Hi everyone,
Illumina provides a list of primers to amplify with high taxonomic coverage the ITS1 region for further fungal sequencing, but I cannot find the exact amount necessary of each primer. I understand then that have to mix them equimolarly, am I right? Has anyone used them?
Thx!
Information about primers, concentrations and volumes are provided in user manual.
https://emea.support.illumina.com/content/dam/illumina-support/documents/documentation/chemistry_documentation/metagenomic/fungal-metagenomic-demonstrated-protocol-1000000064940-01.pdf
Dear Laia, you can use a mixture of forward primers containing overhang sequences (1 uM concentration) and reverse primers containing overhang sequences (1 uM concentration) and 5 ul from pool of f primers and 5 ul from pool of rev primers for each sample dna for amplicon PCR
Dear Laia,
When you synthesize a degenerated primer, default is to make all the variants equimolar, just as you thought. Take a look at my paper, where methodology of generating Illumina ITS1 libs is described: Article How Does Salinity Shape Bacterial and Fungal Microbiomes of ...
Best regards,
Marcin Golebiewski
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