Total Newbie and need help/direction

[browntree]

Hi all,
I'm so pleased to be part of the community and I hope you can help me. Our company have just purchased a Miseq analyser and I need to get a handle on how it works and the protocols for setting up mRNA sequencing analysis, could you please point me in the direction. I should be looking to understand about sample prep and isolation of mRNA from total RNA. mRNA integrity analysis and anything else you may find helpful to get me started. I hope you wont mind me asking stupid questions (I'm sure there will be a lot) until I get a hang of what's going on. I think were also looking at getting an Agilant 2100 analyser too, so anything on that would be good as well. So anything on pitfalls or recommended equipment would be a huge help.

Many thanks

Brian

[Ben3]

Hi Brian,

Welcome to the community. There's no such thing as a stupid question, so feel free to ask away!

I can definitely help guide you on where to begin, especially with mRNA sequencing. I've done a lot on the NextSeq 500 and a bit on the MiSeq as well.

First thing, if you want to understand the MiSeq you should really start with Illumina's official resources. They have a comprehensive MiSeq System Guide (https://support.illumina.com/sequenc...mentation.html) which is a goldmine of information. It covers everything from setting up the system to troubleshooting. Even when I was comfortable with everything, I still kept a printout next to me for every sequencing run.

Illumina also provides detailed protocols for mRNA sequencing. Check out their mRNA-Seq Library Prep Guide (https://www.illumina.com/techniques/.../mrna-seq.html) for specifics on sample prep and isolation of mRNA from total RNA. This will also be really helpful for the library preps you're doing and understanding what you're doing to the sample.

The Agilent 2100 Bioanalyzer, you mentioned you might be getting, is perfect for mRNA integrity analysis. Again, check out Agilent's user manuals and application notes. They are a great resource to understand how to use it effectively and troubleshoot any problems you have.

If you want to learn more, check out videos on YouTube. When I first started I watched videos from Illumina on how to load the instrument and everything else. You can probably find a video there for anything else you want to learn. If you have any specific questions, just ask here and we should be able to help. Additionally, Illumina has a really helpful tech support and I've asked them a million questions over the years so don't be afraid to ask them for help, that's literally their job.

Good luck, and we're here to help if you need it!

[browntree]

Hi Ben,
thanks for the welcome and information, I will definitely follow your advice to get started and I've watched a few videos already. I suppose its going to take a while, but I'm sure that with the help of the community, I'll get the hang of it.
Much appreciated mate...thank you!

B