Đề tài Better Breeding of Common carp (Cyprinus carpio L) for Small-Scale Fish

Better Breeding of Common carp (Cyprinus carpio L.) for Small-scale Fish Farmers (CARD 002/04VI) Report Capacity Development Christopher M Austrin, Tuan Anh Pham, Binh Thanh Thai June 2007 Acknowledgement The project team would like to gratefully acknowledgement the funding support by AusAID. Many people supported this project and special appreciation is extended to Dr Nguyen Xuan Ly, Head of Technology and Science department, Ministry of Fisheries and Dr Le Thanh Luu, Director, RIA1 for their encouragement, provision of facilities and support in conducting this project. Deakin University and Charles Darwin University are acknowledged for their provision of facilities and in kind support for project activities. The success of the training programs would not be possible without the hard work and dedication of a number of people. Thus we would also like to extend our gratitude to Dr Matthew Baranski (formerly with Deakin University now with Akvaforsk, Mr Mark Schultz (Charles Darwin University), Mr Nguyen Huu Ninh, Mr Nguyen Xuan Chuc, Mr Ngo Si Van, Mr Le Quang Hung, Ms Nguyen Thi Tan, Nguyen Thi Ninh, Mr Tran Vu Hung, Mr Le Anh Thuy, Mr Nguyen Thanh Hai (RIA1) and Mr Nguyen Van Gioi and his staff (Thai Nguyen Aquaculture Center, Thai Nguyen) for their assistance in relation to the organising of the Thai Nguyen workshop. The project team would also like to extend its appreciation to all farmers, hatchery managers, and community representatives for their cooperation and participation in the training activities supported by this project. Lastly we would like to thank Ms Nguyen Thi Khoa and staff of CARD project office in Hanoi for their ongoing assistance and advice. Better breeding of common carp for small scale fish farmers ________________________________________________________________ Christopher M Austin, Tuan Anh Pham, Binh Thanh Thai 2 Contents List of Figures............................................................................................................3 List of Tables .............................................................................................................4 1. Intensive training in molecular genetics for aquaculture in Australia ............5 1.1 Aims of training ..............................................................................................5 1.2 Outcomes .........................................................................................................5 1.3 Benefits of training courses............................................................................6 1.4 Young research training course reports .......................................................7 1.4.1 Training course report of Hung and Tan ...................................................7 1.4.2 Training course report of Mr. Khuong.....................................................10 1.4.3 Training course report of Mr. Bon...........................................................13 2. Workshop for young researchers on fish genetics and selective breeding in Vietnam........................................................................................................16 2.1 Aims of training ............................................................................................17 2.2 Outcomes .......................................................................................................17 2.3 Benefits of training course ...........................................................................20 3. Workshops for farmers and hatchery managers on common carp breeding and broodstock management.....................................................................26 3.1 Aims of training ............................................................................................26 3.2 Outcomes .......................................................................................................27 3.3 Benefits of training........................................................................................28 3.4 Suggestions from fish farmers and hatchery managers ............................28 Appendices...............................................................................................................35 Better breeding of common carp for small scale fish farmers ________________________________________________________________ Christopher M Austin, Tuan Anh Pham, Binh Thanh Thai 3 List of Figures Figure 1.1. Young researchers received training at Charles Darwin University in 2006. From left to right, Mr Dinh Van Khuong and Mr Le Quy Bon. ...........5 Figure 1.2. Young researchers received training at Deakin University in 2005. Mr Le Quang Hung at Deakin University’s Murry Cod hatchery........................5 Figure 2.1. Participants in