Illumina Sequencing of Rhyacophila sp.
Genomic libraries were prepared from 27.4 ng (Rhy_25_7) and 30.4 ng (Rhy_31_5) sheared gDNA, respectively, using the NEBNext Ultra II DNA Library Preparation Kit for Illumina (New England Biolabs, Ipswich, MA, USA) following the manufacturers manual. Adapters were diluted 1:10 as recommended for low input libraries, and size selection was omitted due to the low DNA amounts. A dual indexing PCR was run for 7 cycles on a Mastercycler (Eppendorf, Germany), using NEBNext Multiplex Oligos for Illumina (Dual Index Primers Set 1; New England Biolabs, Ipswich, MA, USA). After cleanup, library integrity was verified on a 2200 TapeStation with a High Sensitivity D1000 Tape (Agilent, Santa Clara, CA, USA), and shipped for 150 bp paired-end sequencing (ordering 30 Gbp output) on a partial lane of an Illumina NovaSeq 6000 platform (San Diego, CA) at Novogene (Cambridge, UK).