Reporting
Project Information
MACPROGEN
Grant agreement ID: 229458
Project closed
Start date1 April 2009
End date30 June 2012
Funded under
- Specific Programme „Capacities“: Research potential of Convergence Regions
Total cost€ 1 319 600,00
EU contribution€ 893 452,00
893 452,00
426 148,00
Coordinated by
MACEDONIAN ACADEMY OF SCIENCES AND ARTS
North Macedonia
Final Report Summary – MACPROGEN (National reference centre for genomics and proteomics)
Executive summary:
The MACPROGEN project has been a vital resource for the Research Centre for Genetic Engineering and Biotechnology (RCGEB) ‚Georgi D. Efremov‘, Macedonian Academy of Sciences and Arts (MASA) that was aiming to transform it into a National Reference Centre for Genomics and Proteomics. The main objectives of MACPROGEN were establishing a technological platform for high throughput genomics and proteomics research, networking with EU research institutions in order to foster collaborative activities, disseminating knowledge and expertise and ultimately building an interactive and competitive research environment.
Upgrading the infrastructure of RCGEB by obtaining state-of-the-art equipment was one of MACPROGEN’s key objectives and was supported both by MACPROGEN and by the Government of the Republic of Macedonia. The major genomic equipment included: microarray system, genetic analyser, real-time polymerase chain reaction (PCR) and bioanalyser, while the proteomic equipment included: two-dimensional (2D) difference in gel electrophoresis (DIGE) platform, matrix assisted laser desorption ionisation (MALDI) time of flight (TOF)-TOF mass spectrometer, 2D nano high performance liquid chromatography (HPLC) and Accuspot system. The improved infrastructure has created unprecedented opportunities for planning of competitive research projects. Thus, RCGEB researchers are now able to include state-of-the-art technologies in their research programmes.
During the work on MACPROGEN project a total of nine researchers paid by MACPROGEN were working in RCGEB for a total of 223 months. The MACPROGEN researchers have become an integral part of the research potential of RCGEB. They have been involved in the integration of the newly purchased equipment into RCGEB laboratory practices. New assays have been developed and some of the results obtained have been used as preliminary data for the preparation of research project proposals.
Training and networking with leading European Union (EU) institutions represented one of the specific MACPROGEN objectives. During MACPROGEN project we have been closely collaborating with 12 EU institutions. Knowledge transfer has been achieved through training of RCGEB researchers in the new genomic and proteomic technologies introduced to the Centre with the newly purchased equipment. Several expert visits have also been organised with an aim to establish closer professional contacts and facilitate collaboration and to promote MACPROGEN along with the RCGEB. Three workshops were organised at RCGEB during the MACPROGEN project. The theoretical part of the workshops included lectures given by scientists from the EU partner institutions, while the practical part included demonstration of protocols using the major new equipment. MACPROGEN final conference took place in the historic city of Ohrid, Macedonia, from 29 March to 1 April 2012.
promotion and dissemination activities included advertisement of the potential of the national reference centre for genomics and proteomics through website, leaflets, meetings, publications in scientific journals, participation to scientific events etc.
The MACPROGEN activities provided above ensured successful completion of the project. The MACPROGEN-funded introduction of novel genomic and proteomic technologies at RCGEB ‚Georgi D. Efremov‘ has already resulted in strengthening of Macedonian and regional research capacities. Experienced scientists, some of them trained in EU have joined the RCGEB team. Contemporary research projects in collaboration with leading EU institutions have been initiated. It is very much hoped that in the medium term, the modernisation of research along with intensified collaboration will stimulate promising young scientists to remain in Macedonia and establish their research in their home country. It is expected that the improved infrastructure and research environment will have long-lasting effects, such as enhanced levels of science communication and high-impact publications.
Project context and objectives:
The aim of the MACPROGEN project was to upgrade and improve the capacity of the RCGEB ‚Georgi D. Efremov‘, at the MASA (MASA), Skopje, Republic of Macedonia for research and education in the fields of genomics and proteomics.
RCGEB was founded in 1986 as a research unit of the MASA with the main goal of advancing scientific knowledge in the field of protein chemistry, molecular biology, genetic engineering and biotechnology through research, practical training of scientists, postgraduate studies and application. RCGEB was the first institution in Former Yugoslavia that introduced deoxyribonucleic acid (DNA) technologies and was for many years at the forefront of the research, application and education of these disciplines in the region. RCGEB is the major research facility in the field of molecular genetics in the Republic of Macedonia which has reached national, regional and international recognition. RCGEB is one of the few institutions from Macedonia that met the criteria for centre of excellence due to its esteemed scientific and educational record in life sciences.
Since its establishment, the politics of RCGEB has been to continuously upgrade the infrastructure. Therefore, the Centre has been well equipped for research in the field of molecular genetics (ABI 310 genetic analyser, real-time PCR, thermal cyclers, tissue culture laboratory, luminometer, ultracentrifuge etc) and for protein chemistry work (HPLC, amino acid analyser, freeze dryer, beta counter etc). RCGEB has provided access to the infrastructure to many interested institutions and researchers from the country and the region.
RCGEB has been collaborating with numerous institutions from the Republic of Macedonia, neighbouring countries, Europe and worldwid. The collaboration with these institutions encompassed joint research activities, specialisation and training, invited lectures etc. During early 1990s RCGEB has organised a number of very successful advanced theoretical/practical courses in the field of molecular biology and genetic engineering, sponsored by the Federation of European Biochemical Societies (FEBS), the International Centre for Genetic Engineering and Biotechnology (ICGEB)-Trieste, the United Nations Educational, Scientific and Cultural Organisation (UNESCO) etc.
The turbulent political and socioeconomic developments in the Balkan region in the last two decades have had a negative impact on the whole scientific community in Macedonia including the field of biomedical research. The low salaries and lack of carrier path of the scientists have turned research into an unattractive area, resulting in a massive brain drain. The lack of financial resources prevented exchange of academic staff which has also affected the quality of research. In addition, the low throughput equipment limited the scope and capacity of the research and made it ineffective and uncompetitive. The participation of Macedonian institutions/scientists in the Sixth and Seventh Framework Programmes (FP6 and FP7) and particularly in the field of biomedical sciences was more than modest.
