UNITE DE PRODUCTION DES VACCINS (UPV) ...
UNITE DE PRODUCTION DES VACCINS (UPV)
LABORATOIRE NATIONAL DE L’ELEVAGE ET DES
RECHERCHES VETERINAIRES - BP 2057 - DAKAR-HANN
COMPTE RENDU DE MISSION
A I’ELS de San francisco et à l’Université de Californie, Davis, CA, USA
dans le cadre de transfert technologique en recombinaison génétique
pour la mise au point de vaccins recombinants
du 03 avril au 04 août 1999
par M. KONTE
chef de l’Unité de Production de Vaccins
Réf. UPV/Septembre 1999
COMPTE RENDU DE MISSION
*JC***
IDENTITE DU MISSIONNAIRE :
Prénom : Mamady
Nom :
KONTE
OBJET DE LA MISSION :
Formation dans les nouvelles méthodes de Biologie moléculaire, des gènes recombinants en
particulier et transfert de technologies en matière de mise au point de vaccins de type
recombinant. La formation scientifique et technique est précédée d’une participation à un
cours intensif d’anglais pour une remise à niveau linguistique.
PAYS ET LIEUX DE FORMATION
Etats-Unis d’Amérique (USA) : 4 mois : du 03 avril au 4 août 1999 :
Cours intensif d’anglais : ELS Language Centers de San Francisco en Californie : du 05 au 30
avril 1999.
Biologie moléculaire : Laboratoire international de Biologie moléculaire (ILMB), Université
de Californie, Davis : du 03 mai au 03 août 1999
ORGANISATION/PERSONNALITES RENCONTREES
L’organisation des cours intensif d’anglais est parfaite et efficiente.
La formation scientifique et technique a été réalisé à travers l’exécution conjointe de quatre
projets de recherche en attente, préalablement élaborés par les chercheurs du I’ILMB.
L’exécution de chaque projet est dirigé par le tuteur scientifique qui en est l’auteur. La
réalisation de l’ensemble des projets offre l’opportunité de pratiquer toutes les techniques de
biologie moléculaire dans leurs conceptions les plus récentes.
FINANCEMENT
Bourse AIEA, référence C6/SEN/98006R
NATURE DE LA MISSION
Voyage d’étude et de formation spécialisées
POINTS ESSENTIELS
Le financement AIEA est consenti dans l’optique d’un transfert de technologies des pays
développés aux pays en développement d’Afrique, en particulier, en application d’un
principe voulu, exprimé et constamment soutenu par le Directeur de I’ILMB, le Professeur T.
YILMA. Le choix de ce site de formation par 1’AIEA est des plus judicieux.
Les compétences acquises dans le domaine du génie génétique autorise la conception et la
réalisation d’outils biotechnologiques dans le domaine du diagnostic des maladies et
l’identification des agents microbiens ainsi que dans le domaine des vaccins nouveaux de type
recombinant au LNERV/ISRA.
RAPPORT DE MISSION
Deux rapports détaillés, un à mi-parcours et un autre en fïn de formation, ont été produits et
communiqués aux organismes de supervision de la formation (AIEA à Vienne, NRC à
Washington, et ILMB de U C Davis) : copies jointes ci-après.
