've read a few things here and there that suggest that custom recipes can be used, but maybe complete control isn't possible via recipes. But, surely the instrument behaviour isn't hard-coded into MCS.

There's also a folder (on our MiSeq) called D:\Illumina\MiSeq Control Software\CustomRecipe\

That being said, I've never even seen a custom recipe for the MiSeq.

So... has anyone else heard anything different or tried to modify a recipe?

Cheers,

Scott.

[addyblanch]

Hi Scott,

I'm in the process of designing one with a collaborator. I would be interested to hear what you have done.

So far I'm aware that you can use custom primers in the empty wells on the MiSeq cartridge but regarding the actual recipe I'm still in the dark about.

Adam

[windhorse8]

Hi Scott,

I'm in the process of designing one with a collaborator. I would be interested to hear what you have done.

So far I'm aware that you can use custom primers in the empty wells on the MiSeq cartridge but regarding the actual recipe I'm still in the dark about.

Adam

Hi,
I am wondering if you were able to successfully design custom primers for Miseq. I am also planning to do similar thing. Wondering how did your go or if you can shed some knowledge about it ?

[ScottC]

Hi,

If you just want to use a custom primer, you can simply add these to the existing primer tube in the MiSeq catridge... as long as the current mix of primers that are in there won't interfere or prime elsewhere on your library construct.

Cheers,

Scott.

[windhorse8]

Hi,

If you just want to use a custom primer, you can simply add these to the existing primer tube in the MiSeq catridge... as long as the current mix of primers that are in there won't interfere or prime elsewhere on your library construct.

Cheers,

Scott.

Hi Scott,
Thanks for your reply.
I am wondering if you published your work ? I have been looking for publications ( preferably methods paper) that have used custom primers and adapters to prepare their amplicon libraries (without using Illumina kit) and run them successfully on Miseq. I can't use kit to make amplicons for my project. So far, no luck. Any advice ?
Thanks again.

[ScottC]

Hi,

A good place to start is the Illumina adapter sequence letter:

http://supportres.illumina.com/documents/documentation/chemistry_documentation/experiment-design/illumina-customer-sequence-letter.pdf

This is what you need for using custom primers with the MiSeq:

http://supportres.illumina.com/documents/documentation/system_documentation/miseq/miseq_using_custom_primers_15041638_b.pdf

There is also plenty of information on this site, and on various blogs and pages around the web that talk about designing custom primers and adapters. For example, you can search for the Earth Microbiome Project and you'll see the sequences they use to produce amplicons for 16S sequencing. You can change the 16S component of these primer to whatever you need.

Cheers,

Scott.