Traits

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Traits

• Qualitative traits
• Quantitative traits

• Different phenotypic classes
• Continuous phenotypic distribution

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Types of populations

• F2
• BCn
• RILs
• DHs

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Making a linkage map

• Develop the population
• Genotype individuals
• 3 point linkage analysis

• Based on recombination freq..
• Needs adjustment
• I.e. 80 loci3160 pair wise combinations

Computers will do the job, if you know what you
are doing!
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Types of Markers

• Morphological markers
• Molecular markers
• Proteins
• Isozymes
• Anonymous proteins
• Storage proteins

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Types of Markers (cont.)

• DNA markers
• RADPs
• RFLPs
• AFLPs
• SSRs
• STSs, ESTs
• SSCPs
• SNPs

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RFLPs

• DNA Extraction
• DNA restriction
• Agarose gel electrophoresis
• Southern blotting
• Probe hybridization
• Washing film exp.
• Film development
• Total time 7-10 days

• RsN/4b
• rice (4.5x109 bp)17x106 rs
• Probe can be known DNA or anonymous
• Uses 32P
• Requires large amount of DNA (15 µg)

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Parental Screening Blot
P1
P2
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a
b
1
1 2 3
-
c
2
b
c
a
3

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The PCR
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SSRs

• DNA extraction
• DNA amplification (PCR)
• Polyacrilamide gel electrophoresis
• (Staining) (not in fluorescence systems)

• Simple sequence repeats
• 1-5 bp
• I.e. (CA)n (CGT)n
• Requires primers flanking the repeats

Total time 1-2 days
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SSRs
P1
P2
Segregating population
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AFLPs

• DNA extraction
• DNA restriction (2 enzymes)
• Ligation of adapters
• Amplification
• Selective amplification (primers 3 select. Ns)
• Polyacrilamide gel electrophoresis

• Dominant
• Low reproducibility

Total time 5-7 d.
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AFLPs
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Comparison
labor extraction cost not included
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Sources of Markers

• SSRs Publications
• STS/ESTs Genomic databases (http//www.ncbi.nlm.n
  ih.gov)
• RFLPs Publications clone repositories
  authors (http//wheat.pw.usda.gov)
• AFLPs Publications, yourself

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Association between markers and traits

• Bulk segregant analysis techniques
• Through linkage maps
• QTL analysis

• Good phenotypic characterization

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Bulk segregant analysis

• Develop population
• Evaluate trait/s
• Phenotypic classes
• Genotyping individuals
• Option Bulk DNA within classes

• No need for linkage map
• Very appropriate for qualitative traits
• For quantitative traits use extreme individuals
• ANOVA, t-test

From Kaeppler et al, TAG(1993)85
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QTL analysis

• Develop population
• Evaluate trait
• Several environments
• Develop linkage map
• Statistical analysis

• Requires good map and good phenotype
• Statistical analysis
• Single makers
• Interval mapping

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QTL estimation

• Size and type of population
• Number and location of true loci
• Trait heritability
• Genome coverage
• QTL analysis

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Markers in Plant Breeding

• Genetic knowledge
• Number and effects of genes involved
• Aid in selection
• Morphological markers, storage proteins already
  being used

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Using molecular markers in a plant breeding
program

• Limitations
• Cost
• Integration in the program (Timing)
• Markers are source depending

• Potentials
• Money/time savers
• Increasing selection pressure
• Earlier selection
• Introgression of new alleles in the elite
  germplasm (backcross breeding)

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Two forms of Marker Assisted Selection

• Background selection
• Using markers through the genome in order to
  recover as much of the recurrent parent as
  possible
• Foreground selection
• Using 1-2 markers per gene (or QTL) in order to
  select those individuals homozygous for the
  desired allele

A third form background and foreground selection
combined
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Foreground selection for qualitative traits

• Recessive alleles
• Traits expressed after flowering
• Costly phenotypic screening

In backcross
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Foreground selection for qualitative traits
(cont.)

• Two issues in marker assisted backcross breeding
• Number of lines needed
• Distance from the marker/s to the locus

We need to be certain to be able to find at least
one line with the desired genotype for the marker
locus
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Foreground selection for quantitative traits

• Most important traits are quantitative
• Introgressing a QTL in elite germplasm (similar
  to foreground selection for qualititative traits)
• Application to recurrent selection?

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Relative efficiency (RE) of MAS for QTLS

• RE increases with lower heritability
• RE increases with higher proportion of additive
  variance associated with markers
• RE increases with fewer QTLs
• RE increases when marker and QTL are in coupling
  phase vs. repulsion phase

Simulation studies
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How would you integrate MAS in your breeding
program?
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Some reading anyone?

• Mol. Breeding (1999) 5143
• TAG (1998) 96147
• TAG (1992) 84803
• Heredity (1998) 80489
• Euphytica (1981) 30227
• Statistical Genomics. B.H. Liu

• http//nbpgr.delhi.nic.in/mmarker
• Molecular dissection of complex traits. Paterson
  A.H. (ed)
• Crop Sci. (1997) 371686
• Genetics (1997) 1471469
• Crop Sci. (1998) 381164

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More reading!

• Crop Sci. (1999)39967
• New Phytologist (1997) 137 (1)165
• TAG (1993) 85. Kaepler et al