Progress 01/01/09 to 12/31/12
Outputs
Progress Report Objectives (from AD-416): Develop a set of standardized genetic markers for coca DNA that will make it possible to conclusively identify the different coca species in Colombia. Approach (from AD-416): These coca leaf samples will be used for DNA and RNA extractions, as well as DNA and cDNA sequencing. Selected genes with unique regions from the resulting DNA and cDNA sequences will be used for identifying varietal and species differences. The project will look for short sequence repeats (SSRs) and single nucleotide polymorphism (SNPs) markers. Once identified these marker regions will be validated and screened for their ability to show varietal differences within the coca samples. This interagency agreement provided funds to develop molecular markers and conduct the genetic diversity analysis of Erythroxylum species. Through this project new plant varieties were acquired and placed into the Beltsville collection. Short sequence repeats or SSR markers were developed resulting in 15 primers that can separate both E.c.c. and E.n. varieties of Erythroxylum into separate groupings. An additional 30 primers were screened to improve the resolution of the SSR analysis. One hundred and fifty single nucleotide polymorphic or SNP primers were screened revealing 85 functional markers. These primers were validated for their ability to show differences within specific populations of coca plants. After comparison of both marker systems the SNP markers were found to provide much more information and better separation at lower costs. This information will be used by researchers and drug control specialists to understand the genetic diversity that exists within this plant species.
Impacts
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Progress 10/01/10 to 09/30/11
Outputs
Progress Report Objectives (from AD-416) Develop a set of standardized genetic markers for coca DNA that will make it possible to conclusively identify the different coca species in Colombia. Approach (from AD-416) These coca leaf samples will be used for DNA and RNA extractions, as well as DNA and cDNA sequencing. Selected genes with unique regions from the resulting DNA and cDNA sequences will be used for identifying varietal and species differences. The project will look for short sequence repeats (SSRs) and single nucleotide polymorphism (SNPs) markers. Once identified these marker regions will be validated and screened for their ability to show varietal differences within the coca samples. This interagency agreement provides funds to develop molecular markers and conduct the genetic diversity analysis of Erythroxylum species. Through this project new plant varieties were acquired and placed into the Beltsville collection. Short sequence repeats or SSR markers were developed resulting in 15 polymorphic primers that can separate both E.c. c. and E.n. varieties of Erythroxylum into separate groupings. Additional primers are being developed to add more differentiation power to the SSR analysis. One hundred and fifty single nucleotide polymorphic or SNP primers have been identified from more than 60,000 SNP variations identified from Solexa RNA sequencing of 10 libraries of E. c. c. and E. n. varieties. These primers will be screened and validated for their ability to show differences within specific populations of coca plants. This information will be used by researchers and drug control specialists to understand the genetic diversity that exists within this plant species.
Impacts
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Progress 10/01/09 to 09/30/10
Outputs
Progress Report Objectives (from AD-416) Develop a set of standardized genetic markers for coca DNA that will make it possible to conclusively identify the different coca species in Colombia. Approach (from AD-416) These coca leaf samples will be used for DNA and RNA extractions, as well as DNA and cDNA sequencing. Selected genes with unique regions from the resulting DNA and cDNA sequences will be used for identifying varietal and species differences. The project will look for short sequence repeats (SSRs) and single nucleotide polymorphism (SNPs) markers. Once identified these marker regions will be validated and screened for their ability to show varietal differences within the coca samples. This interagency agreement provides funds to develop molecular markers and conduct the genetic diversity analysis of Erythroxylum species. Through this project 14 new plant varieties were acquired and placed into the Beltsville collection. Ten of those varieties survived the move and are growing in the greenhouse. SPCL has dried leave material for all 14 varieties. SSR markers were developed resulting in 18 polymorphic primers and 28 monomorphic primers for E. c. c. and 24 polymorphic and 20 monomorphic for E. n. The polymorphic primers are being tested for their cross reactivity. All confirmed polymorphic primers will be tested on the 14 new varieties from Colombia. Ten varieties of E. coca and E. novo were subjected to RNA sequencing by Solexa to develop SNP markers. Sequencing was completed in June and the bioinformatics selection of these markers has started. Once identified the all molecular markers will be validated and screened for their ability to show differences within specific populations of coca plants. This project was monitored by e-mail and phone calls between the principal investigator and the principal collaborators listed for this project.
Impacts
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Progress 10/01/08 to 09/30/09
Outputs
Progress Report Objectives (from AD-416) Develop a set of standardized genetic markers for coca DNA that will make it possible to conclusively identify the different coca species in Colombia. Approach (from AD-416) These coca leaf samples will be used for DNA and RNA extractions, as well as DNA and cDNA sequencing. Selected genes with unique regions from the resulting DNA and cDNA sequences will be used for identifying varietal and species differences. The project will look for short sequence repeats (SSRs) and single nucleotide polymorphism (SNPs) markers. Once identified these marker regions will be validated and screened for their ability to show varietal differences within the coca samples. Significant Activities that Support Special Target Populations This interagency agreement provides funds to develop molecular markers and conduct the genetic diversity analysis of Erythroxylum species. Once identified, the molecular markers will be validated and screened for their ability to show differences within specific populations of coca plants. This project was started in January of 2009 and the initial development stages have started. Through this project, new plant material will be acquired in July for the Beltsville collection. This project was monitored by e-mail, and phone calls between the principal investigator and the principal collaborators listed for this project.
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