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The coat protein gene of Apple stem grooving virus (ASGV) was amplified by RT-PCR, cloned, sequenced and subcloned in the expression vector pMal-c2. This plasmid was used to transform Escherichia coli BL21c+ competent cells. The ASGV coat protein (cp) was expressed as a fusion protein containing a fragment of E. coli maltose binding protein (MBP). Bacterial cells were disrupted by sonication and the ASGVcp/MBP fusion protein was purified by amylose resin affinity chromatography. Polyclonal antibodies from rabbits immunized with the fusion protein gave specific reactions to ASGV from infected apple (Malus domestica) cv. Fuji Irradiada and Chenopodium quinoa at dilutions of up to 1:1,000 and 1:2,000, respectively, in plate trapped ELISA. The ASGVcp/MBP fusion protein reacted to a commercial antiserum against ASGV in immunoblotting assay. The IgG against ASGVcp/MBP performed favorably in specificity and sensitivity to the virus. This method represents an additional tool for the efficient ASGV-indexing of apple propagative and mother stock materials, and for use in support of biological and molecular techniques. O gene da capa protéica de Apple stem grooving virus (ASGV) foi amplificado por RT-PCR, clonado, seqüenciado e subclonado no plasmídeo pMal-c2. A capa protéica de ASGV foi expressa em Escherichia coli como proteína de fusão contendo um fragmento do gene da proteína de ligação a maltose de E. coli (MBP). Células bacterianas foram rompidas por sonicação e a proteína de fusão ASGVcp/MBP foi purificada por cromatografia de afinidade em resina de amilose. Anti-soros policlonais de coelhos imunizados com a proteína de fusão reagiram especificamente com extratos de maçãs (Malus domestica) cv. Fuji Irradiada e Chenopodium quinoa infetadas com ASGV em diluições de 1:1000 e 1:2000, respectivamente, em ELISA indireto. A proteína de fusão ASGVcp/MBP reagiu positivamente com anticorpos comerciais produzidos contra ASGV em testes de "immunoblotting". A imunogamaglobulina contra ASGVcp/MBP teve desempenho favorável em especificidade e sensibilidade ao vírus. Este método representa um instrumento adicional para a eficiente indexagem de ASGV em material propagativo e básico de maçã e como complemento de métodos biológicos e moleculares

Polyclonal antibodies to the coat protein of Apple stem grooving virus expressed in Escherichia coli: production and use in immunodiagnosis