Yes, it is a requirement to include the BioAnalyzer information for the pools. Do not send Bioanalyzer information for each individual library in the pool. We prefer to have the Bioanalyzer of the pool. Please email it with your submission form.
Nucleotide diversity is the equal proportions of A, C, G and T nucleotides at each base composition in a sequencing library.
Illumina platforms require sufficient nucleotide diversity for effective template generation.
Sequencing a low diversity library is challenging, but can be accomplished by introducing a proportional amount of spike-in PhiX control in the library prior to the denaturation step. The lower the nucleotide diversity, the more spike-in PhiX control will be introduced into the library. The purpose is to balance the diversity of your library using the PhiX control. Another alternative is to use a well balanced library to spike-into the low diversity nucleotide libraries.
At the end, your library will be sequenced with optimal QC30 scores and higher percentage of passing filter reads. You will also see a percentage of PhiX control reads in the sequencing results that easily can be filtered out during data analysis.
We prefer you use a Bioanalyzer or a similar instrument to estimate the fragment size of your library. Please see the figure to help you to estimate your library fragment size. If you use a Bioanalyzer, use the region table on the area where the majority of your fragments are gathered.
The GSSC will accept libraries with less than 0.5% of adapter dimer contamination. If you are over this threshold, please do an AMPure bead clean up to remove the adapter dimers.
It is very important to minimize the percentage of primer dimer molecules in your libraries to less than 0.5% because adapter dimers are very competitive molecules, especially if you want to sequence your libraries in an Illumina HiSeq4000 platform.
As you increase the adapter dimer contamination in your final libraries, you increase the chance that adapter dimers will compete with your library fragments. With just a 5% adapter dimer contamination you will end up with 50% of your reads being an adapter dimer sequence. It is especially important to minimize the adapter dimer contamination for Illumina Hiseq4000 because of the Exclusion Amplification chemistry during cluster generation.
If you are a new GSSC customer make sure to create an account, which is free only for the first month. Please make sure to download the files you need because you will only have access to your data for 30 days, starting from when you received an email notification from Paul Billing, our Bioinformatics specialist.
We try to sequence your libraries as soon as possible. The sequencing queue time depends on which Illumina platform you use. Please check the table below for an idea of the sequencing queue times for each Illumina platform.
It is very important to plan your experiment ahead to make sure you cover an acceptable amount of reads. Please see the table below with all of the types of runs across different Illumina platforms. The data in this table is an estimation depending on the quality and performance of your library. At the GSSC we make an effort to give you the maximum amount of reads with the highest quality possible.