WSO2 Micro Integrator for Enterprise Integration in a Decentralized, Microser...
Role of molecular marker for the improvement of Potato (Solanum tuberosum )
1. Doctoral Seminar- IInd
On
Role of molecular marker for the improvement of
Potato (Solanum tuberosum )
PJaygendra Kumar
Id. No. 3974
Ph.D. Ag. Biotechnology
CDr. R. S. Sengar
Professor
Department of Ag. Biotechnology
2. Introduction
Potato (Solanum tuberosum) is a member of family
Solanaceae and originated in South America
It is third most important food crop in the world after rice
and wheat in terms of human consumption
Potato economical food contains 20.6% carbohydrate,
2.1% protein & 0.3% fat. It is a rich source of starch,
vitamins (C, B1) and some essential amino acid like
leucine, tryptophane, isoleucine Potato is an important
source of food
3. Molecular marker
Molecular markers are specific fragments of DNA that can be
identified within the whole genome
They allow detection of the sequence differences between two
or more individual
Molecular markers are found at specific locations of the
genome
They are used to 'flag' the position of a particular gene or the
inheritance of a particular characteristic
4. Polymorphic
Co-dominant
Reproducible
Robust
Cost effective
Easy to use
A prefect molecular marker should has the
following character as-
5. Classification of Molecular Markers
1. Morphological Markers
2. Biochemical/Protein Markers
3. DNA markers / genetic markers
6. 1. Morphological Markers
Botanical descriptors of plant which are visually or
phenotype characterized
Visible traits, such as leaf shape, flower color, pod color,
seed color, seed shape, fruit shape and stem length
7. 2. Biochemical/Protein Markers
Protein markers may also be categorized into molecular
markers though the latter are more referred to DNA markers
Isozymes are alternative forms or structural variants of an
enzyme that have different molecular weights and
Electrophoresis mobility but same catalytic activity or
function
Isozyme markers can be genetically mapped chromosomes
and then used as genetic markers to map other genes
They are also used in seed purity test in plant breeding
8. 3. DNA markers /Genetic markers
Genetic markers are the biological features that are
determined by allelic forms genes
They can be transmitted from one generation to another,
and used as experimental probes or tags to keep track of an
individual, a tissue, cell, nucleus, chromosome and gene
9. Types of DNA Markers
Non-PCR Based,
RFLP- Restriction fragment length polymorphism
PCR Based
RAPD- Random amplification of polymorphic DNA
AFLP-Amplified fragment length polymorphism
SCAR-Sequence characterize amplified region
CAPS-Cleaved amplified polymorphic sequences
SSR-Simple sequence repeats
EST-Express sequence tags
SNP-Single nucleotide polymorphism
10. Restriction Fragment Length Polymorphism (RFLP)
Organism can be differentiated by analysis of patterns derived from
cleavage of their DNA
Technique is mainly based on the special enzyme called Restriction
endonucleases
In RFLP restriction enzyme digested DNA is resolved by Gel
electrophoresis and nitro cellulose membrane
Different size or length of restriction fragments are produced such
polymorphism are used to difference plant species , genotypes etc..
11. Random Amplified Polymorphic DNA (RAPD)
It is a PCR based technology
This procedure detects nucleotide sequence polymorphism in DNA
It is used to analyze genetic diversity of an individual by random
primers
If the priming sites are in the amplifiable region a discrete DNA
product is formed through cyclic amplification
Amplified products are separated on agarose gel in presence of
ETBR and view under UV.
12. (Amplified Fragment Length Polymorphism) AFLP
AFLP is based on a selectively amplifying a subset of restriction
fragments from a complex mixture of DNA fragments obtained after
digestion of genomic DNA with restriction endonucleases
Polymorphisms are detected from differences in the length of the
amplified fragments by polyacrylamide gel electrophoresis (PAGE)
The technique involves four steps:
(1) Restriction of DNA and ligation of oligonucleotide adapters
(2) Preselective amplification
(3) Selective amplification
(4) Gel analysis of amplified fragments
13. Sequence Characterized Amplified Region (SCAR)
Based on sequence of polymorphic bands from RAPD/RFLP/AFLP
linked to trait of interest Longer primers (15-30 bp) are designed for
specific amplification of particular locus.
Cleaved Amplified Polymorphic Sequence ( CAPS )
Polymorphisms are differences in restriction fragment lengths caused
by SNPs that create restriction endonuclease recognition sites in PCR
produced by locus-specific oligonucleotide primers.
