GSM6209232: In vitro haustoria, with Host, biol rep3 [H3_S11]; Cuscuta campestris; miRNA-Seq
Total RNA was extracted using bead-beater disruption of tissue coupled with Tri-Reagent (Sigma) per the manufacturer's instructions. 5ul of total RNA, at 100ng/ul, was used for library construction. Small RNA libraries were constructed using a random-splint-ligation protocol, essentially as described by Maguire et al. (2020) [doi 10.1093/nar/gkaa480] except with the inclusion of dual indices. Amplified libraries were cleaned using spin columns, quantified using UV spectrophotometry, and mixed in an equal-mass pool. The pool was size-selected using a BluePippin device set to recover fragments from 150-190bp (corresponding to insert sizes of 12 to 52 nucleotides). The size-selected pool was then sequenced on a NextSeq 2000 instrument using custom oligos and dual indexing.