,
Univers@ of California, Davis
International Atomic Energy Agency
VM-PMI-International Laboratory
P.O. BOX 100, A-1400, Vienna, Austria
Of Molecular Biology
Phone: 431-2600; Fax: 431-26007,
2079 Haring Hall, Davis, CA 95616, USA
Mail:Official.Mail@iaea.org
Phone : 530/752-8306
Fax : 5301752-1354
Institut Senegalais de Recherches
Agricoles (ISRA)-LNERV-
BP 2057-
Dakar (Senegal)- Fax: 221-832 21 18
FIRST TECHNICAL REPORT
Dr. Mamady KONTE
C6/SEN/98006R
Mail : mkonte@ucdavis.edu
FIRST TECHNICAL REPORT
**a****
FELLOW NAME : Dr, Mamady KONTE
FELLOWSHIP NUMBER : C6/SEN/98006R
ADRESS IN USA : Office : c/o Dr. Tilahun YILMA, UCD-ILMB-PMI-VM
2079 Haring Hall, Davis, CA 95616
Home : 717 Adeline Place, Davis, C!A 95616
TRAINING BEGINNING DATE : May 03,1999
TRAINING EXPECTED END DATE : August 03, 1999
TRAINING PLACE : International Laboratory of Molecular Biology
Department of Pathology, Microbiology and Immunology
School of Veterinary Medicine-University of California, Davis
2079 Haring Hall, Davis, CA 956 16 - USA
TRAINING SUPERVISOR : Tilahun YILMA, DVM-PhD, Director and Professor
Training tutors : Shabbir AHMAD, DVM-PhD, Assistant Professor (Research)
Paulo VERARDI, PhD, Post-graduate Researcher
Zhiyun MA, MS
David WADDELL
TRAINING SCHEDULE :
1- PRELIMINARY WORK :
May 03 through May 14,1999 : Administration aid, Housing, Documentation
May 17 through 21,1999 : Polymerase Chain Reaction (PCR) technics and attendance course on
Biosafety (may 18 at 1138 Meyer Hall, UC Davis)
Objective:
Diagnostic applications of PCR
PCR performing : Preparation (extraction) of DNA from cells samples
DNA polymerization by PCR
DNA gel electrophoresis
Tutor : David WADDELL
May 24 through May 28,1999 : PCR technics (cont’d)
Tutor : David WADDELL
II - FIRST AREA OF CONCENTRATION : Sub-cloming a gene
Project 1: Sub-cloning of IFN-gamma in a retrovirus.
Tutor : Zhiyun MA
Objective : Development of live attenuated vaccine by deletion mutation and insertion of human interferon
gamma gene.
June 01 through June 11.1999 :
Preparation of the insert : pLG-Hu-IFN-gamma
Preparation of the vector: pBRp429-3’Nef/NotI
Set up the ligation vector/insert (V-I)
Transformation of E. coli by the V-I
Selection of recombinant colony and extraction plasmid DNA
Recombination check by plasmid and insert isolation
III - SECOND AREA OF CONCENTRATION : Vaceinia Virus as an Expression Veetor
Project 2 : New selection Systems for Vaccinia Virus (Zeocin and Blasticidin)
Objective : TO test the feasibility of using two new antibiotics for selection of vaccinia virus recombinants
in eukaryotic cells. Vaccinia virus recombinants expressing the small zeocin (375 bp) or blasticidin (398
bp) resistance genes under the ~7.5 promoter Will be developed. Future studies Will test their potential use
as a fast and reliable selection method for recombinant vaccinia viruses.
Tutor : Paulo VERARDI
3.1 Project 2 Preliminary work : June 14 through July 02,1999 :
A)- Design primers for PCR cloning of the zeocin and blasticidin resistance genes with engineered Xma 1
(blasticidin) or Pin AI (zeocin) sites. Order primers.
B)- Perform Vent DNA Polymerase PCRs. Clean up PCRs for cloning.
3.2.Project 2 performing Week 1 : Julv 06 throuph iulv 09
C)- Cloning of zeocin and blasticidin resistance genes (PCR products) into the vaccinia virus transfer
vectors pSC 11 (Xma 1 site, under ~7.5). Sequencing of inserts.
3.3. Project 2 performing week 2 : Julv 19 throuph Julv 23,1999 :
D)- Transfection of BS-C-l cells infected with VV WR with ‘pSCl1Zeo’ and ‘pSC11Bsd’ transfer
vectors.
E)- Recombinant vaccinia virus plaque-purification (plaque assay #l) of ‘vWRTK-Zeo’ and ‘vWRTK-
Bsd’ with TK selection and beta-gal screening.
