Genetic Diversity in Crop

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Genetic Diversity in Crop Plants October 2004
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The Issue
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United States Corn Yields by Year
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Genetic Diversity

• Genetic diversity within a crop species is the
  raw material for current plant breeding
• Genetic diversity is the insurance policy to
  enable plant breeders to adapt crops to changing
  environments

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Crop Germplasm and Genetic Resource Management

• Who conserves crop germplasm? A variety of
  governmental, non-governmental, and private
  institutions
• Example Native Seeds/SEARCH, an NGO that
  conserves crop germplasm of Native Americans in
  the southwestern United States and adjacent
  Mexico.

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Crop Germplasm and Genetic Resource Management

• United State Department of Agriculture,
  Agricultural Research Service, maintains the
  National Plant Germplasm System.
• The NPGS conserves in genebanks and orchard
  plantings more than 460,000 different genetic
  types of more than 10,000 species.
• FY04 budget of 40.6 million.
• Genebanks or orchards in more than 20 locations
  throughout the U. S.

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GENEBANK LOCATIONS

• NPGS efforts conducted at 25 U. S. sites
• 450,000 accessions of 10,000 different plant
  species.
• 150,000 samples distributed annually, without
  restriction, worldwide
• 10,000 new introductions into the system per year
  via exploration and exchange
• Conservation/maintenance, characterization,
  evaluation, enhancement, info. management

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Crop Germplasm and Genetic Resource Management

• Acquisition germplasm exchange among genebanks
  plant exploration.
• Distribution NPGS germplasm is distributed free,
  and without restriction, to scientists,
  educators, etc., worldwide.
• 100-150,000 accessions are distributed annually
  40-50 of total available.

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Crop Germplasm and Genetic Resource Management

• Maintenancestorage under slow growth conditions,
  or in field plantings or orchards. When there are
  too few viable propagules, the samples are
  regenerated by controlled pollination or
  vegetative propagation.
• Evaluation and characterizationgenetic
  variability, adaptation, productivity, host-plant
  resistance, nutritional and industrial product
  potential are studied.

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Crop Germplasm and Genetic Resource Management

• Genetic enhancement and breeding--incorporating
  novel genetic diversity into crop genepools, and
  improving their overall agronomic merit.
• Improved germplasm released as lines, varieties,
  or populations for further varietal development.

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Crop Germplasm Databases

• Without preservation of key associated
  information, the value of germplasm diminishes
  substantially.
• GRIN--Germplasm Resources Information System.
  Preserves information about inventory status,
  origin, genetic relationships, and agronomic
  characteristics of germplasm.
• Accessible at http//www.ars-grin.gov

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Gene Pools Harlan and de Wet (1971)

• Primary Consisting of the cultivated species and
  readily crossable relatives with easy gene
  transfer
• Secondary All species that will cross but gene
  transfer is difficult
• Tertiary Can be crossed with difficulty, gene
  transfer requires radical techniques

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Biotechnology

• The use of transgenic approaches allow the
  broadening of the gene pool to all biological
  species.

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DNA

• Hereditary material.
• Contains all information to make proteins.
• Linear polymer of nucleotides.
• Each one has sugar, phosphate and a base.

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How Does DNA Carry Information?

• To answer this question we must take a closer
  look at DNA.
• DNA is a biopolymer
• Polymers are molecules made of repeating units or
  building blocks
• DNA has four chemical building blocks symbolized
  by the letters A,G,C, T
• The letters of your DNA are in a specific order
  that carries information about you!!
• So, a DNA polymer can be represented as a string
  of letters

A G C T T A G G G T A A A C C C A T A T A
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DNA Carries Information in the Sequence of DNA
Letters
. . .A G C T T A G G G T A A A C C C A T A G . . .
A gene

• A gene is a length of DNA letters that contain
• an instruction for a cell to follow.
• The cell uses specially designed protein
  machines
• to read the information in genes.

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The Order of DNA Letters Encodes the Genetic
Information
The order or sequence of the A, G, C and T
letters in the DNA polymer encodes the actual
genetic information

• Example of the DNA letters in a gene
• AGCTTAGGGTAAACCATATAGGGCCATACCCTATCGGTAAGCTT
• AGCTTAGGGAAAACCCATATAGGGCCATACCCTATCGGTAAG

• The specific order of the DNA letters carries
• the information.
• Changing the order of the DNA letters will
  change the information carried by the gene.