the workshop for young researchers...............................16 Figure 2.2. Young researchers practicing molecular genetic techniques .................19 Figure 2.3. Teaching fish morphology and methods of computerised data collection ......................................................................................................................20 Figure 2.4. The four manuals were provided to young researchers participating in workshop.......................................................................................................21 Figure 2.5. Young researchers visited Biotechnology Center in Hanoi ...................21 Figure 3.1. Training in common carp breeding and broodstock management for farmers and hatchery managers held in RIA1 in Bac Ninh and National Broodstrock Center in Hai Duong in 2005. ..................................................26 Figure 3.2. Training in common carp breeding and broodstock management for farmers was held in Thai Nguyen Aquaculture Center, in Thai Nguyen city in 2006. .........................................................................................................26 Figure 3.3. Hatchery manager and farmer are shown common carp breeding techniques in National Broodstock Center in Hai Duong city......................28 Figure 3.4. Manual provided for farmer and hatchery manager workshops.............29 Better breeding of common carp for small scale fish farmers ________________________________________________________________ Christopher M Austin, Tuan Anh Pham, Binh Thanh Thai 4 List of Tables Table 2.1. List of paticipants in workshop................................................................22 Table 2.2. Workshop programs.................................................................................23 Table 3.1. List of participants attended training .......................................................30 Table 3.2. Workshop program 2005 .........................................................................31 Table 3.3. Workshop programs 2006........................................................................33 Better breeding of common carp for small scale fish farmers ________________________________________________________________ Christopher M Austin, Tuan Anh Pham, Binh Thanh Thai 5 1. Intensive training in molecular genetics for aquaculture in Australia 1.1 Aims of training To increase capacity of young Vietnamese researchers to contribute to research and development in the field of genetics of brood-stock management, molecular genetics and analysis of genetic data using various software programs. 1.2 Outcomes Two comprehensive molecular genetic training courses were carried out at Deakin University in 2005 and in Charles Darwin University in 2006. Four young Vietnamese researchers received this training: Ms. Nguyen Thi Tan, Mr. Le Quang Hung from Research Institute For Aquaculture No 1 (RIA1), Bac Ninh, and Mr Le Quy Bon from Research Institute For Aquaculture No 3 (RIA3), Nha Trang, Khanh Hoa and Mr Dinh Van Khuong from Nha Trang University, Khanh Hoa. Figure 1.1. Young researchers received training at Charles Darwin University in 2006. From left to right, Mr Dinh Van Khuong and Mr Le Quy Bon. Figure 1.2. Young researchers received training at Deakin University in 2005. Mr Le Quang Hung at Deakin University’s Murry Cod hatchery. Better breeding of common carp for small scale fish farmers ________________________________________________________________ Christopher M Austin, Tuan Anh Pham, Binh Thanh Thai 6 The young researchers have been trained in advanced genetic techniques and statistical procedures. They practiced in the laboratory common genetic techniques used in aquaculture genetic including genotyping microsatellite, sequencing and single strain confirmation polymorphism (SSCP) analysis. In addition, the researchers were trained in the use of number of genetic statistical softwares including CLUSTAL X, DNASP 4.0, PAUP* 4.0b.10, TFPGA, GENPOP, BioEdit (for more detail see 1.4). During the training in Warrnambool, the young researchers also visited the Warrnambool Trout farm and the Murry Cod hatchery production facility, Victoria and in Darwin they visited the Darwin Aquaculture Center to learn the techniques of broodstock and hatchery management in relation to barramundis and mud crabs. 