The glyceraldehyde 3-phosphate dehydrogenase (gapA) gene codes for a protein involved in the glycolytic pathway and is commonly used in Real-Time RT-PCR quantification studies as housekeeping gene. In this work we cloned and sequenced the full-length gapA gene from Flavescence dore´e phytoplasma (FDp). A _35 kDa recombinant GapA protein was overexpressed in Escherichia coli, purified and used as antigen to raise anti-GapA rabbit polyclonal antibodies. The antiserum detected the GapA protein by western blot analysis of total protein extracts of FDp-infected experimental host (Catharanthus roseus) and grapevine plants collected in the field. We also developed an FDp-specific gapA Taqman Real-Time RT-PCR assay suitable for quantification overtime of gapA mRNA in infected plants.
Cloning of the glyceraldehyde 3-phosphate dehydrogenase gene of Flavescence dorée phytoplasma and development of serological and molecular tools for studying its expression.
PALMANO S;TURINA M
2010
Abstract
The glyceraldehyde 3-phosphate dehydrogenase (gapA) gene codes for a protein involved in the glycolytic pathway and is commonly used in Real-Time RT-PCR quantification studies as housekeeping gene. In this work we cloned and sequenced the full-length gapA gene from Flavescence dore´e phytoplasma (FDp). A _35 kDa recombinant GapA protein was overexpressed in Escherichia coli, purified and used as antigen to raise anti-GapA rabbit polyclonal antibodies. The antiserum detected the GapA protein by western blot analysis of total protein extracts of FDp-infected experimental host (Catharanthus roseus) and grapevine plants collected in the field. We also developed an FDp-specific gapA Taqman Real-Time RT-PCR assay suitable for quantification overtime of gapA mRNA in infected plants.I documenti in IRIS sono protetti da copyright e tutti i diritti sono riservati, salvo diversa indicazione.
