Abstract
The fragment of the VlhA1.2 gene was cloned from a Mycoplasma gallisepticum (MG) DNA library by serial PCRs after the same-sense mutagenesis of three TGA codons encoding tryptophan (Trp). Following transforming the generated plasmid of pKG-VlhA1.2, the recombinant VlhA1.2-GST fusion protein of 92 kDa was induced and recognized by anti-MG sera. After GST-affinity chromatographic purification, the VlhA-based colloidal gold immunochromatography assay (GICA) strips were generated. The GICA strips specifically detected anti-MG antibodies, but not antibodies against Mycoplasma synoviae and other positive sera against non-MG pathogens tested. The GICA strips were 128-fold more sensitive to detect anti-MG antibodies, as compared with traditional serological methods and were stored at 4 °C for 15 months without loss of their sensitivity and specificity. Analysis of sample revealed that, the GICA strips were highly sensitive, specific and stable for the on-site surveillance of MG infections by unskilled users.
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