Why study DNA replication?
Attractive model for multicellular eukaryotes
•Small, sequenced genomes
•Genomic tools available
•Rapidly dividing cell cultures
•Tractable developmental programs
Arabidopsis
Rice
Intercalary meristem
Leaf
blade
Leaf
sheath
Why study DNA replication?
Potential practical applications
•Genetic engineering of artificial chromosomes
•Manipulating the endocycle
•New strategies for geminivirus resistance
Cotton leaf curl disease
Tomato leaf curl disease
Experimental Design
One hour BrdU Pulse
FACS “synchronization”
IP BrdU-labeled DNA
Label IP and Input
DNA with Cy3/5
Hybridize to Chr4
Tiling Array
20000 probes
1 kb PCR products
95% coverage
Normalize
Fix Cells and Isolate
Nuclei
Smooth
Arabidopsis Chromosome 4
genes
nucleolar
organizing region
centromere
transposable elements
Replication Timing Profile
L
E
L
E
L
E
log (early/late)
E
L
chromosome coordinate
L
E
L
Origin assembly
Pre-replication complex
Initiation complexes
assemble in late M
and early G1
licensing
ORC - origin recognition complex
MCM - replicative DNA helicase
CDT1/CDC6 - MCM loader
MCM5/7 are released during mitosis
Merge
MCM5
MCM7
N. tabacum cultured cells (Nt-1)
DAPI
DIC
How is licensing regulated in plants?
X
Proteosome
CDKA
P
?
A different CDT
regulatory protein?
Geminin
ORC1
regulation?
CDT1
MCM subunits
X
cytoplasm
Other
mechanisms?
nucleus
Exploring Genetics across the Middle School
Science and Math Curriculum
Classroom visits
Exploring Genetics across the Middle School
Science and Math Curriculum
Classroom visits
Dr. Jeff Batten - Overview science activities
Genetics Kit Activities
1) Heredity & Genetics Scavenger Hunt
2) Personal Survey of Inherited Human Traits
3) Modeling Mendel
4) Punnett Squares
5) Genetic Offspring Models
6) Extracting DNA from Wheat Germ
7) DNA, Genes, and Proteins (Sickle Cell Disease)
8) A Day in the Life of DNA
9) Genetic Disorders
10)Genetic vs. Environmental Influences on Traits
Activity 2: Survey of Inherited Traits
Activity 6: Extracting Wheat DNA
Activity 5: Genetic Offspring Models
Activity 8: Day in the Life of DNA
Genetics Independent Study Group