3.4. Project 2 performing week 3 : Julv 26 through Julv 30. 1999 :
F)- Recombinant vaccinia virus plaque-purification (plaque a.ssay#‘t) of ‘vWRTK-Zeo’ and ‘vWRTK-
Bsd’ with TK selection and beta-gai screening.
G)- Recombinant vaccinia virus characterization (histochimical staining, DNA analysis, zeocin and
blasticidin resistance gene expression).
IV - THIRY AREA OF CONCENTRATION : Isolating a gene of interest by PCR
Julv 12 through 161999
Project 3 : Isolating and cloning of vif gene of a retrovirus in a plasmid (pVL1393) to construct
pVL1393vif
Objective : Vaccine and pathogenesis studies
Tutor : Shabbir Ahmad & Zhiyun MA
V - FOURTH AREA OF CONCENTRATION : Bacnlovirus
Poject 4 : Construction, isolation, and production of a reeombinant baculovirus expressing a gene of
interest
Tutor : Shabbir AHMAD
June 14 throueh Julv 30,1999
A)- Insect ce11 culture
B)- Molecular cloning of gene of interest into transfer plasmid (From step IV)
C)- Transfection of insect cells with recombinant plasmid (pVL1393vif) and linear DNA of baculovirus.
D)- Identification and plaque purification of recombinant baculovirus
E)- Molecular analysis of baculovirus expressing the recombinant protein.
F)- Production and partial purification of antigen in insect cells and larvae.
G)- Development of ELISA
Last three steps Will be performed if time allows.
FACILITIES AND MAIN LAI3 EQUIPMENT
Electric Freezer, refrigerator
Centrifuges-microfuge-centrefr.
Spectrophometer
Water bath
Hot shaker
Magnetic stirrer
Powerpac 3000 with probe
Bio-Dot Apparatus
Rack LN storage system
Integrated speed vac
Sequencing gel
Sonic dismembratr
Maxima vac pump
New Brunswick sci. incubator shaker
Sterilization equipement
Biological Safety cabinet wl3 petcocks
1
,
Vortex-genie mixer
Napco vacuum oven
Revco Upright 20.2 CU. Ft. freezer
Napco sterilizer/autoclave w/dryer
Incubator
Thermocycler
Ultra violet transilluminator
Electrophoresis material
Electrophoresis photo material
Pipets
Ice machine
Pro-pipets
Others lab material
Chemical products and enzymes supplies
ELISA computer system Iecturer
Computers
THE FELLOW
THE TRAINING SUPERVISOR
Dr. M. KONTE
Pr. T. YILMA
‘University of California, Davis
International Atomic Energy Agency
VM-PMI-International Laboratory
P.O. BOX 100, A-1400, Vienna, Austria
of Molecular Biology
Phone : 43 l-2600 ; Fax : 43 l-26007
2079 Haring Hall, Davis, CA 95616, USA
Mail : Ot&ial.Mail@iaea.org
Phone : 5301752-8306
Fax : 530/752-1354
Institut Sénégalais de Recherches
Agricoles (ISRA)-LNERV, BP 2057
Dakar-Hann (Sénégal)
Tél.:2218322762;Fax:2218322118
FINAL TECHNICAL REPORT
Dr. Mamady KONTE
C6/SEN/98006R
Mail : konte@eIsmaiI.com
FINAL TECHNICAL REPORT
*****JC
FELLOW NAME :
Dr. Mamady KONTE
FELLOW NUMBER : C6/SEN/98006R
ADRESS IN SENEGAL : ISlWUPV
LNERV, BP 2057, DAKAR-H,ANN
Tel.:2218322762;Fax:22183221
18
Mail : konte@!elsmail.com
TRAINING BEGINNING DATE : May 03,1999
TRAINING END DATE : August 03, 1999
TRAINING PLACE : International Laboratory of Molecular Biology
Department of Pathology, Microbiology and Immunology
School of Veterinary Medicine, University of Califomia, Davis
2079 Haring Hall, Davis, CA 95616, USA
TRAINING SCHEDULE :- 1. Diagnostic applications of PCR
-
2. Sub-cloning of IFN-gamma in a retrovirus
-
3. Vaccinia virus as an expression vector : new selection systems for
Vaccinia virus (Zeocin and Blasticidin)
-
4. Isolating a gene of interest by PCR.
-
5. Construction, isolating and production of a recombinant baculovirus
expressing a gene of interest.