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Secret of DNA Fingerprinting Lies in the Ability
to Detect Small Differences in DNA Letters Among
Individual Samples

• Look around the room and see how different we all
  look. Then compare any two human genomes
• The DNA letters are almost the identical order
  (sequence) between any two human genomes!
• A very small number (0.1) of the DNA letters
  differ between any two human genomes.
• Two plants that look very
• similar may be close or
• distantly related because
• humans select for desirable
• traits in new varieties.

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• Molecular Diversity
• - SNP Single nucleotide polymorphism
• InDel Insertion deletion
• - SNPs and Indel are common markers for genetic
  analysis

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Measures of Genetic Diversity

• Two measures of genetic diversity at the DNA
  sequence level are p pi and ? theta.
• AAAAAAAAAAAATTTTTTTTTTAGGGGGGG
• AAAAACAAAAAATTTTTTTTTTTGGGGCGG
• AAAAACAAAAAATTTTTTTTTTTGGGGCGG
• AAAAACAAAAAATTTTTTTTTTTGGGGGGG
• AAAAACAAAAAATTTTTTTTTTTGGGGGGG

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Levels of Genetic Diversity

• AAAAAAAAAAAATTTTTTTTTTAGGGGGGG
• AAAAACAAAAAATTTTTTTTTTTGGGGCGG
• AAAAACAAAAAATTTTTTTTTTTGGGGCGG
• AAAAACAAAAAATTTTTTTTTTTGGGGGGG
• AAAAACAAAAAATTTTTTTTTTTGGGGGGG
• 4
  4 6
• 5 individuals x 30 bases x 10 comparisons 1500
• 14/1500 0.009 p

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Genetic Diversity of Different Species

• Human 0.0008
• Drosophila 0.0070
• Soybean 0.0009
• Corn 0.0096

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How can soybean and corn be so different?

• Corn is an outcrossing species, soybean is a
  predominately a selfing species
• History of selection in the domestication and
  breeding of the species
• Intrinsic differences we dont understand

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Allele

• People have thousands of genes.
• Each gene has one to many alleles.
• An allele is a slightly different DNA sequence of
  a gene.
• Some allelic differences result in different
  phenotypes like brown vs. blue eyes.

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What is selection?

• Nonrandom transmission of alleles from one
  generation to the next.
• Ex. Plants that are very tall with thin stems
  tend to blow over in high winds. The seed from
  these plants usually is missed by the mechanical
  harvester so there is selection for plants
  carrying the alleles for shorter stature.

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The Problem

• To what degree is limiting genetic diversity
• inhibiting genetic improvement in corn?

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Two Views of the Problem

1. Most of the corn germplasm in use in the USA
   today is derived from mixtures of only two major
   races out of 300 races total (Wallace and
   Brown, 1956). The simplest means of correcting
   this situation and of increasing the genetic
   diversity of this important crop is to introduce
   unrelated sources of germplasm from Brown and
   Goodman, 1977, Races of Corn, in Corn and Corn
   Improvement

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Two Views of the Problem
2. From a project comparing sequence diversity
in 21 genes of nine U.S. inbred lines with 16
diversity maize landraces We found that our
sample of U.S. inbreds contained a level of
SNP diversity that was 77 the level of
diversity in our landrace sample. Tenaillon et
al., 2001, PNAS, 989161-9166
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Teosinte
Landraces
Inbreds/Hybrids
Photos courtesy J. Doebley
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Allele Frequencies
Teosintes
Domestication
Landraces
Plant Breeding
Inbreds
Unselected Gene
Domestication Gene
Improvement Gene
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Molecular Diversity in Corn

• How has selection shaped molecular diversity in
  corn?
• Screen 4000 genes for evidence of selection
• Survey SNPs from 1800 genes in diverse maize and
  teosinte germplasm
• Goal Identify genes exhibiting selection
• Domestication, agronomic improvement, and local
  adaptation
• Community resource SNP marker collection

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Genomics

• The science that studies the entire DNA sequence
  on all of the chromosomes in an organism.
• Rather than studying individual genes, new
  technologies allow scientists to study thousands
  of genes at a time.

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Can we develop genomics screens to identify genes
that have undergone selection?

