Polyploidy and molecular cytogenetics in crops: ECA conference Dublin July 2013
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European Cytogeneticists Association Conference June 2013 Dublin Polyploidy: ancient and Polyploidy: ancient and modern signatures in modern signatures in karyotypes of crops and karyotypes of crops and breeding material breeding material Pat Heslop-Harrison [email protected] Website: www.molcyt.com or www.molecularcytogenetics.com UserID/PW ‘visitor’ To download full text of papers
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Transcript of Polyploidy and molecular cytogenetics in crops: ECA conference Dublin July 2013
European Cytogeneticists Association Conference June 2013 Dublin
Polyploidy: ancient and Polyploidy: ancient and modern signatures in modern signatures in
karyotypes of crops and karyotypes of crops and breeding materialbreeding material
Website: www.molcyt.com orwww.molecularcytogenetics.com
UserID/PW ‘visitor’To download full text of papers
Recent polyploidy◦Revealed by cytogenetics and hybridization
Recent rearrangements or duplications◦Revealed by molecular cytogenetics
Ancient, evolutionary polyploidy◦Revealed by sequencing
Understanding polyploidy is important for speciation, evolution and breeding
Different sequence classes evolve at different rates and many are saltatory rather than clocks
Consequences and applications
Dasypyrum Dasypyrum breviaristatum breviaristatum 2n=4x=282n=4x=28Is it Is it AAAAAAAA oror AAAABB??
D. villosum (genomic DNA green) × D. breviaristatum (red)Meiotic metaphase I in a F1 hybrid showing autotetraploid nature
Galasso et al.
Wheat evolution and hybridsWheat evolution and hybridsTriticum uratu
2n=2x=14AA
EinkornTriticum monococcum
2n=2x=14AA
Bread wheatTriticum aestivum
2n=6x=42AABBDD
Durum/SpaghettiTriticum turgidum ssp durum
2n=4x=28AABB
Triticum dicoccoides2n=4x=28
AABB
Aegilops speltoidesrelative
2n=2x=14BB Triticum tauschii
(Aegilops squarrosa)2n=2x=14
DD
TriticalexTriticosecale
2n=6x=42AABBRR
RyeSecale cereale
2n=2x=14RR
Satellite DNA probe green
Evolution of Wheats - PolyploidyEvolution of Wheats - Polyploidy
Common Ancestor2n=2x=14
Aegilops ventricosa2n=4x=28
DDNN
Triticum tauschii2n=2x=14
DDAegilops uniaristata2n=2x=14
NN
Triticum aestivum2n=6x=42AABBDD
Triticum durum
2n=4x=28AABB
Triticummonococcum
2n=2x=14AA
Aegilops sp.2n=2x=14
BB
Aegilops2n=2x=14
Triticum2n=2x=14
Lodging in cereals
Use of repetitive Use of repetitive DNA sequences as DNA sequences as chromosome chromosome markersmarkers
Aegilops tauschii Aegilops tauschii in Iranin Iran
Hojjatollah Saeidi and Pat Heslop-Harrison
Dpta1
dpTa1-Repetitive banding pattern does correlate with taxonomic grouping
Copyright restrictions may apply.
Saeidi, H. et al. Ann Bot 2008 101:855-861; doi:10.1093/aob/mcn042
UPGMA dendrograms of the relationships based on IRAP analysis of (A) accessions of Ae. tauschii subsp
Wild diploid bananaWild diploid banana
Cavendish : ‘Western’banana cultivar 2n=3x=33;AAA genomes
RetroelementsRetroelementsSequences which amplify through an RNA Sequences which amplify through an RNA intermediateintermediate
50% of all the DNA!
Alignment of two BACs shows gaps in both A and B genome
MA4_82I11
MBP_81C12
MuhAT1
MuhAT2a
XX TE (SINGLE)XX TE MITE
XX TE (AGNABI)
MuhAT3 MuhAT4 MITE(MBIR)
XX TE XX TE (MBT)
272 bp 102,190 bp
26, 410 bp 128,068 bp
DNA transposons hAT are particularly frequent
hAT (named from hobo (Drosophila), Ac-Ds (maize), and Tam3 (Antirrhinum) transposon are characterized by an 8 bp TSD8 bp TSD, and short TIRs of 5–27 bp short TIRs of 5–27 bp flanking a transposase (sometimes degenerate) including a DDE site.Non-autonomous (sometimes called MITEs, miniature inverted transposable elements) derivatives of hAT elements are also found, with deletion of most of the coding sequence, and may amplify to huge copy numbers.