SCHEDULE IMPLEMENTATION :
Project # 1 : 1 had the oppotunity to adapt my knowledge (since 1994) to the new approach of PCR
performing and the new equipment actually in use (new thermocycler in particular).
Project # 2 : The objective was the development of a li.ve attenuated vaccine against HIV-2 by
deletion mutation and insertion of human interferon gamma gene.
The technology has been completely perfonned but we doesn’t get the expected result because both
Xhol and KpnI enzymes haven’t allowed to isolate the insert @LG-Hu-IFN-gamma).
SO, research Will
continue to fmd the appropriated restrictions enzymes.
In conclusion, 1 cari say that 1 possess now this technology and 1 cari do it again completely by
myself. That’s positive. However, the designing technic of approptiated primers and restriction
enzymes must be repetead with trainers.
Project # 3 : Obective : to test the feasibility of using two new antibiotics for selection of vaccinia
virus recombinants in eukaryiotic cells and after, to study their potentiel use as a fast and reliable
selection method for recombinant vaccinia virus.
This technic has been performed, since ptimers designing to first plaque assays. The short time
allowed have not permit to perforrn eompletely the necessary others plaque assays and the next stape
which is the study of potential use as selection method for recombinant vaccinia virus.
Project # 4 : Objective : isolating (by PCR) and cloning of Vif gene of SIV in a plasmid (pVL1393) to
construct pVL1393 for vaccine and pathogenesis studies.
The technic have been completely perfonned but no expected pVL1393vif aller three remakes with
different protocols. In conclusion, the test must be done algain using others plasmid and restriction
enzymes. However, 1 possess the technology and 1 cari do it again by myself
Projeet # 5 : We have only start this project with an experilmental infection of an insect (Spodoptera
frugiperda) larvaes which are inoculated, with different recombinant baculovirus, part by injection
with serynge, part by infected food. The dead larvaes proteins were then extracted and stored at -7OOC
(done) for firrther analysis by 1 O%PAGE/western blot (not done).
Because of the absence of the training tutor on vacation, we have not been able to perform this project
which is important for us as it is the basic technic of recombinant vaccine production.
General conclusion : TO master these hasic molecula~r biology technologies, more time is
necessary. SO 1 wish and ask for one more or one and half more month in the next year (2000) to
finaliie the training schedule, important for me and my work
COURSE ATTENDANCE :
1 have completed a course of health and safety instruction entitled : comprehensive Biology and
Medical Waste, on may 18, 1999 (1138 Meyer Hall, UC Davis) (enclosed certificate).
PERSONAL APPRECIATION :
Training quality : excellent ; that gives to Senegal LNERV fa11 profïciency for its molecular biology
laboratory in the areas of disease diagnostic and recombinant vaccine production, also to Dakar
university (Veterinary School) and Nouakchott (Mauritania) university (science school) where 1 am in
charge of molecular biology course since 1995.
MY ACTUAL DUTIES :
1. Head of Microbiology laboratory of ISRA/LNERV : laboratory management, research and
diagnosis of animal diseases agents ; also quality control of veterinary vaccines producted by
1SRAKJPV.
2. Director of the vaccines production unit of ISRA (ISRA/UPV) : unit management : vaccines
production , administration, marketing and comptability
3. Trainer : molecular biology course for Veterinary school students of Dakar univers@ and for
biology science students of Nouakchoot university in Mauritania, since 1995.
