Genetic purity & DUS test , PPV&FRA

For the commercial plant breeding
Uploaded by BS Meena
Student of BAU Sabour Bhagalpur

1. commercial plant breeding S UBMITTED TO: DR. PRAKASH SINGH SIR

2. G enetic purity test comm. of hybrids 01 D u s test 02 R egistration of plant VARIETIES under PPV and FR act 03 Table of contents

3. GENETIC PURITY TESTING OF COMMERCIAL HYBRIDS INTRODUCTION GROW OUT TEST CHEMICAL TEST ELECTROPHORESIS METHOD MOLECULAR MARKER 01

4. Introduction Genetic purity refers to trueness to type, or the degree of contamination of seeds caused by undesired genetic varieties or species. The success of hybrid seed production is dependent on the genetic purity of parental lines. Both outcrossing and the inadvertent mixing of seed can compromise seed quality, therefore genetic purity tests are critical tools for seed producers and plant breeders. Need for genetic purity testing To asses genetic purity of hybrid seeds. To increase crop production at national level To increase farmers income and standard of living. To make IPR (plant breeders right and plant variety protection) part strong. For distinctiveness, uniformity and stability test. G enetic purity test of commercial hybrids

5. It is a test in which appropriate sample of seed are grown to determine the genetic purity of a given seed lot of released and notified variety. Morphological characters are studied in details and compared with standard sample. Seeds are sown in main field in a prevailing environment after proper land preparation. Crop is grown up to peak flowering and FRuiting stage and based on distinguishable characters the hybrids are identified. Standard sample – The standard sample of a cultivar is the official standard against which all other samples of the seed of a particular cultivar will be judged. GROW OUT TEST- standard test for genetic purity

6. A technique that separates charged macromolecules ( DNA,RNA,Proteins ) on the basis of relative migration in an appropriate matrix eg. Agarose subjected to an electric field. Agarose gel electrophoresis: Standard method used to separate, identify and purify DNA Fragments. Due to its negative charge, DNA will towards + ve electrode under influence of electric field. FRagments are separated based on molecular weight or size and travels through pores in agarose gel. To make the bands visible ethidium bromide is used. electrophoresis

7. Polyacrylamide gel electrophoresis (PAGE) it is the latest method of cultivar identification based on protein banding and isoenzyme activity. A few seeds are defatted and extracted for protein and isoenzymes and the extracted materials are separated by polyacrylamide gel electrophoresis. Based on the banding pattern the varieties can be differentiated and identified. ELECTROPHORESIS

8. Some chemical tests rely on the presence of specific enzymes (e.g., peroxidase) or fluorescence chemical compounds within the seeds of different varieties. Other tests would require addition of chemical compounds (e.g., potassium hydroxide and hydrochloric acid) into the seeds to highlight seed features. PEROXIDASE TEST Buttery and Buzzell (1968) separated soybean cultivar into two groups based on the presence of either low or high seed coat peroxidase activity. Equipment and chemical: Guaiacols, hydrogen peroxide, test-tube. Procedure: Place seed coats removed FRom the soybean seeds into a test tube or suitable container C hemical method

9. CHEMICAL method Add 10 drops of 0-5% guaiacal solution to the test-tube After 10 minute add a drop of 0.1% hydrogen peroxide solution One minute after adding the hydrogen peroxide, record the seed coat color as peroxidase positive (high peroxidase activity) indicated by a reddish –brown solution or peroxidase negative (low or no peroxidase activity) indicated by colorless solution in the test tube. FLOUROSCENCE TEST Crops: Ryegrass, oat Equipment: Fluorescence lamp, black paper, germinator etc. Procedures: Place the seeds to be tested on a black background Evaluate the seeds for inflorescence under black light tubes in a room FRom which all other sources of light are excluded. Seeds are considered fluorescence of the lemma or pelea fluoresce or appear light in colour , partially fluorescent seeds should be considered fluorescent. Seed are considered non- fluorescent if the lemma and pelea do not fluoresce and appear dark in colour

10. MOLECULAR MARKERS RAPD Randomly amplified polymorphic DNA ssr Simple sequence repeats Restriction FRagment length polymorphism Snp Sequence nucleotide polymorphism rflp

11. M olecular marker Many DNA markers, such as restriction FRagment length polymorphism (RFLP), random amplification of polymorphic DNA (RAPD), simple sequence repeats (SSR), and single nucleotide polymorphism (SNP), can be utilized for the identification of plant varieties. DNA extraction PCR amplification using nucleotide primer 1. Initial Denaturation 2. Repeated cycles a. Denaturation b. Annealing c. Extension 3. Final extension Electrophoretic run and identification of PCR amplified product.

12. D u s test Distinctness (d) : The variety should be clearly distinguishable FR om any other existing variety at least for one character U niformity (u) : T he variety should be sufficiently uniform to enable its description S tability (s) : T he variety should be stable in its relevant characteristic , that is it must remain true to its initial description even after repeated propogation

13. W hat is dus testing DUS testing is a way of determining whether a newly bred variety differs FRom existing varieties within the same species( the distinctness part), whether the characteristics used to establish Distinctness are expressed uniformly ( the uniformity part) and that these characteristics do not change over subsequent generations (the stability part). DUS tests exist so that new varieties can legally gain access to their markets via registration of plant varieties under PPV and FR Act 2001.

14. Registration of plant varieties under PPV and FR act EVENTS: The PPV and FR act was passed on 30 th October 2001. The PPV and FR rules were verified on 12 th September 2003. The PPV and FR authority was established in 11 th November 2005. Launch of registration of plant varieties was done on 20 th February 2007. Authority initiate process of registration of varieties of notified crops FRom 21 st May 2007. National gene bank authority was established in 2007. National register on plant varieties was opened in 2008. First certificate of registration for extant varieties were issued in 2008. Certificate of registration for new varieties & farmers varieties issued for first time in 2009.

15. brainstorming Registration of plant varieties under ppv and fr act The act seeks to grant protection to plant breeders who have developed plant varieties by the use of their intellectual capabilities so as to boost the agricultural development in the country . The various plant varieties that can be protected through registration under the act are: New variety: these are the varieties that are developed new i.e. they do not exist naturally. The distinguishable characteristics are generally easily discernible in cases of new varieties. Essentially derived variety: These are the varieties that have predominantly been derived from an initial variety. They are different from new varieties as they are fundamentally similar to the initial variety to such an extent that the characteristic that distinguishes them is considerable hard to discern.

16. Registration of plant varieties under PPV and FR act Extant variety: The Indian legislation also provides for existing varieties. These are the varieties that are already in existence but still warrant protection for one reason or another. These include: a. Varieties notified under section 5 of the Seeds Act,1966. b. Farmers variety. c. Varieties in public domain. d. Varieties in common knowledge. Farmers variety: These are the varieties that have been traditionally cultivated or evolved by farmers in their fields and their existence is a matter of common knowledge within the community.

17. WWW.GOOGLE.COM WWW.SLIDESHARE.NET WWW.EUROFINSUS.COM WWW.LIFEEASIBLE.COM WWW.MONDAQ.COM PRACTICAL MANUAL OF PRINCIPLES OF SEED TECHNOLOGY. resources

18. thanks!