Menzel, Schmidt, Nouroz, HH in prep 2013
RetroelementsRetroelements
Homologous BAC sequences from Calcutta 4 Homologous over the full length
except for a 5kb inserta Ty1-copia retroelement
12/04/23 19
Sr. No. Primer Pairs Product Size (bp)
Sequence
1. hAT18486hAT19037
560 ACCCACCTGGCTCTTGTGTCAGCGAATGTGTTTTGACCAC
MBP 81C12 (M. balbisiana) x MA4 82I11 (M. acuminata) BACs.
Musa balbisiana (MBP 81C12)M
usa
acu
min
ata
(MA
4 82
I11)
Transposed Element
hAT 1
hAT 2
hAT 4
Microsatellite (AT)
hAT 3621 bp MBT
384 bp TE + 781 MITE
1676 TE
Microsatellite (AT)
4192 bp TE
12/04/23 20
Sr. No. Primer Pairs Product Size (bp)
Sequence
1. hAT18486hAT19037
560 ACCCACCTGGCTCTTGTGTCAGCGAATGTGTTTTGACCAC
MBP 81C12 (M. balbisiana) x MA4 82I11 (M. acuminata) BACs.
Musa balbisiana (MBP 81C12)M
usa
acu
min
ata
(MA
4 82
I11)
Transposed Element
hAT 1
hAT 2
hAT 4
Microsatellite (AT)
hAT 3621 bp MBT
384 bp TE + 781 MITE
1676 TE
Microsatellite (AT)
4192 bp TE
A-genome specific hAT in three Musa hybrids (2n=3x=33)
Musa ‘Williams Cavendish’ (AAA)
Musa (ABB)
Musa (ABB)
12/04/23 22Dot plot showing the complete Inverted repeat.
12/04/23 23
HP-1 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48
1KB800600400200
hAT1 insertion sites in Musa diversity collectionhAT486F and hAT037RTop bands (560-bp) amplified hAT element and lower bands amplifying the flanking sequences only – Menzel, Nouroz, Schmidt, Schwarzacher, Heslop-Harrison 2013/14
Retroelement Markers
Retrotransposon LTRLTR
Retrotransposon LTRLTR
RetrotransposonLTR LTR
Retrotransposon LTRLTR
Insertion
IRAP – InterRetroelement PCR
Retrotransposon LTRLTR
RetrotransposonLTR LTR
IRAP diversity in MusaIRAP diversity in Musa
Teo, Tan, Ho, Faridah, Othman, HH, Kalendar, Schulman 2005 J Plant BiolNair, Teo, Schwarzacher, HH 2006 Euphytica Teo, Schwarzacher et al. in prep.
12/04/23 26Phylogenetic analysis of Musa genomes – separating species. Teo, Schwarzacher et al.
Diploid 2n=2x=22 Musa / banana metaphase probed red with transposable element
Timing of LTR Retrotransposon insertion in MusaRed 0 to 2 Mya, yellow 2 to 4 Mya, green 4 to 6 Mya and blue olderD’Hont et al. 2012
The Banana GenomeThe Banana GenomeSeven countries + international organization
coordinated by Angelique D’Hont - France (CIRAD, Genoscope)
523 Mb DH-Pahang genome size (flow cytometry)
27.5 million Roche/454 single reads 16 x coverage
2.1 million Sanger reads 4 x coverage50.3 x of Illumina data54 BAC sequences (1%)
A D’Hont et al. Nature 000, 1-5 (2012) doi:10.1038/nature11241
Six-way Venn diagram showing the distribution of shared genefamilies (sequence clusters) among M. acuminata, P. dactylifera,Arabidopsis thaliana, Oryza sativa, Sorghum bicolor and Brachypodiumdistachyon genomes.
A D’Hont et al. Nature 000, 1-5 (2012) doi:10.1038/nature11241
Whole-genome duplication events.