14. Microsatellite polymorphism can be detected by Southern
hybridization or PCR
If nucleotide sequences in the flanking regions of the microsatellite are
known, specific primers can be designed to amplify the microsatellite
by PCR
microsatellite may be identified by screening sequence databases, poly
morphism can detected by gel electrophoresis
Simple Sequence Repeat (SSR) or Microsatellites
15. Single Nucleotide Polymorphisms (SNP)
A SNP is defined as a single base change in a DNA sequence that
occurs in a significant proportion (more than 1 percent) of a large
population. SNPs are co-dominant markers.
Express sequence tags (EST)
ESTs are small pieces of DNA sequence (usually 100 to 800
nucleotides long) generated by sequencing randomly selected cDNA
clones from a library. Bits of DNA sequence , Represent gene
express in single cells, tissues or organ from different organisms.
16. Role of the molecular marker in Potato
Significant correlations exist between the segregation at a certain
genetic marker locus and variation in the trait value, indicating the
presence of a quantitative trait locus (QTL) in the proximity of the
marker locus
Identification of a quantitative trait locus and their large-scale use
will go a long way in developing late blight resistant varieties.
Mapping population at diploid level of Solanum chacoense and
Solanum spegazzinii has been developed for the identification of
QTLs for late blight resistance
17. Protoplast fusion by somatic fusion of leaf mesophyll protoplasts
has provided opportunity to transfer useful genes especially for
disease and insect resistance from wild species and other diverse
sources to cultivated potato.
Genetic transformation through Agrobacterium tuminifaciens in
genetic engineering, viral coat protein mediated cross protection
for control of virus and incorporation of bt gene for insect
control and insertion of genes for herbicide resistance, and high
amino acid contents.
18. Gene Potato species/variety Type of Marker Resistance Reference
Gpa1
Vadg
S. tuberosum SSR Globodera pallida Bryan et al. 2004
Sen1 S. tuberosum PCR based Synchytrium
endobioticum
Gebhardt et al. 2006
Rladg S. tuberosum spp.
andigena
AFLP PLRV resistance Velasquez et al. 2007
Gpa1
Vadg
S. tuberosum CAPS Globodera pallida Molony et al. 2010
Ry-fsto S. tuberosum
(resistance from
S. stoloniferum)
CAPS PVY (extreme) Gebhardt, 2011
Potential markers for trait selection in potato
19. Rych S. chacoense from
resistance
PCR
based
PVY (Extreme) Mori et al. 2011
Gro1 S. tuberoum
(resistance from S.
spegazzinii)
PCR
based
Globodera
rostochiensis
Kuhl, 2011
Gpa2 S. tuberosum
(resistance from S.
tuberosum spp.
andigena)
CAPS Globodera pallida,
pa2
Milzarek et al.
2011
RGp5-
allelic
S. tuberosum SNP Globodera pallida Pardo L. et al.
2013
Rladg S. tuberosum spp.
andigena
SCAR PLRV resistance Mihovi et al.
2014
20. Marker Assisted Selection (MAS)
MAS consists of identifying association between molecular markers
and genes controlling agronomic traits (major genes) , and using
these to improve plant populations
Selection is made on genotype rather than phenotype, which
increases the speed and efficiency of selection
It is used for manipulating both qualitative (disease resistance) and
quantitative (yield) traits
Molecular markers are used to increase the probability of
identifying superior genotypes, by early elimination of inferior
genotypes
22. Quantitative Trait Loci (QTL)
A QTL is the location or region of individual locus or
multiple loci in the genome that affects a trait that is
measured on a quantitative
It can be a single gene or cluster of linked genes that affect
the trait
It is the region of the genome that is associated with an effect
on a quantitative trait
23. Development of mapping population
Genotyping and phenotyping of the mapping population
Construction of genetic maps using molecular markerdata
Detection of QTL Confirmation
and validation of detected QTL
Basic procedure in QTLmapping
24. QTLs have the following characteristics
These traits are controlled by multiple genes, each segregating
according to Mendel's laws
These traits can also be affected by the environment to vary degrees
Many genes control any given trait and Allelic variations are fully
functional
Individual gene effects is small and the genes involved can be
dominant, or co- dominant.
25. Conclusion
Molecular markers technology can benefit breeding
objectives by increasing of the potato variety.
Development to assist selection for disease resistance,
agronomic and quality traits in potato cultivars
increase the starch vitamins (C, B1) and some essential
amino acid like leucine, tryptophane, isoleucine.
Developing late blight resistant varieties.
