1. Will that be allele cache or Mendelian credit
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

2. Arabidopsis wild-type
Fusion mutants

3. FIDDLEHEAD defined a class of mutations that resulted in ectopic organ fusion

4. Two heads are better than one...

5. hothead - phenotype Organ fusion
Pollen hydration on vegetative surfaces
Increased rate of chlorophyll extraction
Limited self-fertility
hth-10

6. All hothead mutations are recessive
Both copies of the gene must have a mutation to reveal the mutant phenotype

7. 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

8. Self-fertilizing verses Cross-fertilizing

9. Progeny should all be homozygous hothead
Stable Inheritance
parent
100%
progeny

10. But, some progeny are wild-type!
90%
10%

11. Arabidopsis wild-type
Mechanism?
Arabidopsis wild-type
Fusion mutant ?

12. 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

13. Using Molecular Markers to Genotype
Allele ‘A’
Allele ‘a’
PCR amplify
(using specific primers)
Restriction Enzyme
Digestion
Gel electrophoresis

14. Does phenotype relate to genotype?

15. Does phenotype relate to genotype?

16. What does the sequence tell us?

17. I don’t know if this is such a wise thing to do, Bob.

18. 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?)

19. Embryonic revertants
hth-4/hth-4 parent
HTH/HTH
HTH/hth
hth/hth
# of embryos 2 57
301 %
<1 16 84
Revertants detected as embryos cannot be due to contaminating seed

20. 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

21. Gene conversion with other family members
HTH ACT GTT GGA ATT ACA
hth ACT GTT GAA ATT ACA
At HTH-like 1 GTT GTT GGG ATT ACC
At HTH-like 2 CCA CCT CAA GTT GTA
At HTH-like 3 CCA CCT CAA GTT GCA
At HTH-like 4 GTC GTG GGT GTT ACT
At HTH-like 5 GCC GTA GGA ATC ACC
At HTH-like 6 GCT GTT GGG ATC ACA
At HTH-like 7 GTC GTT GGA ATC ACA
None of these sequences could provide correction of the mutant nucleotide without introducing other changes

22. Instability of molecular markers in hth background
hth/hth
HTH/HTH
Location in gene AG
4% (7/186)
0% (0/190)
intron
GAPC
4% (9/242)
GL1
1% (1/90)
0% (0/196)
3’-UTR
HTH
2% (10/484)
0% (0/590)
exon
RGA
3% (14/402)
0% (0/386)
UFO
4% (16/438)

23. Where did these sequences come from?
They were not present in the parental genome
molecular and genetic evidence
They were present in the ancestors

24. 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?

25. What might this mean?
Organisms could carry sequence information for more than two alleles
Mechanism might help self-fertilizing species avoid the negative consequences of inbreeding

26. Should you worry about your ancestry?
How long does the allele cache persist?
Is it quality cache?

27. National Science Foundation
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