title: RNA-seq of Leschenaultia sp.: transcriptomic of adult
design: Specimens were collected alive always in pairs by active collection. One was identified at the species level, whenever possible, and the other was torn apart inside a cryogenic tube containing RNALater (Sigma-Aldrich) for storing. The strategy of collecting two individuals of the same species allowed better identification and RNA preservation of the individuals. RNA extraction was performed with a part of a single specimen using TRIzol (Invitrogen) following manufacturers information. We built the libraries using NEBNext Poly (A) mRNA Magnetic Isolation Module kits (New England BioLabs) to isolate only the poly (A) + RNA set. The cDNA library was prepared using NEBNext Ultra RNA Library Prep Kit (New England BioLabs) for Illumina. The necessary adapters and primers for sequencing were added using NEBNext Multiplex oligos for Illumina Index Primers Set 1 (New England BioLabs). RNA extractions were quantified using the Qubit RNA HS kit (Thermofisher Scientific) and DNA libraries with the Qubit dsDNA HS kit (Thermofisher Scientific). Extractions were realized at the Molecular Evolution Laboratory of the Department of Zoology (Institute of Biosciences - University of So Paulo - USP). The sequencing was performed with the NextSeq 500 platform (Illumina) at the Center for Supporting Research Facilities (CEFAP-USP) with NextSeq 500/500 High Output v2 kit (150 cycles) (Illumina).