Hybrid sequencing reveals the full-length Nephila pilipes pyriform spidroin 1 (PySp1)

Araneid spider silk glands can spin seven silk types that have task-specific properties owing to the higher order structure of spider silk proteins. This gives silks superior potential as novel biomaterials. Nephila pilipes, the giant golden orb-weaver, is one of the largest spiders and spins silk with exceptional torsional deformation, toughness, and other properties to support its mass; further investigation relies on a complete amino acid sequence. However, there are no full-length N. pilipes spidroin sequences; in fact, across species, most sequences remain fragmentary because of repetitive region assembly difficulties in short-read sequencing. Here, we develop a hybrid sequencing method that utilizes short-read sequencing to identify seven spidroin terminals in N. pilipes, and long-read sequencing to confirm the full-length pyriform spidroin 1 (PySp1) gene. PySp1 is 11,181 base pairs, with a single exon encoding a 3,726 amino acid protein, the QQ(x)₄Qx motif, and lower repeat homogenization, distinct characteristics of genera Nephilinae PySp1. The full-length N. pilipes PySp1 sequences sheds light on spidroin evolution and demonstrates a helpful strategy to find full-length spidroins.