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

Tomato leaf curl disease

Cotton leaf curl disease

Experimental Design

FACS “synchronization”One hour BrdU Pulse

Fix Cells and Isolate Nuclei

IP BrdU-labeled DNA

Label IP and Input DNA with Cy3/5

Hybridize to Chr4 Tiling Array

Normalize

Smooth

20000 probes1 kb PCR products

95% coverage

Arabidopsis Chromosome 4

nucleolarorganizing region

centromere

genes transposable elements

chromosome coordinate

log

(ear

ly/la

te)

E

L

Replication Timing Profile

E E E EL L L L L

Origin assembly

Initiation complexes assemble in late M and early G1

Pre-replication complex

ORC - origin recognition complexMCM - replicative DNA helicaseCDT1/CDC6 - MCM loader

licensing

MCM5/7 are released during mitosis

MCM5

MCM7

Merge DAPI DIC

N. tabacum cultured cells (Nt-1)

How is licensing regulated in plants?

nucleuscytoplasm

MCM subunits

A different CDT regulatory protein?

Geminin

X

Other mechanisms?

ORC1 regulation?

CDT1

ProteosomeCDKA

P

X

?

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 Activities1) Heredity & Genetics Scavenger Hunt2) Personal Survey of Inherited Human Traits3) Modeling Mendel 4) Punnett Squares5) Genetic Offspring Models6) Extracting DNA from Wheat Germ7) DNA, Genes, and Proteins (Sickle Cell Disease)8) A Day in the Life of DNA 9) Genetic Disorders10)Genetic vs. Environmental Influences on Traits

Activity 2: Survey of Inherited Traits Activity 5: Genetic Offspring Models

Activity 6: Extracting Wheat DNA Activity 8: Day in the Life of DNA

Genetics Independent Study Group