HighResolution Physical Mapping of the Apple Genome by BAC Fingerprinting

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High-Resolution Physical Mapping of the Apple
Genome by BAC Fingerprinting

• Schuyler Korban Jonathan Beever
• University of Illinois at Urbana-Champaign

USDA-CSREES NRI Award 2005-35300-15538
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Apple One of the Three Pillars of the Rosaceae
Model System

• Apple is a diploid with 2n 34
• A relatively small size genome 750 Mb or 1.54 pg
  DNA/2C
• Availability of genetic linkage maps with 840
  molecular markers (RAPDs, AFLPs, and SSRs)
• Map-based cloning strategies have been successful
• Efficient systems for apple transformation and
  regeneration of transgenic lines for multiple
  apple cvs.
• Availability of bacterial artificial chromosome
  (BAC) libraries
• Availability of over 200,000 expressed sequence
  tags (ESTs) in the public database

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Overall Goal

• Generate a whole-genome map for the apple

Rationale
Although availability of molecular markers and
several genetic linkage maps have contributed to
our genetic knowledge of qualitative as well
complex traits, developing a whole-genome
clone-based physical map, consisting of ordered
recombinant genomic clones spanning the entire
genome, is essential for large-scale genome
sequencing, efficient positional cloning of major
genes and QTL, target DNA marker development, and
provides a powerful platform for high-throughput
EST physical mapping
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Objectives

• Construct a genome-wide framework for a
  high-resolution physical map of the apple genome
  by BAC fingerprinting
• Create a panel of BAC DNA pools for PCR-based
  screening of 10X coverage of the apple genome
• Discover, characterize, and map EST-SSRs
• Anchor BAC-fingerprinted contigs to current
  genetic and physical marker maps by STS content
  mapping

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Overall Strategy
Develop a BAC-based physical map using an agarose
gel-based fingerprinting method of 70,000 BAC
clones
Fingerprinting data will be used to cluster
highly-related clones, further refined by manual
clustering, and followed by merging of contigs of
related clones to assemble the BAC map
In parallel, will use the apple EST database to
develop EST-derived SSRs, and integrate them into
a genetic map
Use all genetic markers to anchor apple BAC
contigs to the apple genetic map, apple EST
unigenes will be anchored to BAC contigs, and a
comprehensive apple transcript map will be
developed
A BAC tiling path will be determined, and a set
of BACs with minimal overlap will be established
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Project Deliverables

• A preliminary physical map will be generated by
  the second year of the project
• A final genetic and physical map of the apple
  genome will be completed by the end of the third
  year
• Provide several thousand nucleation points
  suitable for fine mapping, positional cloning,
  and sequence-based identification of genome
  regions containing QTL

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Project Team
Schuyler Korban, PI Jonathan Beever,
Co-PI Yuepeng Han, Postdoc Research Assoc Ksenija
Gasic, Postdoc Research Assoc
Bioinformatics/ Data Management Lei Liu,
Collaborator Jyothi Thimmapuram,
Collaborator Dorrie Main, Collaborator