Dnase 1

The skin tissues during embryonic development were used as the material to extract total RNA. The skin was put into lysing matrix (MP Biomedicals, LLC) containing 1 mL TRIzol reagent and ground twice for 30 s (oscillation speed: 6 m/s) by tissue crushing apparatus (MP FastPrep-24TM). Total RNA was extracted according to the manufacturer’s protocol. The quality and quantity of RNA samples were checked by Spectrophotometer NanoDrop 2000 and denaturing agarose gel electrophoresis. All RNA samples were treated with DNAse-I (TOYOBO, Shanghai, China) for later use. The RNA integrity number was detected by Agilent Bioanalyzer 2100. The quality and integrity of all samples met the requirements of library construction (Supplementary Table S1).

Quantitative reverse transcription PCR (qRT-PCR) with specific primers was carried out to determine the relative abundance of Hc-maoc-1 transcripts in all key stages (i.e. L1s, L2s, iL3s, female L4s, male L4s, female adults and male adults) of H. contortus and some transcripts of candidate gene in Ce-maoc-1RNAi (RNA interference) C. elegans. In brief, total RNA was extracted separately from worms at different developmental stages of H. contortus and Ce-maoc-1RNAi C. elegans employing Trizol reagents (Invitrogen, Shanghai, China), followed by treatment by DNase I (Toyobo, Shanghai, China). First strand cDNA was obtained using ReverTra Ace-α (TOYOBO, Shanghai, China). Gene expression levels were determined by qRT-PCR (25 μl) using SYBR® Green qPCR Master Mix (TOYOBO) and an ABI 7300 thermal cycle. The qRT-PCR reaction procedure was 95 °C for 15 s, 60 °C for 15 s and 72 °C for 30 s for 40 cycles. The dissociation curve was generated under the following conditions: 95 °C for 15 s, 60 °C for 1 min, 95 °C for 15 s and 60 °C for 15 s. Each sample was employed in triplicate using tubulin as an internal loading control using specific primers (Additional file 1: Table S1). The mean threshold cycle (C_q) values were used for further analysis.

Total RNAs were extracted from the leaves or young panicles of the plants using the TRIzol (Invitrogen), and then reverse transcribed using ReverAce (TOYOBO). cDNA was synthesized from 2 μg of total RNA treated with DNase I (TOYOBO) and used as template.

qPCR with specific primers (Hc-clec-160 QF-Hc-clec-160 QR and Ce-clec-160 QF-Ce-clec-160 QR) (Table 1) was carried out to determine the mRNA levels in different stages of growth and development of H. contortus and C. elegans including L1, L2, L3, L4 and adults. Total RNA was isolated with Trizol reagents (Invitrogen, Shanghai, China) according to the manufacturer’s instructions and treated with DNase I (Toyobo, Shanghai, China) to remove the DNA. The qPCR reaction program included: one cycle at 50 °C for 2 min and 95 °C for 1 min; 40 cycles at 95 °C for 15 s, 60 °C for 15 s and 72 °C for 30 s; with a dissolution curve being produced in the last cycle. Each sample was repeated three times using the β-tubulin of H. contortus and actin-1 of C. elegans as internal reference genes and an average threshold (Ct) was taken for data analysis.