Objective A venom allergen-like protein (VAP) gene was cloned from Bursaphelenchus doui with its expressions in organs and function in the nematode-host interaction determined.

Method A specific cDNA library on B. doui was constructed by solid-phase hybridization using cDNA from the anterior end as the tester and the posterior end of nematode as the driver. It was subsequently scrutinized by the reverse-northern blotting to obtain the sequence of positive clones. The full-length cDNA of Bd-VAP-1 gene was cloned using the rapid amplification of cDNA ends (RACE) technology, nucleotide and amino acid sequences analyzed using bioinformatics methods, and gene expression in organs localized by in situ hybridization.

Result A specific cDNA library of the anterior end of B. doui was successfully constructed. The full-length cDNA encoding a putative Bd-VAP-1 was identified by the reverse-northern blotting, sequencing, and RACE technologies. The open reading frame of Bd-VAP-1 was 644 bp in length and consisted of two extrons and one intron. The coding region, 609 bp in length, encoded a 202-amino acid protein with an average molecular weight of 22.32 kD and theoretical isoelectric point of 4.86. The Bd-VAP-1 protein was highly similar in sequence to the MIF homologous proteins of B. xylophilus and B. mucronatus. A secretory protein, it contained a typical CAP conservative domain and a signal peptide without a transmembrane structure. The gene expressed predominantly in the esophagus glands of B. doui as shown by the in situ hybridization.

Conclusion Bd-VAP-1 was successfully cloned and speculated to have its coding protein synthesized in the esophageal gland of B. doui and excreted to affect the host.