Transcript Overview - University of Missouri

Comprehensive Genetic, Physical,
and Database Resources for Maize
University of Missouri and USDA-ARS
Ed Coe
Mike McMullen
Jack Gardiner
Mary Polacco
Karen Cone
Mannie Liscum
Su-Shing Chen
Georgia Davis
Clemson University
Rod Wing
Ralph Dean
University of Georgia
Andrew Paterson
Objectives
Integrated physical and genetic
map resource
Efficient, high-resolution genetic
mapping resources
Enhanced data curation, access,
and representation
Overview
Genetic map
Anchoring via Molecular Markers
Physical map
What is Genomics?
Genomics science that combines
genetics, high through-put robotics and
bioinformatics to derive large amounts of
high quality biological information that
will allow a greater understanding of the
organism being studied.
Structural Genomics
 Science related to
discovery of gene
order and
organization on a
chromosome.
Genetic mapping
Physical mapping
 In situ hybridization
Functional Genomics
Science related to the discovery and
definition of the function of genes
C16
C18
C20
C22
C24
Aldehydes
C26
Alcohols
Wax Esters
Whole plant
Cellular
Biochemical
RNA
The Maize Genome in
Perspective
Maize
Human
Soybean
Sorghum
Tomato
Rice
Bacteria
2500
2500
1500
900
950
450
5
MB
MB
MB
MB
MB
MB
MB
How maize compares to other
species in size
Human
Soybean
Sorghum
Tomato
Rice
Bacteria
equal
1.7X
2.7X
2.6X
5.5X
500X
Maize Genetic Resources
al1
d10
gl2
zb7
kn1
Maize Genetic Resources
mn1
b1 pl1
vp1
te1
tu1
orp1
a1 su1 y1 pr1 sh1 wx1
Maize Genetic Resources
1685 mutants with images in MDB
>5000 mutants available
>10,000 alleles in MDB
www.agron.missouri.edu
May 8, 2000
Maize Genetic Resources
49 QTL experiments in MDB
59 genetic maps in MDB
7850 mapped loci in MDB
May 8, 2000
Mapping Maize at UMC
 Genetic mapping is based on DNA fingerprint
data of offspring from two parents which differ
in their appearance.
Similar fingerprint data for two gene indicates they are
physically close together on a chromosome.
 Physical mapping is a laboratory based
approach and involves identifying small
overlapping DNA fragment to reconstruct whole
maize chromosomes.
Molecular mapping
Digest DNA
Electrophorese
Hybridize
with probe
Southern
blot
+
Scoring data
A
B
H
H
A
B
A
H
Recombination and
Mapping
 Assume that the frequency of
crossing-over is equal along the
chromosome.
 Two genes that are very close
to one another will have a lower
likelihood of having a cross-over
between them than two genes that
are very far apart.
Recombination and
Mapping
 So, we can determine the
relative distance between genes
by counting the number of
recombinant genotypes for each
pair of genes.
 Lots of recombinants = far apart
 Few recombinants = close together
How Maximum Likelihood
Works
BHBBAHBHHBHHBHB
HHBBABBHHBBBBAB
BHBBABHAHHBHBAB
BHBBABBAHHBHBAB
BHBBHBHAHHBHBAB
umc157
umc76
asg45
zb4
csu3
BHBBAHBHHBHHBHB
BHBBABHAHHBHBAB
HHBBABBHHBBBBAB
BHBBABBAHHBHBAB
BHBBHBHAHHBHBAB
umc157
asg45
umc76
zb4
csu3
ESTs Defined
 EST is an acronym for Expressed Sequenced
Tags.
 cDNAs with map locations.
 ESTs represent the genes active in a
particular tissue at specific stage of
development.
 The cellular function of an EST can often be
derived by comparison to sequence
databases.
EST Mapping: Utility
 ESTs are important markers for physical
mapping.
 EST mapping, coupled with trait mapping,
will help link genes to phenotypes.
