GENOME ORGANIZATION AS REVEALED BY GENOME MAPPING WHY

GENOME ORGANIZATION AS REVEALED BY GENOME MAPPING WHY MAP GENOMES? HOW TO MAP GENOMES?

Why map genomes? Understand genome organization Understand genome evolution (comparative mapping) Find and clone important genes Marker assisted selection of important traits

Exploring Genomes: Mapping Genetic mapping l Linkage mapping l l Requires allelic variation and meiotic products Pedigree analysis, LOD scores Physical mapping l l In situ hybridization Somatic cell genetics Radiation hybrids Contig assembly

LINKAGE MAPPING Genetic mapping. Based on recombination in meiosis Requires allelic variation for detection Can be done experimentally with crosses or retrospectively from family studies

Allelic Variation Phenotypic Pigment l Disease l Immunologic Blood type l Tissue antigens l Protein l Electrophoretic variation

Allelic Variation, DNA Restriction Fragment Length Polymorphisms (RFLPs) Microsatellites= Simple Tandem Repeats (STRs) Single Nucleotide Polymorphisms (SNPs)

Pedigree Analysis: AABB x Aa. Bb aabb x aabb 5 children: Aa. Bb, aabb, aa. Bb Are genes A and B linked? Probability assume no linkage: gametes: AB ab a. B Ab (. 25) assume q =. 10 gametes AB ab a. B Ab (. 45) (. 05). 002 liklihood ratio (q =. 10) = (. 25) =. 00097 = 2. 06 Z = 0. 31

Linkage Map: 1 centi. Morgan=1% recombination a b c d __|____________|_____ _ 10 c. M 15 c. M

Linkage map: human chr 12

Interesting notes about linkage maps Different for males and females (in mammals) Female map = 1. 5 x male map Maps are usually presented as sex averaged Different across species Human linkage map = 3200 c. M total Mouse linkage map = 1600 c. M total Different in regions of the same chromosome Not proportional across same physical distance On the average, however: 1 c. M = 1 megabase DNA (human) 1 c. M= 2 megabases DNA (mice)

Somatic Cell Genetics Mapping using interspecific hybrid cells l Advantages Allelic variation not required l Rapid development of a map for any species l l Disadvantages Gene order is difficult to establish l Limited to characters expressed in cell culture l Initially expensive and labor intensive l

Amino Simple Sugars Acids blocked by Aminopterin de novo nucleotide synthesis MAJOR PATHWAY Nucleotides Pyrimidine bases nucleosides Purine bases nucleosides Hypoxanthine Thymidine TK HGPRT SALVAGE PATHWAY HGPRT TK Guanine Phosphorylated Thymidine DNA

Synteny: the occurrence of two loci on the same chromosome, independent of the distance that separates them. Linkage: non-random distribution of alleles in gametogenesis. Tendency of parental combinations of alleles to stay together.

Radiation Hybrids Advantages l l Ordered maps without polymorphism Ordered comparative maps EST maps Integrate EST maps with linkage maps Disadvantages l l Cost of panel construction Differential sensitivity of genotyping markers

Selectable marker Irradiation of donor cells breaks chromosomes Diploid cattle cell line Mutant Recipient rodent cell line Fusion of parental cell lines in PEG Radiation hybrid containing Cattle chromosomal fragments on a rodent genomic background Cattle cells lethally irradiated, ouabain sensitive Non-recombinant rodent cells die in HAT

Ordered Contigs and Sequencing, the Ultimate Physical Map Large Insert Libraries, l BACs and YACs, 200 kb- 1 mb Smaller Insert Libraries l Phage, Plasmids, Cosmids, 20 – 45 kb