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Click to edit Master title style,Click to edit Master text styles,Second level,Third level,Fourth level,Fifth level,*,Role of Genetic Polymorphismsin Responses to Toxic Agents,Definitions,“Forward genetics” and toxicology,“Reverse genetics” and toxicology,Genetic markers,SNPs and their use in toxicology,Ethical, Legal and Social Issues (ELSI),1,“,Toxicology is concerned with the interaction between xenobiotics and biological molecules directly or indirectly coded in the DNA, and can be regarded as a branch of GENETICS,.”,Michael F.W. Festing (2001),Gregor Mendel (1822 1884),四氟板、尼龙棒、有机玻璃板,2,TERMINOLOGY,Gene: A sequence of DNA bases that encodes a protein,Allele: A sequence of DNA bases,Locus: Physical location of an allele on a chromosome,Linkage: Proximity of two alleles on a chromosome,Marker: An allele of known position on a chromosome,Distance: Number of base-pairs between two alleles,centiMorgan: Probabilistic distance of two alleles,Phenotype: An outward, observable character (trait),Genotype: The internally coded, inheritable information,Penetrance: No. with phenotype / No. with allele,Modified from M.F. Ramoni, Harvard Medical School,3,The 80s Revolution and,the Human Genome Project,Genetic Polymorphisms: naturally occurring DNA markers that identify regions of the genome and vary among individuals,The intuition that polymorphisms could be used as markers sparkled the revolution,On February 12, 2001 the Human Genome,Project announced the completion of a first,draft of the human genome and declared:,“A SNP map promises to revolutionize both,mapping diseases and tracing human history”,SNP are Single Nucleotide Polymorphisms subtle,variations of the human genome across individuals,Modified from M.F. Ramoni, Harvard Medical School,4,DISTANCES ON A GENETIC MAP,Physical distances between alleles are base-pairs,But the recombination frequency is not constant,A useful measure of distance is based on the probability of recombination: the Morgan,A distance of 1 centiMorgan (cM) between two alleles means that they have 1% chance of being separated by recombination,A genetic distance of 1 cM is roughly equal to a physical distance of 1 million base pairs (1Mb),Modified from M.F. Ramoni, Harvard Medical School,5,Physical Maps: maps in base-pairs,Human physical map: 3000Mb (Mega-bases),Genetic Maps: maps in centiMorgan,Human Male Map Length: 2851cM,Human Female Map Length: 4296cM,Correspondence between maps:,Male cM 1.05 Mb; Female cM 0.88Mb,MORE TERMINOLOGY,Modified from M.F. Ramoni, Harvard Medical School,6,Single Gene (Mendelian) diseases:,Autosomal dominant (Huntington),Autosomal recessive (Cystic Fibrosis),X-linked dominant (Rett),X-linked recessive (Lesch-Nyhan),Today, over 400 single-gene diseases have been identified,Problem: traits dont always follow single-gene models,Complex Trait: phenotype/genotype interaction,Multiple cause:,multiple genes in several loci determine a phenotype in conjunction with non-genetic factors (accidents of development, social factors, environment, infections, other factors),Multiple effect:,gene causes more than one phenotype,Simple and Complex Traits,Modified from M.F. Ramoni, Harvard Medical School,7,Genetic Markers,Even though we share most DNA, there are variations (polymorphisms),Polymorphic: two or more forms of the same gene, or genetic marker exist with each form being too common in a population to be merely attributable to a new mutation,Classes of polymorphic genetic markers:,Single Nucleotide Polymorphisms (SNP): single base differences in population,Microsatellites:,short tandem repeat (e.g. GATA, 2 6 bp long),Minisatellites: simple sequence repeats (10 40 bp long),Variable Number of Tandem Repeats: the number of repeats may vary,Restriction Fragment Length Polymorphisms: presence/absence of a site,Deletions, Duplications, Insertions: alterations on a chromosome level,Complex haplotypes: combinations of the above,8,Genetic Markers,Coding:,Single Nucleotide Polymorphisms,Restriction Fragment Length Polymorphisms,Deletions, Duplications, Insertions,Non-coding:,Microsatellites,Minisatellites,Variable Number of Tandem Repeats,Restriction Fragment Length Polymorphisms,Single Nucleotide Polymorphisms,Deletions, Duplications, Insertions,9,Polymorphisms (allelic variations) are essential to:,Study inheritance patterns,Map phenotypes and anchor genes to the genetic map by co-segregation analysis,Determine change in function: resistant/sensitive populations,Genetically determined variability among humans is due to a difference in 0.1% of the genomic sequence!,Polymorphisms can be silent, or be exhibited at levels of:,Morphology,Protein,DNA,Genetic Markers,10,Insertion Deletion,Chromosomal rearrangements: Deletions, Duplications, Insertions,Deletions:,a certain part is lost, for example,abc,ac,Insertions:,a part is added, for example,ac,abc,Duplications:,can be tandem, for example,abc,abbc, or not, for example,abc,abcabc,Reversals:,a part is turned around, head to tail,abc,cba,Transpositions:,two parts change places, for example,abcd,acbd,11,Copy Number Variability (CNVs),CNV are DNA segments at 1kb or larger with a variable number of copies in comparison with a reference genome. CNV can have dramatic phenotypic consequences as a result of altering gene dosage, disrupting coding sequences, or perturbing long-range gene regulation.,There are several well-known examples of CNV, including,CYP2A6, CYP2D6, GSTM1, GSTT1, SULT1A1, SULT1A3, UGT2B17,and also the nearby,UGT2B7, UGT2B10,and,UGT2B11 genes,. All these genes are deleted at a relatively high frequency in at least one ethnic group. In addition,CYP2A6, CYP2D6, SULT1A1,and,SULT1A3,can also present duplications and even multiduplications.,Pharmacology & Therapeutics 116, Issue 3, 2007, Pages 496526,12,Original DNA fingerprinting technique,Relies on stretches of tandemly repeated sequences (usually 15 - 100bp),Alleles show high variability in numbers of repeats,Genotyping using minisatellites:,Digest genomic DNA,Run out on gel,Southern blot and probe with radiolabelled repeat DNA,Individuals appear with a set of bands unique to them, although each band is shared with one of their parents,Minisatellites,13,Microsatellites,Number of repeats varies greatly between individuals,Make up to 10-15% of the mammalian genome,Believed to have no function,Have high mutation rates,Used in forensic analysis,Can be amplified by PCR fragments that are generated have different length due to different number of repeats,14,Microsatellites are highly polymorphic due to potential for,“skipping” during DNA replication,15,
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