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Comparative Genomics of Fagaceae

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Comparative Genomics of Fagaceae Comparative Genomics of Fagaceae Images.google.com TM www.quia.comLinkage Map www.clipartlord.com Comparative Genomics of Hardwood Tree Species http://www.hardwoodgenomics.org Selection of mapping parents SM2 SM1 Predominant pollinator? Comparative Genomics of Hardwood Tree Species http://www.hardwoodgenomics.org Progeny Exclusion for Full Sib Linkage Mapping Year Acorns SM2 genotyped pollinated 2000 326 101 2003 674 468 2007 362 110 2010 549 399 • All acorns genotyped across 6 microsatellite markers. • Also all potential pollinators within 200 m of SM1 • Exclude all acorns except those pollinated by SM2 • Parentage analysis done with CERVUS. Comparative Genomics of Hardwood Tree Species http://www.hardwoodgenomics.org Pollination rates in SM1 by SM2 across years SM2 paternity (%) across years Our current mapping population composed of 510 full sibs. Comparative Genomics of Hardwood Tree Species http://www.hardwoodgenomics.org SSR markers developed and mapped Developing codominant SSRs Primary amplification test. Temperature gradient. Test of informativeness in parents. Informative markers configurations ab x cd, ef x eg, hk x hk, nn x np, lm x ll SSR Mapping 37 gSSRs ( 26 with CA repeats, 11 with GA repeats) 39 Q. rubra EST-SSRs from FGP ( 181 screened & 148 amplified) 31 Q. robur EST-SSRs ( 113 screened & 99 amplified) 1 EST-SSR from Castanea mollissima. Comparative Genomics of Hardwood Tree Species http://www.hardwoodgenomics.org SSR Framework Map of Q.rubra 1 (6) 2 (8) 3 (1) 4 (2) Comparative Genomics of Hardwood Tree Species http://www.hardwoodgenomics.org 5 (5) 6 (12) 7 (4) 8 (7) Comparative Genomics of Hardwood Tree Species http://www.hardwoodgenomics.org 9 (3) 10 (9) 11 (10) 12 (11) Comparative Genomics of Hardwood Tree Species http://www.hardwoodgenomics.org Comparative Mapping Marker Name Linkage Gr. in Q. rubra Linkage Gr. in Q. robur FIR 021, FIR027, GOT040, 1 6 GOT063, PIE111, VIT142 PIE099,PIE101 2 8 PIE020 3 1 FIR065, GOT066,POR023 4 2 FIR053, PIE100 5 5 FIR008,GOT009, PIE039, 6 12 PIE126, PIE 236 PIE002, PIE228, WAG016 7 4 FIR030, FIR095, GOT004, 8 7 PIE164 PIE027 9 3 FIR026, PIE028 10 9 POR006 11 10 FIR104 12 11 Comparative Genomics of Hardwood Tree Species http://www.hardwoodgenomics.org Fagaceae Mapping Parents Screening Panel Screening Panel included mapping parents from: American white oak (Quercus alba) American beech (Fagus grandifolia) Chinese chestnut (Castanea mollissima) American chestnut (Castanea dentata) English oak (Quercus robur) Sessile oak (Quercus petraea) Interspecific cross of Quercus robur x Quercus petraea Comparative Genomics of Hardwood Tree Species http://www.hardwoodgenomics.org Amplification of Q.rubra markers across Fagaceae Mapping Parent Test Panel 90 80 70 60 50 CA_gSSRs 40 EST-SSRs markers markers amplified % 30 20 10 0 Quercus alba Fagus Castanea Castanea Quercus Quercus Q.robur x grandifolia mollissima dentata robur petraea Q.patrea Comparative Genomics of Hardwood Tree Species http://www.hardwoodgenomics.org High density RAD-tag Mapping in Quercus rubra Experimental approach: • High quality DNA was extracted using QIAGEN for 250 full sibs. • Genomic DNA was double digested with EcoRI-MseI for producing RAD-tags. • Illumina HiSeq platform used for sequencing. • High coverage parent lane (one lane/2 parents) • 5 lanes of 50 progeny each. Bioinformatics: • Reference loci identified from high coverage parent allele calls. • Full sibs genotyped relative to parental allele calls. • Data formatted and changed into a JoinMap readable format. Comparative Genomics of Hardwood Tree Species http://www.hardwoodgenomics.org JoinMap 4.1 • Pre-mapping quality control Missing data Extreme deviation from Mendelian expectation Similarity of loci. • Grouping LOD value of 10-20 for grouping test statistic. Grouping tree visually checked for selection of groups. Q. rubra and Q. robur gSSR and EST-SSR markers used as anchor tags for identifying LGs. • Mapping Maximum likelihood and Regression mapping algorithm with default criteria. Comparative Genomics of Hardwood Tree Species http://www.hardwoodgenomics.org Comparative Genomics of Hardwood Tree Species http://www.hardwoodgenomics.org Construction of RAD-tag pipeline SAM Tools GATK Total # markers : 32,976 Total # markers : 8,607 >30% missing data: >30% missing data: Oak2 20259 3529 >10 chi square value: > 10 chi square value : 7994 1367 Total # markers : 6977 Total # markers : 5,844 >30% missing data: >30% missing data: Oak3 1740 1432 >10 chi square value: > 10 chi square value : 6 100 Comparative Genomics of Hardwood Tree Species http://www.hardwoodgenomics.org Mapping RAD-tag markers 7994 RAD TAG + 108 SSRs 2nd: 108 SSRs 3rd:108 SSRs 4th:108 SSRs END st 1 : 108 SSRs +200 old RAD +200 RAD from+200 RAD from +392 RAD +200 new RAD 2nd 3rd +200 new RAD +200 new RAD Comparative Genomics of Hardwood Tree Species http://www.hardwoodgenomics.org In silico Comparative Mapping Quercus Castanea Quercus robur rubra mollissima Mapped Mapped Mapped SNP WGS WGS EST-SSRs EST-SSRs Sequence EST-SSRs X X X X RAD- tags X X X X X Comparative Genomics of Hardwood Tree Species http://www.hardwoodgenomics.org QTL Mapping Phenotypes Bud burst Marcescence Leaf morphometrics Leaf nitrogen and carbon content Clonally propagated Q.rubra Phenotypic Data Collection mapping population Bud burst and marcescence 2012, 2013 and 2014 Leaf morphometrics 2013 and 2014 SM1 SM2 Comparative Genomics of Hardwood Tree Species http://www.hardwoodgenomics.org Paper Submission “Development and cross species transferability of genomic and EST microsatellite markers in Quercus rubra. L for comparative mapping in the Fagaceae” ready for submission in Plant molecular biology reporter. First high density northern red oak mapping paper Fall 2014 Bioinformatics' RAD-tag paper Fall 2014 Comparative mapping papers Spring 2015 QTL paper 2015 Comparative Genomics of Hardwood Tree Species http://www.hardwoodgenomics.org JRS lab and ND Acknowledgements Professor. Jeanne Romero Severson Lab Technician: Lauren Fiedler Graduate Students: Daniel Borkowski, Andi Noakes Undergraduate: Rebecca Bullis Collaborators: Dr. Scott Emrich, Olivia Choudhury Notre Dame Genomics and Bioinformatics core facility: Brent Harker Melissa Pullins Grant # TRPGR IOS- 1025947 Comparative Genomics of Hardwood Tree Species http://www.hardwoodgenomics.org.
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