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Transcript
The Role of Genes AT5G48150 and AT2G04890 in Arabidopsis thaliana Seed Development Jennifer Huynh June 8, 2006 Honors Collegium 70AL Professor Bob Goldberg Overview AT5G84150 AT2G04890 GRAS Family mRNA Accumulation Future Research Promoter Regions Conclusions Knockout of Gene 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: AT5G48150 RT-PCR Results Control cDNA: 603 bp GeneChip Data Control Genomic: 922 bp AT5G48150 Gene: 323 bp mRNA Accumulation: AT2G04890 RT-PCR Results Control cDNA: 603 bp GeneChip Data 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 DNA AT5G48150 Plasmid: ~3500 bp Promoter: 2923 bp Gel Bands: 1600 bp, 1300 bp Plasmid: 3500 bp Promoter: 3022 bp Gel Bands: 1800 bp, 1200bp AT2G04890 What happens if the genes are knocked out? Genotyping Insertion Lines AT5G48150 Wildtype: 1508 bp Salk band: 756 bp Found band: 880 bp Wildtype: 1796 bp Salk band: 1740 bp Found band: 1739 bp AT2G04890 Genotyping Insertion Lines AT5G48150 Wildtype: 1508 bp Salk band: 756 bp Found band: 880 bp Wildtype: 1796 bp Salk band: 1740 bp Found band: 1739 bp AT2G04890 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 Globular Heart Torpedo Late Torpedo Wildtype Globular Heart Mutant 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
