mutation detection systems navi
TRANSCRIPT
PREPARED BY
NAVEENA GIRISH
DEPARTMENT OF PLANT SCIENCE
CENTRAL UNIVERSITY OF KERALA
2 TYPES
VISIBLE SLCBIOCHEMICAL
SLC
WIDELY USEDUses 5 or 7 loci
Uses up to 20 enzymes
T O DETECT QUANTITATIVE MUTATION IN MAMMALIAN
GERMLINE CELLS
A visible specific locus mutation is a genetic change that
alters factors responsible for coat color and other visible
characteristics of certain mouse strains
principle
to cross individuals who differ with respect to the genes
present at certain specific loci, so that a genetic alteration
involving the standard gene at any one of these loci will
produce an offspring detectably different from the standard
heterozygote.
FEMALE
HOMOZY
GOUS
MALE +
TEST
REAGENT
PROGENY
SCREENING FOR
MUTATION
BIOCHEMICAL SPECIFIC LOCUS TEST
A biochemical specific locus mutation is a genetic change resulting from a DNA lesion causing alterations in proteins that can be detected by electrophoresis methods.
The principle of the MBSL is that heritable damage to the genome can be detected by electrophoresis analysis of proteins in the tissues of the progeny of mice treated with germ cell mutagens
Treated males are then mated to untreated females to produce F1 progeny. Both blood and kidney samples are taken from progeny for electrophoresis analysis. Up to 33 loci can be examined by starch-gel electrophoresis and broad-range isoelectric focusing. Mutants are identified by variations from the normal electrophoresis pattern. Presumed mutants are bred to confirm the genetic nature of the change.
Single generation & visual detection is used to identify
mutation – SO USEFUL
OFFSPRING EXAMINATION
BIRTH AND WEANING
TISSUE SAMPLING
ELECTROPHORESIS
MUTANT IDENTIFICATION
BLOOD -6 LOCI
KIDNEY – 27 LOCI
AMES TEST
Industrial chemicals potentially harmful
to humans
We can know it using lab animals
But it is time consuming and expensive
Bruce Ames 1974
PRINCIPLEDamage to DNA mutation Cancer
90% carcinogen = mutagenMutation in bacteria = carcinogenesis in humans
I used Salmonella typhimurium
with dna repair system
inactivated and defects in
lipopolysacharide
4 strains used
carcinogenic compounds in hair dyes removed by the help of
Ames test
One of the four strains detect base pair substitutions
Other three detect frameshift mutation
DROSOPHILA
MELANOGASTER
DEFINED MUTATION RATE
INTIAL EXPERIMENTS WAS
FAIL
BUT 1927 BERLIN
CONFERENCE HE SURPRISED
OTHERS
X- RAYS INDUCES
MUTATION
7 YEARS
BUT HE SAID NO QUALITATIVE DIFFERENCE BETWEEN SPONDANEOUS AND
INDUCED MUTATIONS
ClB METHOD – TO STUDY MUTATIONS HAPPENING ON X CHROMOSOME
He received Nobel prize in 1945
MULERS -5 METHOD
TO TEST X CHROMOSOME
The progeny of F2 generation indicate
whether or not lethal mutations occurred on
males x chromosomes
If lethal mutation occurred on wild type male
– it die
C – prevents crossing over & recombination
So lethality reside in one chromosome itself
C – prevents cross over
There is 2 markers
B – bar eye
Wa for apricot
recessive mutation can only express if
there is homozygosity
Attached x – to screen F1 mutations on x
of males
X^X – REPRESENTATION
Compound chromosome 2 x chromosome
attached
Inheritance as single unit
X^X female
ATTACHED X CHROMOSOME