FINE STRUCTURE OF GENE Seymour Benzer (1950-60) Bead Theory: The gene is the fundamental unit of...

FINE STRUCTURE OF GENE Seymour Benzer (1950-60)

Bead Theory: The gene is the fundamental unit of • Structure• Change &• Function

Benzer: if a gene were a sequence of bases it could be divisible at many different locations

Benzer studied of the rII region of T4:(rII mutant causing rapid lysis of E. coli than wild type)

Two types of traits:

plaque morphology

Host range property

1. Permissive host E. coli B; all (rII- & rII+phages grow.

2. Restrictive host E. coli K12; rII+ recombinants grow.

Recombinants of two rII mutants of T4.

Gene order is determined by frequency of recombinants.

If recombination rate is high, genes are far apart.

If recombination rate is low, genes are close together.

Recombination frequency

Intragenic RecombinationRecombination between two mutants to give a

wild type (non-mutant) form of the gene.

O

O XMutant 1

Mutant 2OO

Wild type

Double Mutant

Summary of fine structure mapping of rII region in T4

Seymour Benzer’s conclusion:

Benzer studied 3000 rII mutants showing nucleotide deletions at different levels of subdivision & determined that the rII region is sub-divisible into >300 mutable sites by series of nested analyses (ANOVA) and comparisons.

• hypothesis #1: gene is fundamental unit of change.. ? is wrong..

• Nucleotide pair is the fundamental unit of change/mutation (muton)

Seymour Benzer’s conclusion:

• Benzer’s recombination frequency between some pairs of these was as low as 0.02.

• The T4 genome has 160,000 base pairs of DNA extending over ~1,600 centimorgans (cM).

• So 1 cM 100 base pairs≅• So 0.02 cM represents a pair of adjacent nucleotides.

1. hypothesis #2: gene is fundamental unit of structure…indivisible by crossing over ? is wrong..

2. Nucleotide pair is the fundamental unit of structure (recombination: recon)

Complementation The production of the wild-type phenotype by a cell that contains two mutant genes. The deficiency of one homologue is supplied by the normal allele of the other

F complementation... Fprotein level, mixing of gene productsFno change in genotypes relative to parentsF requires 1 generation of crosses to investigate

F recombination... FDNA levelF results in new genotypes relative to parentsF requires 2 generations of crosses to investigate

complementation or recombination ?

F hypothesis #3: gene is fundamental unit of function... parts of genes cannot function in complementation tests ?

GENE = UNIT OF FUNCTION?

F hypothesis #3: gene is fundamental unit of function... parts of genes cannot function in complementation tests ?... Correct

F a unit of DNA sequence that codes for a single polypeptide or protein

F Cistron: Term coined by Benzer for the smallest genetic unit that does NOT show genetic complementation when two different mutations are in trans position; but shows wild-type phenotype when the same mutations are in cis.

GENE = UNIT OF FUNCTION?

Alternative Splicing ? ? ?(human genes ~30,000 & proteins ~1,20,000 ???)

One Gene / One EnzymeOne Gene / One Polypeptide

“One Gene / One set of connected transcriptsThe definition of a gene by Gerstein et al. [2007] asF ‘a union of genomic sequences encoding a coherent set of

potentially overlapping functional products’

allows genes to have an overlapping sequence, to be alternatively spliced and to exert functions other than protein coding...

GENE = UNIT OF FUNCTION?

Gene definition:Definition 1860s–1900s: Gene as a discrete unit of heredityDefinition 1910s: Gene as a distinct locusDefinition 1940s: Gene as a blueprint for a proteinDefinition 1950s: Gene as a physical moleculeDefinition 1960s: Gene as transcribed codeDefinition 1970s–1980s: Gene as open reading frame (ORF)Definition 1990s–2000s: Annotated genomic entity,Definition (Gerstein et al. 2007) The gene is a union of

genomic sequences encoding a coherent set of potentially overlapping functional products.

TIME LINE OF GENE DEFINITION

GENE = UNIT OF FUNCTION?After ENCODE (Encyclopedia of DNA Elements) PROJECT

FINE STRUCTURE OF GENE Seymour Benzer (1950-60)

Revised Bead Theory:The nucleotide pair is the fundamental unit of • Structure• Change &The gene is the fundamental unit of

FunctionNew Terms: cistron (unit of function), recon (unit of recombination) and muton (unit of mutation).