Therefore, in spite of the strengths, international reputation and established collaborations of the RCGEB, MASA, in the last years, the Centre could not compete fully at international level. Upgrading and expanding the major facility in the field of molecular genetics in Republic of Macedonia and transforming it into a contemporary National Reference Centre for Genomics and Proteomics could greatly enhance the quality of research and the potential of Macedonian researchers for competitive applications to FP7 and other international funding sources.
The main objectives of MACPROGEN project were:
1. to upgrade the RCGEB and to provide a technological platform for high throughput genomics and proteomics research that will meet the needs of the scientific community in Republic of Macedonia and the region
2. to foster the networking with EU research institutions through exchange of visits, training of local scientists in leading European centres and preparation of collaborative projects of mutual interest
3. ro enhance the participation of Republic of Macedonia in the FP7 and other international funding schemes
4. to promote the new trends in the field of genomics and proteomics and to disseminate the knowledge
5. to create an improved interactive and competitive research environment that will increase the scientific carrier opportunities and will slow down the brain-drain phenomena.
Project results:
The realisation of MACPROGEN project contributed to the strengthening of the Macedonian research and technological development (RTD) capacities and enhancement of Macedonian human research and development capacities. The MACPROGEN activities provided a technology platform and critical human research potential that should facilitate the EU integration of our country in the future.
The work within the MACPROGEN project has been organised in five work packages (WPs) designed to support the main objectives of the project: management and coordination; technological platform and employment; networking and training; workshops and symposia; and promotion and dissemination.
management and coordination
The activities within MACPROGEN project were coordinated by thesteering committee which composed of the coordinator of the project and the WP Leaders. Professor Dr Dijana Plaseska-Karanfilska was a coordinator of the MACPROGEN project and a Leader of WP3 networking and training. WP2 technological platform and employment was headed by the late Academician Georgi D. Efremov, who was also a Director of the RCGEB (until May 2011) and by Prof Dr Dijana Plaseska-Karanfilska (since May 2011). WP4 workshops and Symposia were headed by Dr. Katarina Davalieva and WP5 promotion and dissemination by Dr Emilija Sukarova-Stefanovska. Thesteering committee worked closely with the administration and financial departments of MASA, as well as the RCGEB scientific committee. Thesteering committee met regularly each month to enable a smooth operation of all foreseen activities and to fulfil the objectives of the project.
In particular, Academician Efremov, the director of RCGEB and Dr Dijana Plaseska-Karanfilska, MACPROGEN coordinator have been responsible for overall project management, coordination of the activities according to predefined timetable, ensuring on-time implementation of project activities, organisation of regular monthly meetings, efficient administration of the project, overall financial administration, including establishing and maintaining financial records, monitoring of expenses against budget allocations, preparation of financial summary sheets.
Dr Dijana Plaseska-Karanfilska, was also a contact person of RCGEB for communication with the European Commission and was responsible for the technical management of project activities, drafting of the minutes of the steering committee meetings and preparation of reports for the Commission.
The WP leaders were in charge of monitoring the activities within a respective WP including planning, performance and reporting; coordination of the activities within the WP according to predefined time table previously approved by the management team; close cooperation with the participants involved in the respective WP; ensuring on-time implementation of activities within the WP; regular reporting on the work performed within the WP; preparation of reports concerning the respective WP.
The leader of WP2 was responsible for all activities within this WP, such as employment of researchers, purchasing equipment and implementation of the technologies. The leader of WP3 was responsible for the international communication, training of the researchers in EU partner institutions and expert visits. The leader of WP4 was responsible for the organisation of the genomics and proteomics workshops and the final MACPROGEN conference. The leader of WP5 was responsible for the promotion and dissemination activities and in particular for the development of a website and preparation of leaflets.
All members of the team, including the newly hired researchers, participated in the different activities of the WPs, such as operational use of the equipment and preparation of protocols, project-related networking (training and expert visits), preparation of short summary reports related to their training and/or expert visit; in the promotion and dissemination activities and in the organisation of the workshops and final conference.
Technological platforms
Updating the infrastructure of RCGEB by obtaining state-of-the-art equipment has been one of MACPROGEN’s strategic objectives. This has been a priority for the centre during the first half of the project. The feasibility study was performed at the very beginning of the project with the aim to evaluate the equipment to be purchased in RCGEB. After careful consideration of future research plans the MACPROGENsteering committee highlighted the need to obtain some major proteomic equipment, such as 2D DIGE platform and MALDI-TOF mass spectrometer, and genomic equipment, such as microarray platform, DNA sequencer with higher throughput, real-time PCR system, as well as some additional small equipment to support the major genomic and proteomic platforms, such as lab-on-chip system for DNA, ribonucleic acid (RNA) and protein analysis (bioanalyser), liquid scintillation counter, spectrophotometers, centrifuges, biorobot for automated nucleic acid extraction, PCR thermal cyclers, etc.
Two major sources of funding were envisaged for purchasing of the required equipment. The first one was the FP7 MACPROGEN project and the second one the project entitled ‚Further development of the Research Centre for Genetic Engineering and Biotechnology, MASA‘ constituting part of the initiative of the Macedonian Government to support the modernisation of top research institutions in the Republic of Macedonia. The purchase of the equipment followed national procurement procedures and practices.
The equipment purchased and installed at the very beginning of MACPROGEN project included: a versatile DNA microarray system allowing researchers to design and perform large-scale genomic studies, a 2D DIGE system for proteomic analyses, a genetic analyser for sequencing of genetic material, a real-time PCR system, bioanalyser, spectrophotometers and mirocentrifuges, all complementing the genomic facility, and a liquid scintillation counter used for analysis of radioactively labelled biological material.
The upgrading of the RCGEB proteomic infrastructure during second half of MACPROGEN project was supported by the Government of the Republic of Macedonia by purchasing MALDI-TOF-TOF mass spectrometer including Mascot and Saramis software for protein and microbiological identifications, 2D nano HPLC and Accuspot system.
The 2D DIGE platform represents one of the major platforms of the proteomic facility at the RCGEB. The 2D-DIGE system became an integral part of the majority of research programmes in RCGEB. Two RCGEB researchers were trained in EU laboratories with extensive experience in 2D DIGE technology. This has enabled the performance of several successful 2D and 2D DIGE experiments at RCGEB.