OBSERVATIONS/SUGGESTIONS :
1 . Excellent choice for the training place
2. Training schedule and trainers qualities: Perfect
3. Qualities and suitabilities of equipments : Perfect
4. Living conditions : acceptable ; lodging conditions to improve
5 . Luggage excess weight and books allowances must be increased
6. Authorities help : correct.
Dakar, september 09,1999
Dr. M. KONTE
LABORATOIRE NATIONAL DE L’ELEVAGE ET DES
RECHERCHES VETERINAIRES - BP 2057 - DAKAR-HANN
COMPTE RENDU DE MISSION
A I’ELS de San francisco et à l’Université de Californie, Davis, CA, USA
dans le cadre de transfert technologique en recombinaison génétique
pour la mise au point de vaccins recombinants
du 03 avril au 04 août 1999
par M. KONTE
chef de l’Unité de Production de Vaccins
Réf. UPV/Septembre 1999
COMPTE RENDU DE MISSION
*JC***
IDENTITE DU MISSIONNAIRE :
Prénom : Mamady
Nom :
KONTE
OBJET DE LA MISSION :
Formation dans les nouvelles méthodes de Biologie moléculaire, des gènes recombinants en
particulier et transfert de technologies en matière de mise au point de vaccins de type
recombinant. La formation scientifique et technique est précédée d’une participation à un
cours intensif d’anglais pour une remise à niveau linguistique.
PAYS ET LIEUX DE FORMATION
Etats-Unis d’Amérique (USA) : 4 mois : du 03 avril au 4 août 1999 :
Cours intensif d’anglais : ELS Language Centers de San Francisco en Californie : du 05 au 30
avril 1999.
Biologie moléculaire : Laboratoire international de Biologie moléculaire (ILMB), Université
de Californie, Davis : du 03 mai au 03 août 1999
ORGANISATION/PERSONNALITES RENCONTREES
L’organisation des cours intensif d’anglais est parfaite et efficiente.
La formation scientifique et technique a été réalisé à travers l’exécution conjointe de quatre
projets de recherche en attente, préalablement élaborés par les chercheurs du I’ILMB.
L’exécution de chaque projet est dirigé par le tuteur scientifique qui en est l’auteur. La
réalisation de l’ensemble des projets offre l’opportunité de pratiquer toutes les techniques de
biologie moléculaire dans leurs conceptions les plus récentes.
FINANCEMENT
Bourse AIEA, référence C6/SEN/98006R
NATURE DE LA MISSION
Voyage d’étude et de formation spécialisées
POINTS ESSENTIELS
Le financement AIEA est consenti dans l’optique d’un transfert de technologies des pays
développés aux pays en développement d’Afrique, en particulier, en application d’un
principe voulu, exprimé et constamment soutenu par le Directeur de I’ILMB, le Professeur T.
YILMA. Le choix de ce site de formation par 1’AIEA est des plus judicieux.
Les compétences acquises dans le domaine du génie génétique autorise la conception et la
réalisation d’outils biotechnologiques dans le domaine du diagnostic des maladies et
l’identification des agents microbiens ainsi que dans le domaine des vaccins nouveaux de type
recombinant au LNERV/ISRA.
RAPPORT DE MISSION
Deux rapports détaillés, un à mi-parcours et un autre en fïn de formation, ont été produits et
communiqués aux organismes de supervision de la formation (AIEA à Vienne, NRC à
Washington, et ILMB de U C Davis) : copies jointes ci-après.
