Glyceraldehyde-3-phosphate dehydrogenase (GAPDH) in the downregulation of Bamboo mosaic virus and its associated satellite RNA Replication

Binding of host proteins to viral replicase complexes has been demonstrated with a number of positive-strand RNA viruses (1,24), including bamboo mosaic virus (BaMV). In BaMV, chloroplastic phosphoglycerate kinase (PGK) (25) and HSP 90 (Huang et al., Unpublished data) have been reported to be necessary for efficient accumulation of BaMV. Proteins involved in the replication of satBaMV RNA have not yet been recognized.

The glyceraldehyde-3-phosphate hydrogenease gene (GAPDH) is an important enzyme housekeeping gene that catalyzes an important step in glycolysis and is found in all phylogenetic development. Genes can be extracted and isolated from plant gDNA using PCR. After cloning, the GAPDH gene is sequenced and finally analyzed by bioinformatics for further study. The grass studied through the experiment is as follows. Known as kangaroo glass Themeda Triandra is native to Australia and grown in all states and regions (Unkown, nd. Native Seeds). This grass is a perennial cluster growing to a height of 5 meters and a width of 0.5 meters. This special grass is gray / green leaves very rough, with seeds in their heads to make a very distinctive red / brown spike (Jennifer Liles, 2004).

Rabies virus (RABV) is a minus-strand RNA virus. Its genome is tightly packed with templates for transcription and replication of viral nucleoprotein (N), viral RNA-dependent RNA polymerase (L) and its cofactor phosphoprotein (P). We propose the molecular, structural and cellular aspects of transcription and replication of RABV. First, we summarize the features and molecular biology of the two RNA synthesis processes. We then explained the biochemical and structural data of viral proteins (N, P, L) and their interactions in the transcription and replication of viruses. Finally, we review the evidence that transcription and replication of rabies virus occurs in cytoplasmic inclusion bodies formed in RABV infected cells and discuss the role of this cell compartmentalisation.

Cells were lysed with protease inhibitor for 20 minutes at 4 ° C. in a buffer containing surfactant. Glyceraldehyde-3-phosphate dehydrogenase (GAPDH) was used as reference protein. After lysis, cell lysates were collected and centrifuged at 14,000 rpm for 10 minutes. The supernatant was transferred to a new tube and protein quantitation was performed using the Pierce BCA protein assay kit (Thermo) and 25 μg aliquots were loaded on a 10% gel for SDS-PAGE. After electrophoresis, proteins were transferred to a polyvinylidene difluoride (PVDF) membrane (Millipore). Membranes with EAAC1 or GAPDH exit at intermediate positions, blocked respectively, and incubated with different antibodies. Nonspecific binding sites were blocked by incubation for 1 hour at room temperature in Tris buffered saline containing 5% bovine serum albumin (BSA). After washing, blots were exposed to goat anti-rabbit IgG HRP diluted 1: 5000 with blocking solution for 1 hour at room temperature.