During the work on MACPROGEN project, we have established protocols for whole proteome analysis of two types of samples: breast tissues and seminal plasma. The preliminary results from the analysis of breast cancer tissues served as a basis for preparation of two research project proposals submitted for funding to the Ministry of Science and Education, Republic of Macedonia and to the Science Funds of the MASA. The gained experience in using 2D DIGE platform led to collaboration with institutions that have already expressed interest to run experiments on the platform, such as Faculty of Pharmacy at the University ‚St Cyril and Methodius‘, Skopje, Republic of Macedonia.
The 2D nano-HPLC is valuable equipment as part of the proteomics platform at RCGEB. In the period following the installation, this system was used mainly for peptide mixtures and ensymatically digested proteins. The advantage of this instrument is that it allows much faster separation of complex protein mixtures than conventional gel based techniques. Also, sample requirements for this instrument are far more less than in 2D electrophoresis, which is very useful since in some cases the quantity of material available for analysis is very limited.
Accuspot system allows spotting of the fractions separated by 1D or 2D nano HPLC. The separated fractions are directly spotted on MALDI targets, covered by matrix solution and subsequently analysed on MALDI-TOF-TOF instrument, using mass spectrometry (MS) or MS-MS module of the instrument. Thus far, we have been working to develop several procedures for separation of tryptic digestion of complete proteome of different tissues types, which will hopefully become the basis for development of large-scale research projects in the near future.
The Axima Performance MALDI-TOF-TOF mass spectrometer has complemented the 2D DIGE platform, allowing identification of the differentially expressed proteins detected by the 2D-DIGE experiments. It has been also used for microorganism identification using the SARAMIS database. Following the installation of the mass spectrometer, several researchers-experts in mass spectrometry visited RCGEB laboratories. The hands-on trainings during this visits ensured that RCGEB researchers gain basic experience in several procedures that form an integral part of unknown protein identification. We have gained experience both in peptide mass fingerprinting (PMF) and tandem mass spectrometry of peptides (MS/MS) using Post Source Decay (PSD) and/or collision induced dissociation (CID) method. For the pilot PMF run, we had chosen to do the complete identification of the seminal plasma proteome, previously resolved by 2D electrophoresis.
A number of meetings with different institutions have been organised in order to inform the Macedonian scientific and medical community about the RCGEB potential in the fields of proteomics. Once sufficient experience is gained in the newly acquired proteomics technologies at RCGEB promotional material will be prepared and distributed to the research and health institutions in the Republic of Macedonia. We believe that the commercial usage of the proteomics equipment for protein identification and microbiological identification will ensure sufficient funding for full usage and maintenance of the instruments in the future.
The DNA microarray system analyses were considered as one of the most important field in the genomics for which the RCGEB researchers need to be trained, since there was no prior experience in this technology in RCGEB. Three RCGEB researchers were trained in several EU laboratories with extensive experience in this technology. The trainings abroad were highly important and beneficial for gaining experience and skills to practically perform all steps of these sophisticated experiments at RCGEB laboratories. The trainings in EU laboratories were followed by an on-site training at RCGEB, during which the RCGEB scientists performed the procedures learned in EU laboratories at home laboratory using the new MACPROGEN equipment. The microarray technology was part of all three workshops in RCGEB. A number of array comparative genomic hybridisation (CGH) experiments were performed using the new microarray platform at the RCGEB in the field of mental retardation, male infertility and breast cancer research. The second main application based on microarray technology in RCGEB was related to micro RNA (miRNA) profiling. Most of this work was devoted to the miRNA expression in breast cancer tissues. All these activities ensured that at the end of MACPROGEN, RCGEB researchers possess not only the skills and confidence necessary to run properly the microarray system, but also are able to explore its built-in flexibility.
Both applications array CGH and miRNA profiling will be integrated into future research projects in RCGEB. We are also planning to commercially exploit the array CGH analysis in diagnosis of mental retardation and hope that this will secure funding for consumables and optimal usage of the equipment in the future.
The new 7 500 fast real-time PCR system became a vital tool in a number of applications at RCGEB. During the work on MACPROGEN project, several protocols were developed using this equipment, such as quantification of human miRNAs, globin messenger RNAs (mRNAs) and detection and quantification of some viruses, such as hepatitis C (HCV) and hepatitis B. The real-time PCR has been also used to confirm the results of the first array CGH experiments. It is expected that the usage of the real-time PCR will grow steadily in the months to follow.
The 3130 genetic analyser has been used extensively during the MACPROGEN project. Many protocols were developed that should become part of different research projects and applications in the future. These protocols include both sequencing analysis and fragment analysis, such as Microsatellite, SNaPshot, SNPlex, LOH and multiplex ligation-dependent probe amplification (MLPA). Special emphasis was also given on the use of different additional softwares such as SeqScape and Coffalyser, which enabled better and easier interpretation of the sequencing and MLPA results, respectively. At the end of MACPROGEN project a genetic analyser with eight capillaries has been purchased by RCGEB funds. Also, RCGEB is considering a purchase of a third generation sequencer. For that purpose a feasibility study has been already initiated.
The Agilent 2100 bioanalyser has been used for the analysis of DNA and RNA samples, including miRNA as well. The bioanalyser has become an integral part of several procedures such as the preparation of samples for microarray hybridisation.
During the work on MACPROGEN projects we have developed a number of different protocols for the newly acquired equipment that will become an integrative part of the research and applicative projects and programmes at RCGEB. Altogether, the equipment purchased during the MACPROGEN project has significantly improved the capacity of the RCGEB laboratories and have created unprecedented opportunities for the development of contemporary research programmes resonating with one of the major goals of the centre, namely, modernisation of its research and better level of integration into the European scientific community.
MACPROGEN researchers
The introduction of several new techniques in the laboratory poses special requirements on the researchers working at the centre. MACPROGEN therefore envisaged the appointment of new members of staff who would provide the expertise required for the operation of the new technologies at RCGEB.
During the work on MACPROGEN project a total of nine researchers paid by MACPROGEN were working in RCGEB for a total of 223 months: Lybomira Chakalova, PhD (33 months), Svetlana Madsunkova (33 months), MD, MSc, Predrag Noveski (38.5 months), MSc, Sanja Kiprijanovska (38.5 months), Ivana Maleva (38.5 months), MSc, Biljana Atanasovska (18 months) and Svesdana Moneva (13 monts), MSc, Svetlana Pecioska (5 months) and Ognen Spiroski (5 months).
