Will that be allele cache or
Mendelian credit? Evidence for
the inheritance of ancestral
sequences in Arabidopsis.
Susan J. Lolle
Purdue University
National Science Foundation
Arabidopsis wild-type
Fusion mutants
FIDDLEHEAD defined a class of mutations
that resulted in ectopic organ fusion
Two heads are better than one...
hothead - phenotype
hth-10
• Organ fusion
• Pollen hydration on
vegetative surfaces
• Increased rate of
chlorophyll extraction
• Limited self-fertility
All hothead mutations are recessive
Both copies of the
gene must have a
mutation to reveal
the mutant
phenotype
Fundamentals of Mendelian
Genetics
• Two alleles per locus
• Alleles can be dominant
or recessive
• Alleles are stable from one
generation to the next, whether or
not they contribute to the observed
phenotype
Self-fertilizing verses
Cross-fertilizing
Progeny should all be homozygous
hothead
Stable Inheritance
100%
parent
progeny
But, some progeny are wild-type!
90%
10%
Mechanism?
Arabidopsis wild-type
Fusion mutant
Sequencing of HOTHEAD
•All mutant alleles contain a single point
mutation in the HOTHEAD gene
•Columbia and Landsberg wild-type alleles
very similar; Ws quite different
Using Molecular Markers to Genotype
Allele ‘A’
PCR amplify
(using specific primers)
Restriction Enzyme
Digestion
Gel electrophoresis
Allele ‘a’
Does phenotype relate to
genotype?
Does phenotype relate to genotype?
What does the sequence tell us?
I don’t know if
this is such a
wise thing to
do, Bob.
Is reversion really contamination?
Is this a mistake?
•Seed contamination due to wild-type
plants grown in close proximity to
mutants
•Elevated levels of outcrossing in
hothead mutants (who is the daddy?)
Embryonic revertants
hth-4/hth-4
HTH/HTH
parent
HTH/hth
hth/hth
# of
embryos
2
57
301
%
<1
16
84
•Revertants detected as embryos cannot be
due to contaminating seed
Explanations for Genetic Instability
What does the sequence tell us?
•Transposon-induced alleles
•Inverted or direct repeats at the locus
•Epigenetic alleles
•High rate of random mutation
•Gene conversion
Gene conversion with other family
members
HTH
hth-10
At HTH-like
At HTH-like
At HTH-like
At HTH-like
At HTH-like
At HTH-like
At HTH-like
1
2
3
4
5
6
7
ACT
ACT
GTT
CCA
CCA
GTC
GCC
GCT
GTC
GTT
GTT
GTT
CCT
CCT
GTG
GTA
GTT
GTT
GGA
GAA
GGG
CAA
CAA
GGT
GGA
GGG
GGA
ATT
ATT
ATT
GTT
GTT
GTT
ATC
ATC
ATC
ACA
ACA
ACC
GTA
GCA
ACT
ACC
ACA
ACA
•None of these sequences could provide correction
of the mutant nucleotide without introducing other
changes
Instability of molecular markers in hth
background
Marker
hth/hth
HTH/HTH
Location in
gene
AG
4% (7/186)
0% (0/190)
intron
GAPC
4% (9/242)
0% (0/190)
intron
GL1
1% (1/90)
0% (0/196)
3’-UTR
HTH
2% (10/484)
0% (0/590)
exon
RGA
3% (14/402)
0% (0/386)
exon
UFO
4% (16/438)
0% (0/196)
exon
Where did these sequences come
from?
• They were not present in the parental
genome
 molecular and genetic evidence
• They were present in the ancestors
 molecular and genetic evidence
A DNA template cannot be
detected in the parental genome
Evidence strongly supports a
‘template-directed’ process.
Is the template RNA?
Where is it?
How long does it persist?
What might this mean?
“@$#?!$”
•Organisms could carry
sequence information for
more than two alleles
•Mechanism might help selffertilizing species avoid the
negative consequences of
inbreeding
Should you worry about your
ancestry?
How long does the allele cache persist?
Is it quality cache?
Acknowledgements
National Science Foundation
Collaborator:
Bob Pruitt
People:
Katie Krolikowski
Ryan Lee
Tina Nussbaum
Wagler
Liese Pruitt
Trulie Thorley
Mirayda
Torres
Jen Victor
Jessica
Young