Maize Genetics, Genomics, Bioinformatics workshop Doreen Ware ARS USDA Cold Spring Harbor...
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Transcript of Maize Genetics, Genomics, Bioinformatics workshop Doreen Ware ARS USDA Cold Spring Harbor...
Maize Genetics, Genomics, Bioinformatics workshop
Doreen Ware ARS USDA
Cold Spring Harbor Laboratory
The Plan
• Acknowledgements• Maize Sequences • Massaging of data sets
– Maize FPC, MMP unigene alignments
• Comparative map views– Pair wise alignments of maize genetic map t to
the rice pseudomolecule, Gramene CMap views
• How might this be useful?
CSHL
Wei Zhao
Kiran Ratnapu
Lincoln Stein
?LennyTeytelman
KenClark
?
Cornell
Noel YapSusan McCouch
Maize sequences
Red genomic, Blue expressed, Green clustered, Black genetic map Pink FPC
ESTs (Expressed Sequence Tag)
• Clusters of ESTs– Tug- Plantgdb (www.plantgdb.org) – GI-TIGR (www.tigr.org)– Unigenes (Maizegdb) private sequences
included deposited in Genbank– NCBI has unigenes
Genomic Sequence
• Complete BACs
• Genomic Survey Sequences (GSS)– BAC ends– 100 skim bacs – Methyl filtered– Hi-Cot filtered
Rice and Maize Synteny Analysis public data sets
• Maize Mapping Project (MMP)– AGI Maize FPC map Cone et al., 2002
• www.maizemap.org, www.genome.arizona.edu/FPC/maize/
• IBM 2 neighbors Map– www.maizegdb.org
• International Rice Sequencing Project (IRGSP) – Rice Genome Sequence
• rgp.dna.affrc.go.jp/IRGSP/
– TIGR rice assembly• www.tigr.org
• Rice sequenced based maps• Cereal genetic and FPC maps • Establish common anchor points between
the genetic and physical maps• Extend information available from the
genetic maps of each species to the physical maps (leverage work of genetic systems)– Quantitative Trait Loci (QTL) and Mutants
Leverage synteny of cereal genomes in the absences of
complete sequence
Maize FPC map
• Identify features to use – Maize unigene overgos (MMP unigenes)– Maize genetic markers
• Identify high confidence features for correspondence to rice.
• Assign a position for the feature on a contig
• FPC maps consist of contigs of clustered BACs
• The BACs are represented by imaged bands
• Bands represent restriction enzyme digest fragments of the BAC clones• cccG/ATATCcccgga…ccggatcaG/ATATCacc
• Features are anchored to BACs in the contigs
Finger Print Contig Maps
What is an overgo?
• ~42 bp oligo sequence used for hybridization
• Derived from the MMP unigenes
• Gene specific?
MMP overgo positions on the maize FPC map
contig
BACsovergos
How many MMP overgo positions are on the maize FPC map?
• All MMP Unigene overgo positions – 15,574
contig
overgos
4 overgo positions on two contigs using 3 overgo probes
Contig A Contig B
How many FPC contigs does an overgo hit?
# of contigs and MMP overgo hybridizes to in the maize FPC map
0
1000
2000
3000
4000
1 3 5 7 9 11 13 15 17 19 21
# of MMP overgos
# o
f B
AC
co
nti
gs
Series1
How many BACs does an overgo hit in a contig?
# of BACs a MMP overgo hybridized to in a FPC contig
0
5000
10000
15000
1 3 5 7 9 11 13 15 17 19
# of BACS hybridized to in a contig
# o
f M
MP
o
verg
os
Series1
Select overgos with at least two BACs hybridized in an FPC contig.
• To remove potential false positives, require an overgo be found on more than one BAC in a Maize FPC contigs.
• Red and pink are accepted blue is rejected– 8864
contig
BACsovergos
Generate a position for the overgo on the FPC contig
• To establish a rough order of the overgo on the contig the position of the BACs the overgo hit in the FPC contig is used. This establishes a relative order of the overgos within the contig– Maximum start position of a BAC
– Minimum end position of a BAC
contig
BACsovergos
Align MMP clusters to the rice genome
• Sequenced based alignments of features to the rice genome using BLAT.