These researchers have become an integral part of the research potential of RCGEB. The Presidency of MASA in coordination with RCGEB is making efforts to obtain permanent positions paid by the Government of the Republic of Macedonia for the researchers who were working on the MACPROGEN project. Until this is realised, RCGEB has ensured that funds are available for their salaries for the period after completion of the MACPROGEN project.
Before joining the RCGEB Dr Lybomira Chakalova has been working for nine years at the Babraham Institute, Cambridge, United Kingdom (UK) and had been involved in a number of studies on various aspects of the regulation of the mammalian globin gene loci. Her experience has complemented the research on hemoglobinopathies at the RCGEB, a field in which the centre has long-lasting traditions and international reputation. This, in addition to her experience in construction of complex libraries, single-cell techniques and fluorescent microscopy, has resulted in the preparation of two highly competitive project-proposals (European Research Council starting grant, FP7 Ideas work programme and project grant to support collaborations between academics in the UK or Republic of Ireland and scientists from developing or restructuring countries, Welcome Trust, UK). The two project proposals received largely favourable reviews, however, in both cases they were competing at a high level and neither one was funded.
Dr Chakalova’s technical experience in molecular and cell biology, especially in real-time PCR, has been highly beneficial for various research projects at RCGEB. Her laboratory experience and knowledge of various genome databases has also been very important for the performance, data analysis and interpretation of the microarray experiments as well as the design of copy number variation (CNV) real-time and MLPA follow-up analyses.
Svetlana Trivodalieva (Madjunkova) is a medical doctor with several years of experience in molecular diagnostics. During the work on MACPROGEN project, Dr Svetlana Trivodalieva (Madjunkova) has actively participated in strengthening of the competences of RCGEB in the field of genomics.
She was mainly involved in the studies of the genetics of spontaneous abortions and non-invasive prenatal diagnosis. Her involvement in the studies of the genetics of spontaneous abortions has resulted in successful defence of her Master thesis ‚Genetic basis of spontaneous abortions‘ in March 2011 at the Medical Faculty in Skopje, University ‚St. Cyril and Methodius‘ – Skopje, Republic of Macedonia. The research in the field of non-invasive prenatal detection of fetal sex, common monogenic diseases, RHD status and chromosomal aneuploidies is part of her PhD thesis. Part of her research interest involves proteomic analysis of the placenta and aneuploidy biomarker discovery. Dr Madjunkova has also participated in the research on different monogenic diseases and breast and prostate cancers.
The experience of Dr Trivoladieva in a wide spectrum of molecular biology techniques has enabled her successful training in microarray technology and transfer of this technology in RCGEB. She also successfully transferred the knowledge on the methods for non-invasive prenatal determination of fetal sex and RHD status using cell free fetal DNA from maternal plasma, employing the real-time PCR technology, acquired during the training at the University of Plymouth, UK.
During the MACPROGEN project Predrag Noveski has been mostly involved in the project on male infertility. He has been investigating the Y chromosome changes not only in male infertility, but also in different malignancies such as prostate cancer and colon cancer. Predrag was in charge for the high-throughput genomics data analysis, mainly R project package for microarray and MLPA data analysis and the statistical package for the social sciences (SPSS) package for statistical analysis. He has been indispensable in the data analysis of the miRNA microarray experiments on breast cancer samples.
He has also done population studies using markers on the Y chromosome as a part of his MSc thesis ‚DNA markers on Y chromosome in population of Republic of Macedonia‘ which was successfully defended in December 2011. As a continuation of his research activities he has started the work on his doctoral dissertation entitled ‚Y chromosome structural and functional changes in malignant diseases‘.
During the MACPROGEN project, Ivana Maleva has been working mainly on the project on genetics of breast cancer and comparative genomic hybridisation analysis in patients with mental retardation and male infertility. She has been trained in microarray experiments and was instrumental in the set-up of these analyses at RCGEB together with Svetlana Madjunkova. Ivana Maleva also stayed for one-month in Dr Dork’s Laboratory, where she improved her skills and gained novel ideas and expertise in various methods in breast cancer field. While there, she attended training in single nucleotide polymorphism (SNP) genotyping and targeted resequencing on Fludigm platform.
In December 2011 she defended her thesis on the subject: ‚Mutation in genes associated with breast cancer in Macedonian patients‘ and obtained the Master’s degree in molecular biology. She continued the research on breast cancer as part of her doctoral thesis with a title ‚Identification of genetic and epigenetic causes of BRCAness profile of tumour tissue in breast cancer patients‘. Ivana has also contributed to different other research projects in RCGEB.
Sanja Kiprijanovska has actively participated in strengthening of the competences of RCGEB in the field of proteomics. During MACPROGEN project she received several trainings on different proteomics technologies. She was instrumental in the set-up of the proteomics analysis, especially 2D-DIGE experiments and preparation of proteomics working protocols. She was involved in the experiments using MS/MS analysis, as well. She also received a two weeks training in proteomics and genomics analysis of spermatosoa during which she acquired new knowledge in proteomic identification of human sperm proteins and gained practical experience which was very useful for the future research on male infertility in RCGEB.
She has also actively contributed to other research programmes at RCGEB, such as chromosome aneuploidies, HCV infection and polymorphisms associated with human papillomavirus (HPV).
The work of Biljana Atanasovska during MACPROGEN project was focused on detection of abnormal haemoglobin and determination of different types of thalassemia. Working with Dr Chakalova and other team members she was involved in designing and optimising a multiplex single-nucleotide primer extension assay for identification of common Mediterranean beta-thalassaemia mutations and in the optimisation of a multiplex fluorescent PCR assay for identification and typing of HPV. Biljana was also involved in the preparation of the working protocols based on MLPA analysis and real-time PCR, especially those related to haemoglobin work.
Svesdana Moneva joined MACPROGEN in the final year of the project, after she returned to Macedonia from Denmark. She majored in the field of Biochemistry and Cell Biology at the ‚Jacobs University Bremen‘ in Bremen, Germany. She pursued her Master of Science degree at the University of Copenhagen, where she was awarded grant scholarship in the Master of Excellence programme in Human Biology. Her practical and research experience involves different branches of the fields of Cellular and Molecular Biology and Microbiology.
With a previous academic background in molecular and cell biology, her initial project at RCGEB was mainly focused genetic polymorphisms linked to obesity and male infertility.