 ESTs are the basic building blocks for high
density “gene chips”
Efficient Markers for Maize
Simple Sequence Repeats(SSRs)
Develop SSR markers
Map SSRs genetically
Use SSRs to anchor physical
map to genetic map
Use SSRs to map QTLs more
efficiently
SSRs: What and how?
Dinucleotide and trinucleotide
repeats. ex. CACACACACA
Different corn lines have different
numbers of repeats. (CA)10, (CA)14
These differences can be seen on
a high resolution agarose gel.
Can be used for mapping.
SSR Mapping
C A
T
G
Trait Mapping
Links phenotypes to genetic
locations
Precursor to linking phenotypes
to genes
Provides tools for functional
analysis of gene expression
Aspergillus flavus
Infection via
silk channel
wounds in kernels
cracks in kernels
Aflatoxin
Aflatoxin B1
Potent carcinogen.
Associated with liver cancer
in humans.
Molecular mapping
Gene 1
Lo
Va35
Mp313E
DNA fingerprint
Hi
1
2
3
Int. Hi Lo
4
5
Int. Lo
6
Int.
Gene 2
Lo Hi
Int. Hi
Lo Int. Lo
Int.
Compare the DNA
fingerprint with
grain toxin levels
Look for bands on
fingerprint
associated with
high toxin and
those associated
with low toxin
Mapping Maize at UMC
 Genetic mapping is based on DNA fingerprint
data of offspring from two parents which differ
in their appearance.
Similar fingerprint data for two gene indicates they are
physically close together on a chromosome.
 Physical mapping is a laboratory based
approach and involves identifying small
overlapping DNA fragment to reconstruct whole
maize chromosomes.
Mapping Maize at UMC
Physical Mapping
 Large fragments are cloned into Bacterial
Artificial Chromosomes (BACs).
 DNA fragments can easily be retrieved for gene
hunting experiments.
 A complete physical map is needed before DNA
sequencing of the chromosomes can begin.
Maize HindIII BAC library
Made at CUGI
Average insert
size is 135 kb.
Total coverage is
14X.
Fingerprinting
and anchoring of
5X underway.
Physical Map
Constructed in the laboratory by assembling overlapping DNA
fragments. Each fragment can be easily retrieved for experiments
which can localize genes of interest.
DNA fragments cloned into
Bacterial Artificial
Chromosomes (BACs)
Chromosome 1
Integrating the Genetic and Physical
Maps to Form a Single Map
 Molecular markers serve as anchor points for the
two maps since they can be mapped both
genetically and physically
 Types of Molecular Markers
RFLPs
Simple Sequence Repeats (SSRs)
Expressed Sequence Tags (ESTs)
Markers from other species
Maize Physical Mapping
 Whole-genome
characterization of
HindIII in progress.
 Anchor with AFLPs
and low copy RFLPs
 Considerations:
 Repetitive elements
 Syntenic duplicated
regions
B73 Maize BAC library probed
by asg20 single copy clone
Maize Physical Mapping
CentA-LTR sequence
bnl6.16 - Single copy RFLP
Integrated Genetic and Physical Map
Genetic
Physical
Plant Height
Disease Resistance
Kernel Color
Tassel Branching
Drought Resistance
Unknown Function
(EST)
Chromosome 1
Synteny in Crop Plants
 The order of genes along chromosomes is
conserved in many crop plants.
Maize-Sorghum-Rice
Tomato-Pepper-Potato
 This is known as synteny and occurs in animals
as well.
Mouse-Rat-Humans
 The Missouri Maize Project is working to
characterize maize-sorghum synteny.
Maize Synteny With Close Relatives
The Missouri Maize Project is working to characterize
maize-sorghum-rice synteny. This will allow desirable plant
traits to be transferred across species.
Maize
Sorghum
Rice
Informatics Tasks
What good is the data if you
can’t access it?
Project data curation
Community curation via MaizeDB
Dissemination from MaizeDB
Project Data Curation
Excel Spreadsheets
Scripts
LabDB
Scripts
MaizeDB
What do we use the tools for?
Identifying genes for important
agronomic traits.
Insect resistance
Disease resistance
Drought tolerance
Cold/heat tolerance
Grain quality