• Single best match in the rice genome
• 6,771 maize unigenes (63%)
Filter based upon match-lengthDistribution of maize unigene cluster hit length
0
200
400
600
800
1000
1200
0-10
0
150-
200
250-
300
350-
400
450-
500
550-
600
650-
700
750-
800
hit length
# o
f fe
atu
re h
its
Series1
Matches with less than 150bp match-length were removed leaving 7,770 hits.
0
100
200
300
400
500
600
700
800
900
1000
43 58 62 65 68 71 74 77 80 83 86 89 92 95 98
Percent Identity
Ma
pp
ed
Co
rns
en
su
s U
nig
en
es
Count
Distribution of percent identity of Maize Cornsensus unigenes mapped to rice
The hits represent 6,692 unique Clusters (62% of the total 10,678).
0
200
400
600
800
1000
1200
1400
1 2 3 4 5 6 7 8 9 10 11 12
Rice Chromosome
Mai
ze C
orn
sen
sus
Un
igen
es
MAPPED TO RICE ON MAIZE FPC UNIGENES PER 10MB
What is the distribution of the maize unigenes across the rice
genome?
Rice
Maize
Calculate adjacent distances between pairs of unigenes on maize contigs and
their distance of the rice genome
50Kb_1MB
Unigene clusters pairs found on maize contigs and the distances within 50Kbp and 1Mbp on
rice chromsome 1 ,4 and 10
050
100150200250300350400450
5000
0
1500
00
2500
00
3500
00
4500
00
5500
00
6500
00
7500
00
8500
00
9500
00
distance of pairs in bps on rice
# o
f m
aiz
e c
on
tig
pa
irs
Colinear gene pairs 400,000 bps or lower
Unigene clusters pairs found on maize contigs found within 200kb on rice chromosome 1, 4,
10
020406080
100120140160180200
1000
0
3000
0
5000
0
7000
0
9000
0
1100
00
1300
00
1500
00
1700
00
1900
00
distance of pairs in bps on rice
# o
f m
aiz
e c
on
tig
pai
rs
90% of colinear maize overgos contig-pairs fall under 400,000 bps are found within
165,000 bps on rice
Rice
Maize
A maize contig span will be defined as syntenic if it contains a unigenes pair that is less than 400 kb apart on the rice genome
Maize Contig 417
CTG 417 Anchored to the maize chromosome 8 by marker UMC1905
Plot the colinear rice and maize spans
that contain genetic marker information • What regions of the rice genome are
syntenic with maize?
• What regions of the rice genome have no synteny with maize?
• What regions of the maize genome have no synteny to rice?
Maize-Rice Colinear intervals with anchored maize genetic position
Genetic loci with sequence mapped to rice genome
Blue - Genetic markers Red -CommonGreen- Colinear FPC contigs
What can rice do for maize?
• Where colinear regions exist to rice– Provide potential genetic neighborhood of unanchored
maize contigs– Provide link to other cereal genetic and physical maps– Provisional order of cereals features where no orientation
is known. (Wheat deletion bin map, Maize filtered reads?)– Candidate sequence for marker screening development
• Where limited or no colinear regions exist to rice– Rethink candidate gene approach?
Provide potential genetic neighborhood of unanchored maize contigs chromosome 8 (red), 3 (blue) and unknown (black)
Comparative maps with other cereals
• Wheat EST deletion map (new)
• Sorghum RFLP map from Paterson (new)
• Rice TIGR assembly (new)
• Maize curated FPC
Rice chromosome 1 to Wheat deletion map,
Sorghum genetic map, and Maize FPC map
Provisional order of cereals features where no orientation is known.
SummaryThe maize physical map can provide a provisional
order for the maize sequences that have been anchored.
In syntenic regions the rice sequence can serve as an anchor to define contact points between cereal genomes
In syntenic regions the rice sequence can provide a provisional order to cereal sequences