Networking and training
A specific objective related to RCGEB’s infrastructural expansion and scientific directions was to facilitate scientific training and foster networking with other institutions. The ultimate goal was to establish close collaboration with leading EU institutions for transfer of knowledge and initiation of collaborative projects. During MACPROGEN project we have been closely collaborating with 12 EU institutions: University of Copenhagen, Denmark; Hannover Medical School, Germany; University of Barcelona, Spain; University of Verona, Italy; University of Copenhagen, Denmark, Institute Paoli Chalmette, Marseille, France, Catholic University, Leuven Belgium; Institute Cochin, the Institute National de la Recharche et la Sante Medicale (INSERM), Paris, France; Henri Mondor Hospital, Creteil, France, University of Plymouth, UK; University Medical Centre Ljubljana, Slovenia, Vienna University of Technology, Austria and Clinical Proteomics Centre, Luxembourg.
Since the starting date of the project, researchers on the MACPROGEN team have applied for a total of 18 research projects. The majority were collaborative projects between researchers from RCGEB and other institutions, either from the Republic of Macedonia, foreign institutions or both. Seven of the project applications were successful, five were not selected for funding and the remaining four are awaiting the decisions of the respective funding bodies. Work on funded projects has commenced in accordance with the respective contracts. Details of the projects are available on the MACPROGEN website.
Knowledge transfer has been achieved through training of MACPROGEN researchers in leading EU institutions in the new genomic and proteomic technologies as well as through expert visits and organisation of workshops.
Training of MACPROGEN researchers in the new technologies was considered one of the priorities towards reaching high levels of expertise. Several trainings of researchers from RCGEB in EU laboratories had been organised.
These trainings aimed to ensure that RCGEB researchers are familiarised with the new technologies in the fields of interest of RCGEB, primarily through practical experience. RCGEB researchers have been trained both in genomics (microarray technology gene expression profiling and array comparative genomic hybridisation, high resolution melting, real-time expression and copy number analysis) and proteomics technologies (2D differential gel electrophoresis, MALDI-TOF-TOF mass spectrometry and nano HPLC). Bioinformatics analyses of the data produced by microarray, 2D DIGE and mass spectrometry have been also covered as part of the trainings.
Several expert visits have also been organised. These visits were primarily aiming to establish closer professional contacts and facilitate collaboration, to present and promote MACPROGEN along with the RCGEB and to improve the operational use of the new sophisticated equipment, such as microarray and mass spectrometry.
Several visits were organised to our EU partnering laboratories in order to establish closer collaboration in some particular subject of high importance to RCGEB. Some of the visits were organised in order to initiate new collaboration with institutions with great expertise in the newly acquired technologies at RCGEB, such as 2D DIGE, MALDI-TOF-TOF mass spectrometry and microarray, as well as novel technologies that were considered for future upgrading of RCGEB laboratories, such as next generation sequencing technologies.
Seven experts/scientists visited RCGEB during the MACPROGEN project. One visit to RCGEB was related to the genomics part, i.e. work with the microarray equipment, two included theoretical and practical training in proteomic techniques using newly acquired MALDI-TOF-TOF instrument, while four were primarily focused on establishment of closer collaboration with institutions with common scientific interest.
Workshops and symposia
During MACPROGEN project three workshops were organised at the RCGEB. The theoretical part of all workshops included lectures given by scientists from the EU partner institutions, while the practical part included demonstration of protocols using the major new equipment. The lectures of all three workshops were attended by more than 120 scientists from the Republic of Macedonia, mainly members of the Macedonian Biochemical Society and Macedonian Society of Human Genetics as well as clinicians from several public and private health institutions.
The programmes of the workshops, including the names of the invited speakers, titles of their lectures and practical sessions were published in workshop practicum’s and are available on the MACPROGEN website. MACPROGEN Final Conference took place in the historic city of Ohrid, Macedonia from 29 March to 1 April 2012.
First theoretical/practical genomics and proteomics workshop
The first genomics and proteomics workshop was held from 22 to 26 November 2010. The workshop comprised theoretical lectures and practical sessions. The lectures were presented by invited speakers from the MACPROGEN partner institutions (Prof Niels Tommerup and Jan Gorodkin, University of Copenhagen, Denmark, Dr Thilo Dork, Hannover Medical School, Germany; Dr Cristina Patusso, University of Verona, Italy; Prof Rafael Oliva, Faculty of Medicine, Barcelona, Spain; Dr Max Chaffanet, Institute Paoli-Chalmette, Marseille, France). The practical part of the workshop included demonstration of several protocols using the major new equipment, such as the real-time PCR system, the ABI 3130 genetic analyser, the microarray and 2D DIGE platforms. Each practical session focusing on one technology was led by one or two demonstrators selected from among the participants. In addition, a representative of applied biosystems took on the lead role during select practical sessions involving the use of equipment supplied by applied biosystems. A practical would typically start off with an overview of the technology followed by a demonstration of the techniques and procedures involved. Whenever possible, the participants were given the opportunity to take part in setting up the experiments. The practical part of the workshop was attended by all scientists working at the RCGEB as well as a number of scientists from several Macedonian research institutions.
Second theoretical/practical genomics and proteomics workshop
The second genomics and proteomics workshop also comprised theoretical lectures and practical sessions and was held from 27 to 29 June 2011.
The lectures were presented by three invited speakers from the MACPROGEN partner institutions (Thilo Dork, Hannover, Germany; Neil Avent, Plymouth, UK and Paul Bradey, Leuven, Belgium) and RCGEB scientists. The three lectures given by the scientists from MACPROGEN partner institutions presented various applications of key modern genomics and proteomics technologies in the context of their own research.
A round table session, attended by the guest speakers and RCGEB scientists was also organised with short talks given by a few RCGEB researchers with an aim to present the ongoing research projects performed in RCGEB in the field of genomics and proteomics that represent potential fields of collaboration with the guest speaker’s institutions. The talks were followed by a fruitful discussion including the possibilities for collaboration.
The practical part of the workshop included demonstration of several protocols using the real-time PCR system, the microarray and 2D DIGE platforms. It was attended by all RCGEB scientists and students that were staying in the laboratories at that period. The practical part was a great possibility for exchange of ideas and knowledge between RCGEB scientists and invited guests from EU partnering institutions. The practical in microarray technology included overview of the technology, troubleshooting of the most often encountered technical problems during performance of an experiment, web resources, data analysis and possibilities for custom array design. The practical in 2D DIGE technology included some theoretical and practical aspect of performing an experiment and data analysis using DeCyder and image master platinum software. Real-time PCR technology was addressed in terms of gene expression analysis and CNV validation.