• Invariant SSR approach
• What proportion of the genes that have low
  allelic diversity among inbreds result from
  selection for domestication?
• Contrast sequence diversity among teosintes,
  landraces, and inbreds

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Simple Sequence Repeat Markers
-----------ATATATATAT----------------- --------
---ATATATATATATATAT-----------
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Screening SSR primers against 12 inbred lines
Invariant SSR
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Invariant SSR Screening

• Screened 480 invariant SSR primer sets from ESTs
• against six teosinte and six exotic accessions
• a. 321 monomorphic throughout
• b. 60 polymorphic in both exotics and teosintes
• c. 14 polymorphic in exotics only
• d. 75 polymorphic in teosintes only
• (designated Class II SSRs)
• Analyzed sizes 31 normal SSRs and 44 Class II
  SSRs in 44 teosinte and 45 landrace accessions

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Class II
Non Class II
Class II
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Genetic diversity in ssp. parviglumis vs.
landraces
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AI737167

• Encodes MADS-box (transcription factor gene,
• RNA was found in immature ear library)
• Maps to a gene for ear structure
• between teosinte and maize
• Sequenced 600 bp region from 15 teosinte and
• 16 landrace accessions, in teosintes ? 0.011,
• in landraces ? 0.000
• 4. Attempting to define function,
  ?

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SNP Objectives

• Develop SNPs to help make an integrated
• genetic/physical map of corn
• Develop SNPs in a large set of random maize genes
• to access genetic diversity across Zea mays
• Develop SNPs in candidate genes to support
• trait analysis in maize
• Use contrasts in levels of SNP diversity in
  identify
• genes that have signatures of selection for
  either
• domestication or improvement

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Haplotypes

• AAAAAAAAAAAATTTTTTTTTTAGGGGGGG
• AAAAACAAAAAATTTTTTTTTTTGGGGCGG
• AAAAACAAAAAATTTTTTTTTTTGGGGCGG
• AAAAACAAAAAATTTTTTTTTTTGGGGGGG
• AAAAACAAAAAATTTTTTTTTTTGGGGGGG
• X
  X X
• Three haplotypes genetic diversity of the whole
  sequence is 0.64

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Distribution of SNP haplotypes among 470 maize
unigenes
- Mean haplotype 4.46

• gt 80 of unigenes have 2 to 7
• haplotypes
• Haplotype diversity 0 1
• - Average 0.629
• For each gene, a few haplotypes
• account for much of the diversity

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Are genes with low inbred diversity enriched for
domestication and improvement candidates?
(Masanori Yamasaki)
Chose 36 genes with no diversity among the MPZ
inbred set. Sequenced same region in 16 haploid
landrace samples, 16 teosinte partial inbreds and
a Tripsacum dactyloides sample.   Test for
selection on inbreds, landraces and teosintes
compared to four neutral genes.  
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Summary of Selection Tests on 36 Genes.
Able to get out-group sequence (Tripsacum) for
33. For 5 of the genes at least ¾ tests were
significant in both the inbreds and the landraces
(evidence for domestication genes).   For 7 other
genes at least ¾ tests were significant in the
inbreds but not the landraces (evidence for
improvement genes). One additional gene was
classified as either domestication or improvement
depending on which Tripsacum allele was used.  
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Not significant
Not tested
Improvement
Domestication
Dome.
Impr.
66 of reduction of diversity was lost between
teosintes and exotics
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Diversity in maize inbreds vs. teosinte
0.07
0.06
0.05
0.04
0.03
q mze
0.02
0.01
0
0
0.02
0.04
0.06
0.08
q teo
Average q.mze/q.teo 0.57 Excluding q.mze0
values 0.63
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On a genomic scale.

• Assume 40,000 genes in maize
• 40,000 x 0.05 2000 selected genes
• Before genomics, 11 genes had been identified as
  selected by population genetic approaches
• By sequencing 1000 genes, have 50 novel
  candidates
• These genes need to be divided between
  domestication and improvement

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Corn Genome
Teosintes
Domestication
Landraces
-40
Plant Breeding
-20
Inbreds
Unselected Gene
Domestication Gene
Improvement Gene
38,000 genes
1,000 genes
1,000 genes
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Signatures of Selection

• Hypothesis manipulation of the expression of
  domestication and improvement genes will alter
  key agronomic traits
• Methods use genetic and transgenic approaches to
  examine teosinte, exotic, and inbred alleles

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ITS A FINE DAY FOR SCIENCE