Molecular Markers

Morphological Markers

Recessive in nature Mutations - deleterious phenotype Problems with epistasis, pleiotrophy,

incomplete penetrence Influenced by environment Transitory phenotype Difficult to combine

Characteristics of Ideal Polymorphic Markers

Co-dominant expression Nondestructive assay Complete penetrance Early onset of phenotypic

expression High polymorphism Random distribution throughout

the genome Assay can be automated

Isozymes

• The granddaddy of molecular markers

• Lewontin and Hubby 1966– Amino acid substitutions shift

mobility of enzyme through gel

• Folding and charge• Still used today

Isozymes

• Pros:– Moderately easy, well developed protocols– Don’t need genome information– Decades of data to tie into

• Cons:– Low variation– Lots of fresh tissue needed– Many hazardous chemicals

Now 3 Methods of Detection

Restriction fragment length polymorphism and Southern blotting (RFLP)

Polymerase chain reaction (PCR)

Sequence information

Southern blotting

Isolate DNA Digest DNA w/ restriction

enzyme Size fractionate DNA Denature DNA Blot SS DNA to membrane

Methodology

Prepare a probe Label Denature

Hybridize probe with membrane

Rinse Autoradiography

Disadvantages:

The technique is laborious

Time-consuming Expensive May use isotope

Hypervariable Sequences - VNTRs - Minisatellites

Some VNTRs detect polymorphisms at single specific loci.

Other VNTRs detect many bands, making them more useful for forensics.

Microsatellites

Advantages Easy to detect via PCR Lots of polymorphism Co-dominant in nature

Disadvantages Initial identification, DNA sequence information

necessary

Others

AFLPs RAPDs

Single Nucleotide Polymorphisms (SNPs)

SNPs

Polymorphism most used in human genomics

2/3 C → T Coding and non-coding regions Sequence information required High through-put analysis

Nonpolymorphic Markers

Can be used for positional cloning, gene isolation

ESTs (expressed sequence tags) STSs (sequence tagged sites)

Conclusions

Many types of molecular markers available

Type(s) chosen for use will depend on many factors

Dominant or co-dominant, co-dominant preferable

Conclusions, cont.

Now, markers where there is sequence information are preferred to anonymous markers, for sharing, PCR

Polymorphism is necessary for genetic mapping, not for physical mapping

Conclusions, cont.

All molecular markers are not equal. None is ideal. Some are better for some purposes than others. However, all are generally preferable to morphological markers for mapping.