A Hypothesis for the function of gene AT4G23180 in A. thaliana By 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 Will Effectively Knock Out Our Gene SALK is within an exon and will likely cause a knockout mutation of At4g bp 1395bp Key T-DNA Insertion Site SALK R L

MUTANT PLANT OBSERVATIONS Lab 3.2

Clear Difference in Growth and Development SALK Plants Day 7 - Hypocotyl present Day true leaves Wild Type Plants Day 7 - Cotyledons & 2 true leaves emerging Day true leaves & 2 more emerging

SALK Plants Have Slower 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

Location of At4g23180 Expression

1 st 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 TESTING Lab

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 ADBCADBC

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): 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): 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): Print.