Third theoretical/practical genomics and proteomics workshop
The theoretical/practical third genomics and proteomics workshop was held from 1 to 6 March 2012. The lectures given during this workshop were presented by two invited speakers from the MACPROGEN partner institution (Max Chaffanet and Stephane Audebert, both from Marseille, France). The lectures presented applications of microarray and proteomics technologies in the context of the most up-to-date cancer research.
The practical part of the workshop included demonstration of several protocols using the real-time PCR system, the ABI 3130 genetic analyser, the microarray and MALDI-TOF-TOF mass spectrometer. It was attended by all RCGEB personal, both scientists and students that were staying in the laboratories at that period. Each practical session focusing on one technology was led by one demonstrator selected from the RCGEB scientists and by one of the three guest scientists from the partner institution Paolli Chalmette Cancer Institute, Marseille, France (Max Chaffanet, Jose Adelaide and Stephane Audebert). The practical in microarray technology included overview of the DNA methylation arrays and data analysis. Real-time PCR practical included overview and troubleshooting of high resolution melting (HRM) analysis. Practical using the ABI 3130 genetic analyser included designing and performing MLPA analysis. The practical in proteomics included theoretical overview of MALDI-TOF-TOF technology, sample preparation, analysis in Linear and Reflectron mode of proteins and peptides respectively, MASCOT database searches and bacterial identification using MALDI-TOF-TOF instrument and Saramis database.
Final MACPROGEN conference
MACPROGEN final conference took place in the historic city of Ohrid, Macedonia from 29 March to 1 April 2012. A total of 38 participants were present at the Conference. Of them, 10 were representatives from our EU partner institutions (Prof Massimo Delledonne, University of Verona, Functional Genomics Center, Italy ; Prof Niels Tommerup, Wilhem Johannsen Centre for Functional Genome Research, University of Copenhagen, Denmark; Dr Natalia Bogdanova, Gynaecology Research Unit, Hannover Medical School, Germany; Dr Cristina Patusso, Department of Life and Reproduction Sciences, University of Verona, Italy; Prof Rafael Oliva, Human Genetics Laboratory, Dept. of Physiological Sciences, Faculty of Medicine, Univ. of Barcelona, Spain; Dr Max Chaffanet, Institute Paoli-Chalmette, Molecular Oncology Department, Oncogenomic Group, Marseille, France; Prof Neil Avent, University of Plymouth, Plymouth, UK ; Prof Martina Marchetti-Deschmann, Vienna University of Technology, Institute of Chemical Technologies and Analytics, Austria; Prof Borut Peterlin; University Medical Centre Ljubljana, Institute of Medical Genetics, Slovenia; Prof Bruno Domon, Luxembourg Clinical Proteomics Centre, Luxembourg). The other participants were RCGEB scientists and representatives from the Macedonian scientific community and public and private health institutions with whom RCGEB collaborates on various research projects.
Representatives from our partner institutions and RCGEB scientists participated in the Conference with lectures and short talks that represented their ongoing research. The lectures presented application of the most up-to-date genomics and proteomics technologies in the context of several research areas such as breast and ovarian cancers, male infertility, non-invasive prenatal diagnosis, thalassaemias, deafness. Also, several lectures overviewed the current technological state and possibilities of the latest -omic technologies. Recent developments in mass spectrometry based proteomics approaches, microarray technology, RNA-sequencing (RNA-seq) technology and integratomics were discussed in detail. The conference was also, an excellent opportunity to review and discuss the accomplishments of the collaborative activities initiated under MACPROGEN as well as to update our future plans and goals.
Most of the lectures presented on the MACPROGEN final conference were published as manuscripts in the special Supplement of the Balkan Journal of Medical Genetics, Proceedings of the MACPROGEN final conference which is available on the MACPROGEN website. The list of the participants and the programme of the conference, including the names of the speakers and titles of their lectures are also part of the Balkan Journal of Medical Genetics (BJMG) Supplement and are available on the MACPROGEN website.
Promotion and dissemination
Under promotion and dissemination activities our objectives focused on advertisement of the potential of the National Reference Centre for Genomics and Proteomics in order to strengthen collaboration with Macedonian and foreign institutions. The website at http://www.manu.edu.mk/macprogen was launched during the second month of the realisation of the MACPROGEN project. It was hosted by the web server of the MASA. The website contained all relevant information related to the MACPROGEN project, such as the project general structure and main objectives, participants, EU partners, WPs, equipment purchased, researchers hired, expert visits, trainings, publications, workshops etc. The website was regularly updated to reflect the progress of MACPROGEN project activities.
One thousand copies of leaflets have been published three times during MACPROGEN project. The leaflets included general information regarding MACPROGEN, such as the main objectives and impact of the project, as well as the information on the already completed activities. They are available on the MACPROGEN website. The leaflets were distributed during different scientific meetings by RCGEB staff and also to the participants of different events organised by RCGEB and MASA. The leaflets were also given to the visitors of the RCGEB and were also sent to the collaborating institutions in the country and abroad and to the members of the editorial board of the Balkan Journal of Medical Genetics, a journal that is published by the MASA.
Meetings with clinicians, health professionals and research scientists from several institutions were organised on a regular basis to discuss the centre’s policies for achieving optimal translation of its potential. These meetings represented an opportunity to promote the field of genomics and proteomics among scientific community in the country. In most instances these meetings resulted in initiation of new collaborative programs, activities and projects with different scientific and medical institutions. Thus, these activities were the basis for a wider use of the improved MACPROGEN facility.
RCGEB scientists have participated in several scientific meetings and published 35 conference abstracts in the proceedings of different scientific events, such as European Human Genetics Conference 2009, 2010, 2011 and 2012, eighth and ninth Balkan Meeting of Human Genetics, 12th International Congress of Human Genetics, 35th FEBS Congress, 17th Congress of European Hematology Association and EMBO 2011 Meeting. The list of abstracts is available on the web. These events were an opportunity to promote the MACPROGEN project and to exchange ideas with colleagues from different countries.
