2. Geneticist Definition of Gene  If you ask 10 different geneticist to define gene, you would get 10 different answers  Genes must involve information  Information must be contained in a molecule  Need to define nature of information  Then look at molecules

3. Mutations as a Tool  To geneticist mutations are data  Mutations shows us the effect of removing or disrupting a process  If you are interested in a process  Then make mutants to disrupt the process  Number of different mutants will tell you steps in the process  Can be used to determine order of events  Combinations of mutants: double mutants

4. Functional Test for Allelism  Two mutants with the same phenotype  Are they alleles of one gene?  Do they represent alleles of two different genes?  We could isolate the genes, sequence them and compare...  Can we do this with simple genetic crosses?  Complementation Test

5. Complementation Test  Only works with recessive alleles  Logic is simple:  Cross homozygous mutants—>heterozygote  Assess phenotype  If wild type (doubly heterozygous)  Then different genes, we say they complement (provide wild type function)  If mutant (heteroallelic)  Then alleles of same gene, we say they fail to complement (no wild type function)

6. Treat with mutagen Isolate (recessive) mutants Make homozygous Mutant 1 Mutant 2 Mutant 3 Mutant 4 How many genes do these represent?

7. Consider Single Test X X w1 w2 + w1 + w2 w1a w1b w1 w2 + w1 + w2 w1a w1b complements (+) fails to complement (-)

8. Test All 4 Mutants M1M2M3M4 M1 M2 M3 M4 – + + – – – + – + – Allelic Relationships: 2 genes, each with 2 alleles Gene 1: with alleles M1, M4 Gene 2: with alleles M2, M3

9. Genes and Proteins  Relationship of genotype and phenotype  First clue came from Beadle and Tatum:  Studied synthesis of arginine in Neurospora  Isolate mutant alleles of genes  Correlate with biochemical pathway  Mutants identified by failure to make Arg  Call this kind of mutant auxotroph  Supplement media with Arg = growth  No Arg in media = no growth  Supplement with intermediates = pathway

10. precursor Media Supplement Mutant Ornithine Citrulline Arginine Wild type+ + + Mutant A+ + + Mutant B- + + Mutant C- - + ornithinecitrullinearginine CBA

11. Conclusions?  Supplement with intermediate after block in pathway = growth  Can dissect pathways using genetics  Beadle and Tatum concluded each gene encoded a single enzyme in the pathway:  One gene-one enzyme  One gene one polypeptide is better  Even this is an oversimplification  Begins to get at the relationship

12. Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display. Chromo- some Gene 1 Enzyme E GlutamateOrnithineCitruline Argino- succinate Arginine Enzyme FEnzyme GEnzyme H Encoded enzyme biochemical pathway Gene 2 Gene 3 & 4 arg-H arg-G arg-F arg-E

13. CDMB  DNA makes RNA makes Protein DNARNAProtein  DNA Stores information, and is replicated  RNA contains information in DNA  RNA is used to direct synthesis of proteins transcriptiontranslation replication

14. CDMB  A more modern version is below: DNA RNAProtein  This encompasses the original idea  DNA can be made from RNA (reverse transcription)