,
Univers@ of California, Davis
International Atomic Energy Agency
VM-PMI-International Laboratory
P.O. BOX 100, A-1400, Vienna, Austria
Of Molecular Biology
Phone: 431-2600; Fax: 431-26007,
2079 Haring Hall, Davis, CA 95616, USA
Mail:Official.Mail@iaea.org
Phone : 530/752-8306
Fax : 5301752-1354
Institut Senegalais de Recherches
Agricoles (ISRA)-LNERV-
BP 2057-
Dakar (Senegal)- Fax: 221-832 21 18
FIRST TECHNICAL REPORT
Dr. Mamady KONTE
C6/SEN/98006R
Mail : mkonte@ucdavis.edu
FIRST TECHNICAL REPORT
**a****
FELLOW NAME : Dr, Mamady KONTE
FELLOWSHIP NUMBER : C6/SEN/98006R
ADRESS IN USA : Office : c/o Dr. Tilahun YILMA, UCD-ILMB-PMI-VM
2079 Haring Hall, Davis, CA 95616
Home : 717 Adeline Place, Davis, C!A 95616
TRAINING BEGINNING DATE : May 03,1999
TRAINING EXPECTED END DATE : August 03, 1999
TRAINING PLACE : International Laboratory of Molecular Biology
Department of Pathology, Microbiology and Immunology
School of Veterinary Medicine-University of California, Davis
2079 Haring Hall, Davis, CA 956 16 - USA
TRAINING SUPERVISOR : Tilahun YILMA, DVM-PhD, Director and Professor
Training tutors : Shabbir AHMAD, DVM-PhD, Assistant Professor (Research)
Paulo VERARDI, PhD, Post-graduate Researcher
Zhiyun MA, MS
David WADDELL
TRAINING SCHEDULE :
1- PRELIMINARY WORK :
May 03 through May 14,1999 : Administration aid, Housing, Documentation
May 17 through 21,1999 : Polymerase Chain Reaction (PCR) technics and attendance course on
Biosafety (may 18 at 1138 Meyer Hall, UC Davis)
Objective:
Diagnostic applications of PCR
PCR performing : Preparation (extraction) of DNA from cells samples
DNA polymerization by PCR
DNA gel electrophoresis
Tutor : David WADDELL
May 24 through May 28,1999 : PCR technics (cont’d)
Tutor : David WADDELL
II - FIRST AREA OF CONCENTRATION : Sub-cloming a gene
Project 1: Sub-cloning of IFN-gamma in a retrovirus.
Tutor : Zhiyun MA
Objective : Development of live attenuated vaccine by deletion mutation and insertion of human interferon
gamma gene.
June 01 through June 11.1999 :
Preparation of the insert : pLG-Hu-IFN-gamma
Preparation of the vector: pBRp429-3’Nef/NotI
Set up the ligation vector/insert (V-I)
Transformation of E. coli by the V-I
Selection of recombinant colony and extraction plasmid DNA
Recombination check by plasmid and insert isolation
III - SECOND AREA OF CONCENTRATION : Vaceinia Virus as an Expression Veetor
Project 2 : New selection Systems for Vaccinia Virus (Zeocin and Blasticidin)
Objective : TO test the feasibility of using two new antibiotics for selection of vaccinia virus recombinants
in eukaryotic cells. Vaccinia virus recombinants expressing the small zeocin (375 bp) or blasticidin (398
bp) resistance genes under the ~7.5 promoter Will be developed. Future studies Will test their potential use
as a fast and reliable selection method for recombinant vaccinia viruses.
Tutor : Paulo VERARDI
3.1 Project 2 Preliminary work : June 14 through July 02,1999 :
A)- Design primers for PCR cloning of the zeocin and blasticidin resistance genes with engineered Xma 1
(blasticidin) or Pin AI (zeocin) sites. Order primers.
B)- Perform Vent DNA Polymerase PCRs. Clean up PCRs for cloning.
3.2.Project 2 performing Week 1 : Julv 06 throuph iulv 09
C)- Cloning of zeocin and blasticidin resistance genes (PCR products) into the vaccinia virus transfer
vectors pSC 11 (Xma 1 site, under ~7.5). Sequencing of inserts.
3.3. Project 2 performing week 2 : Julv 19 throuph Julv 23,1999 :
D)- Transfection of BS-C-l cells infected with VV WR with ‘pSCl1Zeo’ and ‘pSC11Bsd’ transfer
vectors.
E)- Recombinant vaccinia virus plaque-purification (plaque assay #l) of ‘vWRTK-Zeo’ and ‘vWRTK-
Bsd’ with TK selection and beta-gal screening.
3.4. Project 2 performing week 3 : Julv 26 through Julv 30. 1999 :
F)- Recombinant vaccinia virus plaque-purification (plaque a.ssay#‘t) of ‘vWRTK-Zeo’ and ‘vWRTK-
Bsd’ with TK selection and beta-gai screening.
