The document discusses genetic purity testing of commercial hybrids. It describes various methods used to test genetic purity, including grow out tests, chemical tests, electrophoresis, and molecular markers. Grow out tests involve growing a seed sample and analyzing morphological characteristics. Chemical tests examine enzymes or fluorescence. Electrophoresis separates biomolecules like DNA and proteins based on size. Molecular markers like RAPD and SSR analyze DNA patterns to identify varieties. Maintaining genetic purity is important for hybrid seed quality and production.

1. commercial
plant breeding
SUBMITTED TO: DR. PRAKASH SINGH SIR

2. Genetic purity
test comm. of
hybrids
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D u s test
0
2
Registration of plant VARIETIES under PPV and
FR act
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3
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.
Genetic 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
Chemical 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. Molecular 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
FRom any other existing variety at least for one
character
Uniformity (u) :
The variety should be sufficiently uniform to enable
its description
Stability (s) :
The variety should be stable in its relevant
characteristic , that is it must remain true to its
initial description even after repeated propogation

13. What 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 30th October 2001.
• The PPV and FR rules were verified on 12th September 2003.
• The PPV and FR authority was established in 11th November 2005.
• Launch of registration of plant varieties was done on 20th February
2007.
• Authority initiate process of registration of varieties of notified crops
FRom 21st 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.

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

19. thanks!