1.3 Benefits of training courses The young researchers gained from their intensive training course in Australia significantly improved knowledge and technical skills in genetics in relation to fish breeding and population genetic analysis. The knowledge that they gained from the course was designed to help benefit their professional career development in aquaculture generally and specifically in molecular genetic in relation to fish breeding and genetic improvement. After training at Deakin University, Ms Nguyen Thi Tan was successful in her application for a Masters scholarship for study in aquaculture genetics in Norway. Mr Le Quang Hung has been carrying out Tilapia population genetic analysis at RIA1 for development of tilapia suitable for saline water culture. Mr Le Quy Bon has been involved in the genetic improvement of fish and shellfish programs at RIA3. Lastly, Mr Dinh Van Khuong has been working at Nha Trang University as a lecturer in genetics. All have used their increased knowledge of fish and aquaculture genetic gain thought their training course. This enhanced knowledge and skill has been directly relevant to Ms Tan, Mr. Bon, Mr Hung and Khuong as they use genetic on a day to day basis. Better breeding of common carp for small scale fish farmers ________________________________________________________________ Christopher M Austin, Tuan Anh Pham, Binh Thanh Thai 7 1.4 Young research training course reports 1.4.1 Training course report of Hung and Tan Ministry of Agriculture & Rural Development Project Name: Better Breeds of Common carp (Cyprinus carpio L.) for Small- scale Fish Farmers (CARD 002/04VI) Report on the short training course in molecular genetics for aquaculture at Deakin University in Australia from 31/10/2005 to 21/12/ 2005 1. VISITOR DETAILS Name: Ms. Nguyen Thi Tan and Mr. Le Quang Hung Position: Researcher Assistant Address: Research Institute for Aquaculture No.1 Dinh Bang, Tu Son, Bac Ninh Vietnam Telephone: 84-4-8780614 Fax : 84-4-8273070 E-mail: [email protected] 2. VISIT DETAILS Date of Arrival : 31 October, 2005 Date of Departure: 21 December, 2005 Duration of stay: 50 days Institutions to be visited: School of Ecology and Environment Deakin University P.O. Box 423 Warrnambool, Victoria, 3280 AUSTRALIA 3. PURPOSE OF THE VISIT To increase capacity of young researchers and technicians to contribute to research and development in the fields of genetics of broodstock management, molecular genetics and analysis of data using software programs. Better breeding of common carp for small scale fish farmers ________________________________________________________________ Christopher M Austin, Tuan Anh Pham, Binh Thanh Thai 8 Time table Period ( from …….to) Works 01/11 - 13/12/2005 Extraction of genomic DNA (1200 tissue common carp samples) 10/11 - 17/12/2005 SSCP and DNA Sequencing analyses 06/12 - 10/12/2005 Data analysis of sequence 12/12 - 17/12/2005 Microsatellites 16/12 - 20/12/2005 Analysis of data from interviews and microsatellites 4. OUTCOMES During our study period in Deakin University, we practiced in Single Strand Conformation Polymorphism (SSCP), Sequencing and DNA fingerprinting (microsatellites) techniques: SSCP and microsatellites techniques including tasks: 1. DNA extraction 2. Running PCR with common carp primers ( approx 225 samples) 3. Preparation of acrylamide gel 4. Preparation, loading and running of PCR samples 5. Silver staining to visualise genotypes 6. Interpretation and scoring bands DNA direct sequencing including tasks: 1. Isolation of DNA 2. Mix with PCR cocktail 3. Amplification 4. Visualise PCR products 5. Purification 6. Sequencing Data analysis DNA sequences were analysed using CLUSTAL X, DNASP 4.0, PAUP* 4.0b.10, Modeltest software programs for alignment, calculating haplotype, nucleotide diversity, genetic distance, constructing phylogenetic trees of common carp populations. Microsatellites (DNA fingerprinting) analyses used the GENPOP software program. Genetic data analysis included: Estimation of allelic frequencies, genotypic frequencies, test for Hardy-Weinberg proportions, measures of genetic variability, levels of differentiation among populations and cluster analysis. Applications of molecular genetics are used for identifying relationships among individuals or groups, helping to understand the demography of populations. These Better breeding of common carp for small scale fish farmers ________________________________________________________________ Christopher M Austin, Tuan Anh Pham, Binh Thanh Thai 9 genetic markers have been used to analyses population structure, genetic diversity and provide a scientific fundamental for a breeding program. Data collected from 133 interviews with stakeholders growing or breeding carp (including state hatcheries, private hatcheries and farmers) from 21 provinces in Vietnam were summarized and analysed using the SPSS statistical package. We gained from our intensive training course in Australia significantly improved knowledge and technical skills in genetics in relation to fish breeding and population analysis. The knowledge that we gained from the course should benefit professional career and development of aquaculture. Acknowledgements The authors are thankful to Dr. Chris Austin and Thai T.B, Deakin University, for advice and assistance in the molecular techniques. We would also like to thanks Dr. Tuan P.A, Research Institute for Aquaculture No.1 for your kindness help and supports. Thanks to AusAID for financial support. SIGNATURE OF VISITORS …………………………Ms. Nguyen Thi Tan…………….. …………………………Mr. Le Quang Hung..