a hypothesis for the function of gene at4g23180 in a. thaliana by nicole foxworth and deborah lee...

A Hypothesis for the function of gene AT4G23180 in A.

thalianaBy Nicole Foxworth and Deborah

Lee(Ether Fowl Ox)

CREATING A GENE MODEL

Lab 2.2

Predicted Gene Structure

GENE KNOCKOUT MUTATION

Lab 3.1

SALK 116653 Will Effectively Knock Out Our Gene

SALK 116653 is within an exon and will likely cause a knockout mutation of At4g23180.

1860bp1395bp

Key

T-DNA Insertion Site

SALK 116653

R L

MUTANT PLANT OBSERVATIONS

Lab 3.2

Clear Difference in Growth and Development

SALK Plants

Day 7- Hypocotylpresent

Day 15- 4 true leaves

Wild Type Plants

Day 7- Cotyledons &2 true leaves emerging

Day 15- 5 true leaves &2 more emerging

SALK Plants HaveSlower Onset of Leaf

Development

Effect of Low Fe Medium is Inconclusive

Low Fe Medium

Day 9

- No seeds germinated

MS Medium

Day 9

- One SALK plant germinated

RESEARCHING GENE FUNCTION: BIOINFORMATICS

Lab 4.1

Protein Kinase Catalytic Domain is a Conserved

Domain in our Gene

CRK10/RLK4 is a Perfect Match to our Predicted Protein Sequence

RESEARCHING GENE FUNCTION: LITERATURE SEARCH

Lab 4.2

RLK4 is Expressed in the Shoot Apical Meristem and

Lateral Root• Review paper entitled

“Receptor Kinase Signaling in Plant Development” (Becraft 2002)

• Overview of many proteins in the RLK family, but does not indicate a specific function for RLK4

CRK10 Plays a Role in Plant Defense

• Chen et. al. explored entire CRK family–Found that many CRKs are involved

in plant defense responses–CRK10 is not soley responsible for

plant defense, but plays a role. • Could be due to functional redundancy

of closely related genes

RLK4 is Involved in Response to Pathogen Infection

• Du and Chen find evidence that RLK4 is highly expressed in leaves after pathogen infection.

MICROARRAY RESOURCES

Lab 5.1-5.2

Location of At4g23180 Expression

1st Experiment Idea: Seedling Root Response to Salinity

Stress

Focused Experiment on the Effects of Wounding

Gene At4g23180 expression increases in response to leaf wounding/aphid feeding.

HYPOTHESIS TESTINGLab 5.4-5.6

Hypothesis of Gene Function: Plant Defense

• We hypothesize that our gene is involved in plant defensive response to wounding and is expressed in the leaves.

• We designed an experiment to test this hypothesis.– Poked holes in plant leaves and tested

level of gene expression after 30 minutes.

Experiment Yielded Inconclusive Results

A D

B C

Future Directions

• We maintain our hypothesis that our gene is involved in plant defensive response

• Repeat our designed experiment with the following changes:– Bigger sample size– Quality assessment of RNA extracted

through gel electrophoresis– Design new primers within exons that flank

an intron but are not split in half by one

References

• Becraft, Philip W. "Receptor Kinase Signaling In Plant Development." Annual Review of Cell and Developmental Biology 18.1 (2002): 163-92. Print.

• Chen, Kegui, Baofang Fan, Liqun Du, and Zhixiang Chen. "Activation of Hypersensitive Cell Death by Pathogen-induced Receptor-like Protein Kinases from Arabidopsis." Plant Molecular Biology 56.2 (2004): 271-83. Print.

• Du, Liqun, and Zhixiang Chen. "Identification of Genes Encoding Receptor-like Protein Kinases as Possible Targets of Pathogen- and Salicylic Acid-induced WRKY DNA-binding Proteins in Arabidopsis." The Plant Journal 24.6 (2000): 837-47. Print.