MACPROGEN researchers have also published 33 papers in international and national journals, i.e. Balkan Journal of Medical Genetics, Prilosi MASA, Applied Biochemistry and Microbiology, Diversity, Journal of Genetics, Molecular Biology, Forensic Science International, Hemoglobin, Acta Haematologica, Genetic Testing and Molecular Biomarkers, Andrologia, Journal of Andrology.
A list of the publications is available on the web. Ten of these papers were published in a special issue of Balkan Journal of Medical Genetics. These papers represent the spectrum of different research topics performed in RCGEB during the work on MACPROGEN and were also presented as oral lectures during the final MACPROGEN conference.
During MACPROGEN project, RCGEB researchers have given several interviews in different newspapers and radio and television (TV) channels and participated in a television debates. The main aim of these activities has been to communicate science- and medical-related issues to the lay public as well as to popularise the RCGEB and its central role in biomedical research and education.
Potential impact:
The MACPROGEN activities were designed to address all objectives of the project and to bring all expected impacts of the Regpot-2008-1 call, i.e. upgrading the RTD capacity and capability, better integration of the selected research entities in the European Research Area, contribution to regional capacity building and improvement of the potential of research entities to participate in FP7 and other international projects.
Upgrading the RTD capacity and capability
RCGEB has been the most prominent biomedical research institution in Republic of Macedonia with significant influence on the development of the research in the country and wider region. Majority of the activities in the field of molecular genetics has been performed in the RCGEB in collaboration with the Clinics of the Medical Faculty and other health care and scientific institutions.
The upgrading of the RCGEB with high sophisticated genomic equipment, such as microarray platform, genetic analyser, real-time PCR and proteomic equipment, such as 2D-DIGE platform, 2D Nano-HPLC and MALDI-TOF-TOF mass spectrometry technology already have had a significant impact on the research programmes of the RCGEB and the quality of the research in the field of biomolecular sciences in Republic of Macedonia and in the region.
The improved infrastructure has made RCGEB laboratories attractive for many Faculties and Institutions in the country which expressed interest to involve RCGEB staff and infrastructure in their teaching curricula. At present, RCGEB staff is involved in the undergraduate and postgraduate studies of the Faculties of Pharmacy, Natural Sciences and Biotechnology. Dr Dijana Plaseska-Karanfilska has been elected as a Professor at the University ‚St Cyril and Methodius‘. She is participating in the teaching of the Biochemistry and pharmacogenetics courses for undergraduate students at the Faculty of Pharmacy. She also teaches the proteomics and genomics course for undergraduate students of molecular biology at the Institute of Biology, Faculty of Natural Sciences and Mathematics. Prof. Plaseska-Karanfilska is a Course Leader of the ‚Bioinformatic tools for the analysis of nucleic acids and proteins‘ and ‚Molecular diagnostics of inherited, malignant and infectious disorders‘ at the PhD studies at the University ‚St Cyril and Methodius‘. She also participates in several other courses of these PhD studies. Dr Emilija Sukarova and Dr Katarina Davalieva also participate in the practical courses of the PhD studies at the Faculty of Pharmacy, and Dr Katarina Davalieva is also involved in the courses at the Faculty of Biotechnology.
The RCGEB researchers are also one of the institutions-initiators of the new specialisation ‚Clinical laboratory genetics‘ for health workers (medical doctors and pharmacists).
All these activities have attracted many young students to visit and to stay at RCGEB laboratories for a shorter or longer time to work on their graduation, Master or PhD thesis or just for a volunteer work with an aim to gain valuable laboratory experience.
During MACPROGEN project the RCGEB researchers and the improved infrastructure have been focused on well-defined areas of research, which are defined by the current expertise of the participants, such as breast cancer genetics, male infertility, non-invasive prenatal diagnosis and molecular genetics of different monogenic diseases, thus allowing the RCGEB to reach a higher level in the national, regional and European Research Area. Better integration of the selected research entities in the European Research Area Genomics and proteomics are relatively new disciplines for which there are great expectations for their implementation in the everyday clinical practice. They have become a standard approach of a whole genome/proteome analysis in the study of various diseases. The MACPROGEN project has provided contemporary facility, trained and educated researchers, thus enabling competitive research that will ultimately foster the integration in the European Research Area. The improved infrastructure has enabled RCGEB researchers to publish higher number of scientific papers during the work on MACPROGEN project in comparison to the previous several years. The MACPROGEN project has also allowed for increased participation of RCGEB researchers to different Balkan, European and International Conferences. The participation to these meetings was very important to present the results of the pilot experiments performed at RCGEB with the new equipment, but also to keep informed about the newest achievements in different areas of human genetics and to meet and establish contacts with the scientists from the leading European institutions. The RCGEB scientists have given a number of lectures during different events in the country on the Balkan Meetings of Human Genetics, as well as in different EU institutions during the expert visits. During MACPROGEN project three very successful workshops were organised with lecturers given by renowned EU experts in the fields of genomics and proteomics and practical’s presenting the most sophisticated technologies. Thus, MACPROGEN project has helped to restore the reputation of RCGEB as a leading center in the field of molecular research where workshops with cutting edge content are held. Namely, during early 1990’s more than 100 young researchers from 22 different countries from all over the world received advanced practical training in the techniques of genetic engineering and biotechnology at RCGEB during short advanced courses sponsored by UNESCO, FEBS, ICGIB-Trieste, etc. We are convinced that the improved RCGEB infrastructure will enable organisation of theoretical/practical courses in the years following the completion of MACPROGEN project that will be sponsored by different funding agencies and that will contribute to better integration of RCGEB and Macedonian biomolecular researchers into European Research Area.
Contribution to regional capacity building
The reinforcement of research capacities of RCGEB for genomics and proteomics has contributed to the regional capacity building as well. RCGEB has been providing access to the infrastructure for all interested scientists from the country, region and beyond. The reinforced centre for research in genomics and proteomics has further extended the open access politics of RCGEB for researchers from the Republic of Macedonia and the region through fostering joint research activities using the new facility.