…………….. Room 207-208, Building A9, Ministry of Agriculture and Rural Development, No. 2 Ngoc Ha Street, Ba Dinh, Hanoi Địa chỉ mới của Văn phòng Chương trình CARD: Phòng 207-208, Nhà A9, Bộ Nông nghiệp và PTNT, Số 2 Ngọc Hà, Ba Đình, Hà Nội Better breeding of common carp for small scale fish farmers ________________________________________________________________ Christopher M Austin, Tuan Anh Pham, Binh Thanh Thai 10 1.4.2 Training course report of Mr. Khuong Ministry of Agriculture & Rural Development Project Name: Better Breeds of Common carp (Cyprinus carpio L.) for Small- scale Fish Farmers (CARD 002/04VI) Report on the short training course in molecular genetics for aquaculture at Charles Darwin University in Australia from 01/07/2006 to 28/8/2006 1. VISITOR DETAILS Name: Mr. Dinh Van Khuong Position: Lecturer Address: University of Fisheries No 2, Nguyen Dinh Chieu Street Nha Trang, Vietnam Telephone: 84-58-831149 (ext. 171) Fax : 84-4-831147 E-mail: [email protected] 2. VISIT DETAILS Date of Arrival : 02 July, 2006 Date of Departure: 28 August, 2006 Duration of stay: 58 days Institutions to be visited: - School of Science and Primary Industry Charles Darwin University - Arafura Timor Research Facility (ATRF) P.O. Box 41775, Casuarina Darwin, NT, 0811 AUSTRALIA 3. PURPOSE OF THE VISIT Better breeding of common carp for small scale fish farmers ________________________________________________________________ Christopher M Austin, Tuan Anh Pham, Binh Thanh Thai 11 To increase research capacity of young scientists to contribute to research and development in the fields of genetics of broodstock management, molecular genetics and analysis of data using software programs. Time table Period ( from …….to) Works 01/07 - 02/07/2006 Fly from Vietnam to Darwin 03/07 - 03/07/2006 Visiting Charles Darwin University Learning laboratory safety 04/07 - 17/07/2006 Molecular genetic concept Methodologies for DNA-based techniques DNA extraction (about a hundred samples) 18/07-08/08/06 PCR Sequencing 09/08-28/08/06 SSCP techniques Phylogenetic analysis (PAUP) Genetic population analysis (TFPGA, Genpop 3.2) 4. OUTCOMES During our study period in Charles Darwin University, we practiced in Sequencing and Single Strand Conformation Polymorphism (SSCP): DNA direct sequencing including tasks: 7. Isolation of DNA 8. Mix with PCR cocktail 9. Amplification 10.Visualise PCR products 11.Purification 12.Sequencing in ABI 3130 SSCP technique including tasks: 7. DNA extraction 8. Running PCR with common carp primers (approx 100 samples) 9. Preparation of acrylamide gel 10. Preparation, loading and running of PCR samples 11. Silver staining to visualise genotypes 12. Interpretation and scoring bands Data analysis DNA sequences were analysed using CLUSTAL X, DNASP 4.0, PAUP* 4.0b.10, TFPGA. Modeltest software programs for alignment, calculating haplotype, Better breeding of common carp for small scale fish farmers ________________________________________________________________ Christopher M Austin, Tuan Anh Pham, Binh Thanh Thai 12 nucleotide diversity, genetic distance, constructing phylogenetic trees of common carp populations. Applications of molecular genetics are used for identifying relationships among individuals or groups, helping to understand the demography of populations. These genetic markers have been used to analyses population structure, genetic diversity and provide a scientific fundamental for a breeding programs. We gained from our intensive training course in Australia significantly improved knowledge and technical skills in genetics in relation to fish breeding and population analysis. The knowledge that we gained from the course should benefit professional career and development of aquaculture. Acknowledgements I am thankful to Dr. Chris Austin and Thai T.B, Charles Darwin University, for advice and assistance in the molecular techniques. We would also like to thank Dr. Tuan P.A, Research Institute for Aquaculture No.1 for your kindness help and supports. Thanks to AusAID for financial support. SIGNATURE OF VISITORS ………………………Mr. Dinh Van Khuong………….. Room 207-208, Building A9, Ministry of Agriculture and Rural Development, No. 2 Ngoc Ha Street, Ba Dinh, Hanoi Địa chỉ mới của Văn phòng Chương trình CARD: Phòng 207-208, Nhà A9, Bộ Nông nghiệp và PTNT, Số 2 Ngọc Hà, Ba Đình, Hà Nội Better breeding of common carp for small scale fish farmers ________________________________________________________________ Christopher M Austin, Tuan Anh Pham, Binh Thanh Thai 13 1.4.3 Training course report of Mr.