A D’Hont et al. Nature 2012doi:10.1038/nature11241
A D’Hont et al. Nature 2012doi:10.1038/nature11241
10 10 mm
DNA methylation is unevenly distributed DNA methylation is unevenly distributed on on MusaMusa chromosomes chromosomes
copia copia elementselements
in methylated in methylated regions, but also regions, but also in some low in some low methylated methylated regions (arrows) regions (arrows)
5MeC
10 10 mm
C.H Teo and Schwarzacher
5MeC
DNA methylation is unevenly distributed DNA methylation is unevenly distributed on on MusaMusa chromosomes chromosomes
gypsy gypsy elementselements
in methylated in methylated regions, but also regions, but also in some low in some low methylated methylated regions (arrows)regions (arrows)
Teo & Teo & Schwarzacher in Schwarzacher in prep 2013 prep 2013
Arachis hypogaea Arachis hypogaea - Peanut- PeanutTetraploid of recent origin, Tetraploid of recent origin, ancestors separated only 3 My agoancestors separated only 3 My ago
Ana Claudia Araujo, David Bertioli, PHH et al. Embrapa, Brasília. Annals Botany in press 2013
Ara
chis
hypogea 2
n=
4x=
40
pro
bed w
ith
(gre
en
) A
. du
ran
en
sis;
(re
d)
A.
ipaën
sis
Bert
ioli
et
al. In p
ress
20
13
Arachis duranensis BAC probes on A. hypogea chromosomes
Right: green BAC AD52G19;red AD79O23Below: red AD51I17Below right: green AD179B13;red 5S rDNA
paradox in the evolution of genome structure: the predominant repetitive DNA genome fraction is in evolutionary flux, whilst, at the same time, low copy number DNA is conserved over evolutionary time
Size and location of chromosome regions from radish (Raphanus sativus) carrying the fertility restorer Rfk1 gene and transfer to spring turnip rape (Brassica rapa)
DAPI metaphase blueRadish genomic red (labels 2 radish chromosomes and 45S rDNA)Rfk1 carrying BAC green labels sites on radish and homoeologous pair in Brassica
Tarja Niemelä, Seppänen, Badakshi, Rokka HHChromosome Research 2012
Recent polyploidy◦Revealed by cytogenetics and hybridization
Recent rearrangements or duplications◦Revealed by molecular cytogenetics
Ancient, evolutionary polyploidy◦Revealed by sequencing
Understanding polyploidy is important for speciation, evolution and breeding
Different sequence classes evolve at different rates and many are saltatory rather than clocks
Consequences and applications
Anything special about crop Anything special about crop genomes?genomes?
Crop Genome size 2n Ploidy Food
Rice 400 Mb 24 2 3x endosperm
Wheat 17,000 Mbp 42 6 3x endosperm
Maize 950 Mbp 10 4 (palaeo-tetraploid) 3x endosperm
Potato 900 Mbp 48 4 Modifed leaf
Sugar beet 758 Mbp 18 2 Modified root
Cassava 770 Mbp 36 2 Tuber
Peanut 2,800 Mbp 40 4 Seed cotyledon
Oil palm 3,400 Mbp 32 Fruit mesocarp
Banana 523 Mbp 2x 33 3 Fruit mesocarp
Heslop-Harrison 2012. www.tinyurl.com/domest
50 years of plant breeding 50 years of plant breeding progressprogressGlobal production figuresGlobal production figures
Agr
onom
y &
N
itrog
en
Genetics
TransgenicBt insect-resistant maize(& herbicide tolerant)
CytoGenomics …CytoGenomics …
The genepool has the diversity to address these challenges …
New methods to exploit and characterize germplasm let use make better and sustainable use of the genepool
Recent polyploidy◦Revealed by cytogenetics and hybridization
Recent rearrangements or duplications◦Revealed by molecular cytogenetics
Ancient, evolutionary polyploidy◦Revealed by sequencing
Understanding polyploidy is important for speciation, evolution and breeding
Different sequence classes evolve at different rates and many are saltatory rather than clocks
Consequences and applications
European Cytogeneticists Association Conference June 2013 Dublin
Polyploidy: ancient and Polyploidy: ancient and modern signatures in modern signatures in
karyotypes of crops and karyotypes of crops and breeding materialbreeding material
Website: www.molcyt.com orwww.molecularcytogenetics.com
UserID/PW ‘visitor’To download full text of papers