G)- Recombinant vaccinia virus characterization (histochimical staining, DNA analysis, zeocin and
blasticidin resistance gene expression).
IV - THIRY AREA OF CONCENTRATION : Isolating a gene of interest by PCR
Julv 12 through 161999
Project 3 : Isolating and cloning of vif gene of a retrovirus in a plasmid (pVL1393) to construct
pVL1393vif
Objective : Vaccine and pathogenesis studies
Tutor : Shabbir Ahmad & Zhiyun MA
V - FOURTH AREA OF CONCENTRATION : Bacnlovirus
Poject 4 : Construction, isolation, and production of a reeombinant baculovirus expressing a gene of
interest
Tutor : Shabbir AHMAD
June 14 throueh Julv 30,1999
A)- Insect ce11 culture
B)- Molecular cloning of gene of interest into transfer plasmid (From step IV)
C)- Transfection of insect cells with recombinant plasmid (pVL1393vif) and linear DNA of baculovirus.
D)- Identification and plaque purification of recombinant baculovirus
E)- Molecular analysis of baculovirus expressing the recombinant protein.
F)- Production and partial purification of antigen in insect cells and larvae.
G)- Development of ELISA
Last three steps Will be performed if time allows.
FACILITIES AND MAIN LAI3 EQUIPMENT
Electric Freezer, refrigerator
Centrifuges-microfuge-centrefr.
Spectrophometer
Water bath
Hot shaker
Magnetic stirrer
Powerpac 3000 with probe
Bio-Dot Apparatus
Rack LN storage system
Integrated speed vac
Sequencing gel
Sonic dismembratr
Maxima vac pump
New Brunswick sci. incubator shaker
Sterilization equipement
Biological Safety cabinet wl3 petcocks
1
,
Vortex-genie mixer
Napco vacuum oven
Revco Upright 20.2 CU. Ft. freezer
Napco sterilizer/autoclave w/dryer
Incubator
Thermocycler
Ultra violet transilluminator
Electrophoresis material
Electrophoresis photo material
Pipets
Ice machine
Pro-pipets
Others lab material
Chemical products and enzymes supplies
ELISA computer system Iecturer
Computers
THE FELLOW
THE TRAINING SUPERVISOR
Dr. M. KONTE
Pr. T. YILMA
‘University of California, Davis
International Atomic Energy Agency
VM-PMI-International Laboratory
P.O. BOX 100, A-1400, Vienna, Austria
of Molecular Biology
Phone : 43 l-2600 ; Fax : 43 l-26007
2079 Haring Hall, Davis, CA 95616, USA
Mail : Ot&ial.Mail@iaea.org
Phone : 5301752-8306
Fax : 530/752-1354
Institut Sénégalais de Recherches
Agricoles (ISRA)-LNERV, BP 2057
Dakar-Hann (Sénégal)
Tél.:2218322762;Fax:2218322118
FINAL TECHNICAL REPORT
Dr. Mamady KONTE
C6/SEN/98006R
Mail : konte@eIsmaiI.com
FINAL TECHNICAL REPORT
*****JC
FELLOW NAME :
Dr. Mamady KONTE
FELLOW NUMBER : C6/SEN/98006R
ADRESS IN SENEGAL : ISlWUPV
LNERV, BP 2057, DAKAR-H,ANN
Tel.:2218322762;Fax:22183221
18
Mail : konte@!elsmail.com
TRAINING BEGINNING DATE : May 03,1999
TRAINING END DATE : August 03, 1999
TRAINING PLACE : International Laboratory of Molecular Biology
Department of Pathology, Microbiology and Immunology
School of Veterinary Medicine, University of Califomia, Davis
2079 Haring Hall, Davis, CA 95616, USA
TRAINING SCHEDULE :- 1. Diagnostic applications of PCR
-
2. Sub-cloning of IFN-gamma in a retrovirus
-
3. Vaccinia virus as an expression vector : new selection systems for
Vaccinia virus (Zeocin and Blasticidin)
-
4. Isolating a gene of interest by PCR.
-
5. Construction, isolating and production of a recombinant baculovirus
expressing a gene of interest.
