Patent ID: 7303882
Filing Date: 2007-12-04
Classification: C12Q

Abstract:
1. The procedure using the specific biotin label nucleotide probes directed to the baculovirus DNA-binding protein (DBP) gene, Bombyx mori nucleopolyhedrovirus DBP gene (BmNPV DBP gene) and Autographa californica nucleopolyhedrovirus DBP gene (AcNPV DBP gene), for the quantitative determination of baculovirus titer, comprising: The procedure for amplifying the AcNPV DBP gene from the negative control (non-infected) and AcNPV infected samples, based on the polymerase chain reaction (PCR) in the presence of digoxigenin-11-dUTP using the synthesized oligonucleotides (SEQ ID NO. 1) and (SEQ ID NO. 2) for PCR: The procedure for the synthesis of three specific biotin label nucleotide probe 7, synthesized using SEQ ID NOs 1 and 1, probe 8, synthesized using SEQ ID NOs 4 and 5 and probe 9, synthesized using SEQ ID NOs 6 and 2, directed respectively to the three different fragments of 304 bp (fragment 1), 420 bp (fragment 2), and 328 bp (fragment 3) located in the coding region of the AcNPV DBP gene using the synthesized oligonucleotides (SEQ ID NO. 1) and (SEQ ID NO. 3) for fragment 1, (SEQ ID NO. 4) and (SEQ ID NO. 5) for fragment 2, (SEQ ID NO. 6) and (SEQ ID NO. 2) for fragment 3: The ELISA procedure for the determination of the titer of baculovirus using the obtained specific label nucleotide probes 7, 8, and 9, the obtained amplified Ac NPV DBP gene conjugated to digoxigenin-11-dUTP, and the immobilized streptavidin on polystyrene microtitration plates: