The Role of Genes AT5G48150 and AT2G04890

in Arabidopsis thaliana Seed Development

Jennifer HuynhJune 8, 2006

Honors Collegium 70ALProfessor Bob Goldberg

Overview

GRAS Family

Knockout of Gene

Future Research

mRNA Accumulation

Promoter Regions

AT5G84150AT2G04890

Conclusions

What has been already discovered about the genes?

The GRAS Family• Name derives from the three initially

identified members– GAI, RGA, and SCR

• Functions range from meristem maintenance to regulation of hormone and light signal transduction

• PAT1 branch plays role in far red light signaling

• Homologs found in higher plants– Tomato, petunia, lily, rice, barley

The GRAS Family Tree

The GRAS Family• Name derives from the three initially

identified members– GAI, RGA, and SCR

• Functions range from meristem maintenance to regulation of hormone and light signal transduction

• PAT1 branch plays role in far red light signaling

• Homologs found in higher plants– Tomato, petunia, lily, rice, barley

Gene One: AT5G48150• Scarecrow-like transcription factor 21 (SCL21)

Gene Size: 2355 bp Protein Size: 491 aa

Gene Two: AT2G04890• Phytochrome A signal transduction 1 (PAT1)

Gene Size: 2601 bp Protein Size: 414 aa

Where is the gene transcribed into mRNA?

mRNA Accumulation: AT5G48150RT-PCR Results

GeneChip DataControl cDNA: 603 bp

Control Genomic: 922 bp

AT5G48150 Gene: 323 bp

mRNA Accumulation: AT2G04890RT-PCR Results

GeneChip DataControl cDNA: 603 bp

Control Genomic: 922 bp

AT2G04890 Gene: 499 bp

Is the upstream region of the gene responsible for the

transcription of the gene?

Restriction-Digested Plasmid DNAAT5G48150

AT2G04890

Plasmid: 3500 bpPromoter: 3022 bpGel Bands: 1800 bp, 1200bp

Plasmid: ~3500 bp Promoter: 2923 bp

Gel Bands: 1600 bp, 1300 bp

What happens if the genes are knocked out?

Genotyping Insertion LinesAT5G48150

AT2G04890

Wildtype: 1796 bpSalk band: 1740 bp Found band: 1739 bp

Wildtype: 1508 bp

Salk band: 756 bp

Found band: 880 bp

Genotyping Insertion Lines

AT5G48150

AT2G04890

Wildtype: 1796 bpSalk band: 1740 bp Found band: 1739 bp

Wildtype: 1508 bp

Salk band: 756 bp

Found band: 880 bp

AT5G48150: SALK_064220• Wildtype: 1796 base pairs• Salk band: 1740 base pairs • Found band: 1739 base pairs

AT2G04890: SALK_146085

• Wildtype: 1508 base pairs• Salk band: 756 base pairs• Found band: 880 base pairs

Nomarski Photographs

Wildtype

Mutant

Globular

Globular

Heart

Heart

Torpedo

Torpedo Late Torpedo

Late Torpedo

Conclusions• No mutant phenotype on flowers, leaves,

or siliques for either AT5G48150 or AT2G04890

• Possibilities– Genes not vital for seed development– Similar genes exist that compensate– Mutant phenotype not visible in experiments

• More research is necessary

Future Research

• Perform RT-PCR on homozygous mutants seeds to observe transcription

• Fuse GFP marker with promoter to observe expression

• Cross homozygous mutant plants of different knockouts

Acknowledgements

• Bob Goldberg

• Anhthu Bui

• Mike Gavino

• Jonathan Russell

• Ria Yagnik

• Brandon Le

• Xingjun Wang

• Tomo Kawashima

• Jessica Luke

• HC 70AL Classmates