SCHEDULE IMPLEMENTATION :
Project # 1 : 1 had the oppotunity to adapt my knowledge (since 1994) to the new approach of PCR
performing and the new equipment actually in use (new thermocycler in particular).
Project # 2 : The objective was the development of a li.ve attenuated vaccine against HIV-2 by
deletion mutation and insertion of human interferon gamma gene.
The technology has been completely perfonned but we doesn’t get the expected result because both
Xhol and KpnI enzymes haven’t allowed to isolate the insert @LG-Hu-IFN-gamma).
SO, research Will
continue to fmd the appropriated restrictions enzymes.
In conclusion, 1 cari say that 1 possess now this technology and 1 cari do it again completely by
myself. That’s positive. However, the designing technic of approptiated primers and restriction
enzymes must be repetead with trainers.
Project # 3 : Obective : to test the feasibility of using two new antibiotics for selection of vaccinia
virus recombinants in eukaryiotic cells and after, to study their potentiel use as a fast and reliable
selection method for recombinant vaccinia virus.
This technic has been performed, since ptimers designing to first plaque assays. The short time
allowed have not permit to perforrn eompletely the necessary others plaque assays and the next stape
which is the study of potential use as selection method for recombinant vaccinia virus.
Project # 4 : Objective : isolating (by PCR) and cloning of Vif gene of SIV in a plasmid (pVL1393) to
construct pVL1393 for vaccine and pathogenesis studies.
The technic have been completely perfonned but no expected pVL1393vif aller three remakes with
different protocols. In conclusion, the test must be done algain using others plasmid and restriction
enzymes. However, 1 possess the technology and 1 cari do it again by myself
Projeet # 5 : We have only start this project with an experilmental infection of an insect (Spodoptera
frugiperda) larvaes which are inoculated, with different recombinant baculovirus, part by injection
with serynge, part by infected food. The dead larvaes proteins were then extracted and stored at -7OOC
(done) for firrther analysis by 1 O%PAGE/western blot (not done).
Because of the absence of the training tutor on vacation, we have not been able to perform this project
which is important for us as it is the basic technic of recombinant vaccine production.
General conclusion : TO master these hasic molecula~r biology technologies, more time is
necessary. SO 1 wish and ask for one more or one and half more month in the next year (2000) to
finaliie the training schedule, important for me and my work
COURSE ATTENDANCE :
1 have completed a course of health and safety instruction entitled : comprehensive Biology and
Medical Waste, on may 18, 1999 (1138 Meyer Hall, UC Davis) (enclosed certificate).
PERSONAL APPRECIATION :
Training quality : excellent ; that gives to Senegal LNERV fa11 profïciency for its molecular biology
laboratory in the areas of disease diagnostic and recombinant vaccine production, also to Dakar
university (Veterinary School) and Nouakchott (Mauritania) university (science school) where 1 am in
charge of molecular biology course since 1995.
MY ACTUAL DUTIES :
1. Head of Microbiology laboratory of ISRA/LNERV : laboratory management, research and
diagnosis of animal diseases agents ; also quality control of veterinary vaccines producted by
1SRAKJPV.
2. Director of the vaccines production unit of ISRA (ISRA/UPV) : unit management : vaccines
production , administration, marketing and comptability
3. Trainer : molecular biology course for Veterinary school students of Dakar univers@ and for
biology science students of Nouakchoot university in Mauritania, since 1995.
OBSERVATIONS/SUGGESTIONS :
1 . Excellent choice for the training place
2. Training schedule and trainers qualities: Perfect
3. Qualities and suitabilities of equipments : Perfect
4. Living conditions : acceptable ; lodging conditions to improve
5 . Luggage excess weight and books allowances must be increased
6. Authorities help : correct.
Dakar, september 09,1999
Dr. M. KONTE