Fingerprinting of crop varieties using DNA markers

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Lecture 32

Fingerprinting of crop varieties using DNA markers

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DNA fingerprinting

Breeder Seed

+ Control

Seed Lots

Test sample

DNA extraction

DNA Purity

DNA amplification

DNA electrophoresis

Interpretation &

Documentation

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Markers used for Fingerprinting

• RAPD

• RFLP

• SSR

• ISSR

• AFLP

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Repetitive sequence

primer I

primer II

plant A

plant B

microsatellite

plant A

plant B

flanking region IIflanking region I

specific primers were designed corresponding to

flanking sequence of microsatellite

PCR analysis and analyze on 6%denaturing

polyacrylamide gel with silver staining

A B

Schematic of SSR assay

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Gel electrophoresis

Protocol

•

Gel electrophoresis is a technique used to separate DNA samples based on their size.• DNA is loaded onto an agarose gel which is placed in a buffer solution.

• An electric current will cause the DNA (which is positively charged) to migrate to the negativecurrent (down the gel).

• The small DNA molecules will move faster, and this results in small DNA fragments movingfarther down the gel.

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Gel electrophoresis

Ladder

Breeder SeedSample Seed

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Seed Purity Assessment Of Rice Hybrid Using

Microsatellite Markers

Detection of impurities in the Indian rice hybrid-KRH2Through multiplex PCR using the microsatellite markers RM164 and

RM206. M — 50 bp ladder, A — CMS line (IR58025A), H —  Hybrid

(KRH2), R  — Restorer line (KMR3), 221 to 240 —  Samples of hybrid

KRH2 collected from a commercial seed-lot.

Arrow indicates contaminants

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DNA Fingerprint of Maize genotypes

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Problems & Precautions

• Marker identification

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SuitablePrimer !!!

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Advantages of DNA fingerprinting in plants

- identification of genetic diversity within a breeding population.

- RFLP markers are used to detect the genetic distance in wheat.

-RAPD markers are used for characterization, estimation of genetic relatedness

and determination of genetic diversity of tea germplasm.

- to find genetic relatedness and difference in figs.

- AFLP markers help in assessing genetic diversity among cultivars such as

wheat. It also helps detect higher level of polymorphism.

- DNA fingerprinting of herbal drugs useful in authenticating the various claims

of medical uses related to the plants.

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Simplifying the Search

DNA fingerprinting can be of use to plant breeders to simplify their work and reduce

the amount of time it takes to produce crops with desirable new traits.

Guaranteeing Crops

Protecting plant breeders's rights (the breeders' patents on specific types of seed), is

another use for the DNA test. Disputes over the true identify of seed varieties can be

easily resolved, since the test will be able to isolate the specific traits that distinguishone seed variety from another

Building a Library

In order for the seed industry or others to effectively use DNA tests, both private and

government labs are working on building a library of crop DNA profiles. As newsamples are analyzed, a computer scan can produce matches between the samples

and DNA profiles already recorded. These labs plan to offer the testing service at a

reasonable cost. That means DNA testing will likely becomes as commonplace as

other agricultural testing services, like soil sampling and seed germination testing.