The regular meetings with clinicians, health professionals and research scientists during MACPROGEN project represented an opportunity to promote the field of genomics and proteomics among scientific community in the country and to initiate new collaborative programmes that are based on a cutting edge research. In addition several programmes/projects have been initiated in collaboration with the institutions from the neighbouring countries. In particular the project ‚Cobalt compounds – new insights into regulation of iron metabolism‘ with the Bulgarian Academy of Sciences, Sofia, Bulgaria has already been accepted for funding within the Macedonian-Bulgarian cooperation scheme financed by the national Ministries for Science of both countries. The collaboration with the Medical Faculty of Nis and Medical Faculty of Sofia in the field of polycystic kidney disease has already been initiated. This research will be sponsored by the ongoing project financed within the Scopes programme. This is a multilateral project in which the Macedonian principal investigator is Academician Momir Polenakovic, who is the new RCGEB director. A few months stay of young researcher form Nis at RCGEB to investigate the miRNA profiles of polycystic kidney tissues has already been agreed. Several other institutions from neighbouring countries have expressed their interest to use RCGEB infrastructure in the near future, as well. In this way the improved RCGEB facility by the MAPROGEN funds is contributing not only to Macedonian research capacity building, but to regional as well.
Improvement of the potential of research entities to participate in FP7 and other international projects
The upgraded infrastructure of RCGEB as well as the reinforced human resources has provided means for a competitive research in the centre, country and region in the field of molecular biosciences and biomedicine. This area is one of the most promising areas for research with a significant impact on the whole human welfare. The improved RCGEB infrastructure has opened opportunities to Macedonian and regional institutions to better collaborate and compete with European institutions. One of the main objectives of this project was to initiate collaborative projects with EU institutions, thus ensuring a better participation of the reinforced centre in the FP7 programme as well as in other international funding schemes.
Since the starting date of the project, researchers on the MACPROGEN team have applied for a total of 18 research projects, seven of which were successful. The majority were collaborative projects between researchers from RCGEB and other institutions, either from the Republic of Macedonia, foreign institutions or both. The number of submitted and approved projects has increased in comparison to the period before the start of MACPROGEN project. The number of submitted projects would have been even higher if the regularly calls for bilateral projects were opened by the Ministry of Education and Science of Republic of Macedonia as scheduled.
We expect that the number of submitted and funded projects to FP7 and other international calls will increase in the period to come. This is supported by the following facts. The improved infrastructure has enabled participation of RCGEB in an international program-workshop leaded by Plymouth University and with participation of 13 institutions that aims to standardise the procedure for non-invasive determination of fetal rhesus (Rh) status among RhD negative pregnant women. A common programme for identification of genetic and epigenetic causes of BRCAness profile of tumour tissue in breast cancer patients has been agreed with Hannover University. This project is a doctoral thesis of MACPRGEN researcher Ivana Maleva, and will be performed partly at RCGEB in Skopje and partly in Dr Dork’s laboratory at Hannover University. The content of the bilateral project on the proteomics of male infertility has been already agreed with Dr Marcheti-Deschmann that will be applied as soon as the call is opened by the Ministry of Education and Science of Republic of Macedonia and Austrian Federal Ministry of Science and Research (BMWF). The collaboration with Prof Peterlin form the University Clinics of Ljublana has been extended to include the programme on male infertility in addition to the genetics of spontaneous abortions. A collaborative study with participation of RCGEB, University Clinic of Ljubljana and Faculty of Medicine in Belgrade that aims to investigate different polymorphisms in obesity genes associated with male infertility has been already started. Preliminary agreements for closer collaboration on male infertility have been already done with Prof Oliva’s laboratory at the University of Barcelona, as well. RCGEB has also established collaboration with US institution at the Columbia University, New York. The project of schisophrenia research that has been submitted to National Institutes of Health (NIH) has already received favourable reviews and it is expected to start in early 2013.
Use and dissemination of foreground
Spreading of excellence, exploiting results and disseminating knowledge have been an integral part of MACPROGEN project. The knowledge gained within this project has been promoted and disseminated through organisation of workshops, conferences, meetings and other scientific events, training of young researchers in the centre, publications in scientific journals and lay public magazines, web site, promotional material (leaflets), etc.
RCGEB collaborates with scientific institutions and faculties within the Universities as a part of its research activities, as well as with numerous primary, secondary and tertiary health care institutions through the application, i.e. molecular diagnostics. This represented a solid basis for the promotion of the Centre and dissemination of the knowledge within the country. The RCGEB researchers were involved in the Executive Board of the Macedonian Society of Human Genetics (G.D. Efremov, – President; D. Plaseska-Karanfilska, – Secretary General), and Macedonian Society of Biochemistry and Molecular Biology (Emilija Sukarova-Stefanovska – Secretary General) which are important factors in the promotion of biomolecular sciences in the country. The involvement of RCGEB researchers in undergraduate and PhD studies at different faculties have also contributed to the dissemination activities. A number of meetings organised during MACPROEN project has resulted in strengthening of the already existing collaborations and establishment of new ones.
Prof. Plaseska-Karanfilska has also been a Macedonian representative in the European National Genetics Societies for several years, including the whole MACPROGEN period. She has been a member of the scientific committees of the Balkan Meetings of Human Genetics that were held twice during MACPROGEN project (May 2009 and September 2011). She is also a member of the scientific committee of the Balkan Meetings of Human Genetics that will be held in Slovenia in October 2013. These positions have enabled long-term contacts with all relevant professional organisations and individuals in the Balkan and EU and promotion of the National Centre for Genomics and Proteomics, internationally as well.
Networking with leading EU institutions, which has been an integral part of this project has helped in the establishment of firm thematic collaborations with EU partners that should enhance the participation of the Macedonian researchers within FP7 and other international projects in the future. Especially important were contacts with the Hannover University on the field of breast cancer, with University of Barcelona and University Clinic Ljubljana on male infertility, with Plymouth University on non-invasive prenatal diagnosis and on proteomic technologies with Vienna Technology University. MACPROGEN project has also enabled wider participation of RCGEB scientists, especially young one at the International conferences. This has allowed for better communication of the RCGEB potential and MAPROGEN objectives and activities.
The structure and capacity of the National Reference Centre for Genomics and Proteomics has strengthened the role of RCGEB as a Centre of excellence for research, education and applied activities in biomedicine and molecular biosciences in Republic of Macedonia.
List of websites:
MASA
Research Centre for Genetic Engineering and Biotechnology ‚Georgi D. Efremov‘
Contact person: Dr Dijana Plaseska-Karanfilska
Krste Misirkov 2, 1000 Skopje
Republic of Macedonia
Tel: +38-923-235411, Fax: +38-923-115434
Email: gde@manu.edu.mkdijana@manu.edu.mk
Project website: http://www.manu.edu.mk/macprogen
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