Project Description
Objective: Our research focuses on elucidating the molecular mechanisms by which auxin controls megaspore mother cell (MMC) initiation or proliferation during ovule development via employing molecular genetic and cell biological techniques. Recently we found that manipulation of auxin signaling causes more MMCs. Thus, we hypothesize that the auxin maximum in the apical cells of nucellus provides a positional cue to restrict the somatic cells entering into the reproductive meiocyte. Moreover, activation of auxin signaling inside the nucellus will promote the MMC initiation or proliferation.
Methodology: We found that the promoter of KNUCKLES (KNU) gene is active in the MMC. In this project, the student will perform genetic analysis to determine whether the YUCCA1, an auxin biosynthesis gene, driven by KNU promoter will increase the number of MMC.
Tasks and Responsibilites
The student will be performing the following experiments:
1. Plant seeeds of DR5:GFP and KNU:GFP in growth chamber.
2. Clone the YUCCA1 gene into pENTR-KNU to make pENTR-KNU:YUCCA1 vector and then make KNU:YUCCA1 by recombination reaction with the pGBW1 vector.
3. Introduce KNU:YUCCA1 into the Agrobacterium GV3101 and conduct plant transformation using DR5:GFP and KNU:GFP plants.
4. Harvest transgenic T0 seeds and screen T0 seeds on plant medium plates containing the hygromycin antibiotic.
5. Transfer transformants into soil.
6. Extract transgenic plants DNA and perform PCR to confirm the transgene.
7. Analyze the phenotype of transgenic plants to determine the number of MMC, the DR5:GFP and KNU:GFP distribution.
Desired Qualifications
None Listed