Budget Justification
Personnel
Vijaya M. Lella (also known as L. Vijaya Mohan Rao), Ph.D. Principal Investigator, 3.6 calendar months effort and salary. The P.I. will devote 30% of his effort on the proposed grant. He will assume overall responsibility for the proposed project, which includes supervision of all personnel involved in the project, planning, the design of experimental approaches, the close review of the data, data analysis & interpretation, preparation of manuscripts, and decisions as to priorities of activities and effort. The P.I. will also help the post-doctoral fellows and the research associate in purification of FVIIa variants and their characterization.
Usha Pendurthi, Ph.D. Co-investigator, 3.0 calendar months effort and salary. Dr. Pendurthi will help in the overall experimental design of the proposed study, data analysis and manuscript preparation. In addition, Dr. Pendurthi will help the post-doctoral fellow and the research associate in generating novel FVIIa derivatives, including primer design, cloning, transfection and expression of the proteins.
Shiva Keshava, Post-doctoral fellow, Ph.D., 12 calendar months effort and salary. Dr. Keshava has been working in the laboratory for the last 4 years. He is well trained in animal procedures, including tail vein injection, saphenous vein bleeding, and needle puncture knee injury. He is also expert in the immunohistochemistry. He is responsible for performing majority of animal experiments described in the proposal, data analysis of these experiments, and preparation of draft manuscripts. To be named, Post-doctoral fellow, Ph.D. 12 calendar months effort and salary. He/she will be primarily responsible for generating rFVIIa variants, their purification and characterization. The fellow will be responsible for performing most of the experiments requiring cell model systems. He/she will also perform animal experiments described in the proposal. Anuradha Rajulapati, Research Associate, M.S., 6 calendar months effort and salary. Ms. Rajulapati is responsible for breeding animals and genotyping them. She will also assist the post-doctoral fellow in cloning and expression of recombinant proteins, and also in performing animal experiments and immunohistochemistry. She will be responsible for ordering supplies and maintaining inventory related to this grant. Materials and Supplies: $36,000/year. This is based our current expenditure related to this grant. The approximate break-up of the above cost is: Procurement of cells, culture media, and serum, $6, 000. Human and mouse endothelial cells, HEK cells, CHO cells, and fibroblasts. Each vials costs $200 to $500. The culture medium for endothelial cells costs about $100/500 ml. Plastic supplies, $6, 000: Plastic supplies include daily routine plasticware, and culture flasks and dishes for culturing cells, syringes for injections. PCR reagents/molecular biology reagents, $6,000. PCR reagents are needed for constructing mFVIIa variants and genotyping mice. Molecular biology reagents and kits are required for cloning and transfection. Clotting reagents, $6, 000. Clotting reagents include purified factor X, hVIIa, FVIII, thrombin and other reagents, including deficient plasmas, thromboplastin, APTT. Chromogenic substrates, Eliza assay kits and reagents, $6, 000. The Chromogenic substrate is required for measuring factor X activation and also determining APC levels in mouse plasma. Each chromogenic vial costs ~$650. ELIZA multiplex assay kits (each $1500) are required to measure multiple cytokines. Other Eliza assay reagents are needed to measure FVII expression levels and thrombin:antithrombin complexes levels. Chemicals, other reagents and glassware, $6,000. Animals, per diem charges, $30,000/year.
Personnel
Vijaya M. Lella (also known as L. Vijaya Mohan Rao), Ph.D. Principal Investigator, 3.6 calendar months effort and salary. The P.I. will devote 30% of his effort on the proposed grant. He will assume overall responsibility for the proposed project, which includes supervision of all personnel involved in the project, planning, the design of experimental approaches, the close review of the data, data analysis & interpretation, preparation of manuscripts, and decisions as to priorities of activities and effort. The P.I. will also help the post-doctoral fellows and the research associate in purification of FVIIa variants and their characterization.
Usha Pendurthi, Ph.D. Co-investigator, 3.0 calendar months effort and salary. Dr. Pendurthi will help in the overall experimental design of the proposed study, data analysis and manuscript preparation. In addition, Dr. Pendurthi will help the post-doctoral fellow and the research associate in generating novel FVIIa derivatives, including primer design, cloning, transfection and expression of the proteins.
Shiva Keshava, Post-doctoral fellow, Ph.D., 12 calendar months effort and salary. Dr. Keshava has been working in the laboratory for the last 4 years. He is well trained in animal procedures, including tail vein injection, saphenous vein bleeding, and needle puncture knee injury. He is also expert in the immunohistochemistry. He is responsible for performing majority of animal experiments described in the proposal, data analysis of these experiments, and preparation of draft manuscripts. To be named, Post-doctoral fellow, Ph.D. 12 calendar months effort and salary. He/she will be primarily responsible for generating rFVIIa variants, their purification and characterization. The fellow will be responsible for performing most of the experiments requiring cell model systems. He/she will also perform animal experiments described in the proposal. Anuradha Rajulapati, Research Associate, M.S., 6 calendar months effort and salary. Ms. Rajulapati is responsible for breeding animals and genotyping them. She will also assist the post-doctoral fellow in cloning and expression of recombinant proteins, and also in performing animal experiments and immunohistochemistry. She will be responsible for ordering supplies and maintaining inventory related to this grant. Materials and Supplies: $36,000/year. This is based our current expenditure related to this grant. The approximate break-up of the above cost is: Procurement of cells, culture media, and serum, $6, 000. Human and mouse endothelial cells, HEK cells, CHO cells, and fibroblasts. Each vials costs $200 to $500. The culture medium for endothelial cells costs about $100/500 ml. Plastic supplies, $6, 000: Plastic supplies include daily routine plasticware, and culture flasks and dishes for culturing cells, syringes for injections. PCR reagents/molecular biology reagents, $6,000. PCR reagents are needed for constructing mFVIIa variants and genotyping mice. Molecular biology reagents and kits are required for cloning and transfection. Clotting reagents, $6, 000. Clotting reagents include purified factor X, hVIIa, FVIII, thrombin and other reagents, including deficient plasmas, thromboplastin, APTT. Chromogenic substrates, Eliza assay kits and reagents, $6, 000. The Chromogenic substrate is required for measuring factor X activation and also determining APC levels in mouse plasma. Each chromogenic vial costs ~$650. ELIZA multiplex assay kits (each $1500) are required to measure multiple cytokines. Other Eliza assay reagents are needed to measure FVII expression levels and thrombin:antithrombin complexes levels. Chemicals, other reagents and glassware, $6,000. Animals, per diem charges, $30,000/year.