Novel Genetic Tools to Accelerate Our Understanding of Photosynthesis and Lipid Accumulation
Final rept. 15 May 2013-14 May 2014
CARNEGIE INST OF WASHINGTON DC
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Green algae are a promising source of biofuel, but the genetic tools for studying them are very limited. In this AFOSR YIP project, a transformative genetic screening tool, ChlaMmeSeq, was developed to increase the throughput of genetic screens in the green alga Chlamydomonas by 1,000-fold. ChlaMmeSeq can be applied to pools of tens of thousands of mutants to simultaneously i determine the sites of insertion of an exogenous DNA fragment into the genomes and ii track the growth rates of individual mutants. ChlaMmeSeq was used to reveal the insertion sites in tens of thousands of insertion mutants, an unprecedented scale for green algae. The data revealed insights critical for future screens and for improving expression of foreign genes in Chlamydomonas. Furthermore, ChlaMmeSeq was used to quantify the growth rates of 10,000 mutants in a pool, under photosynthetic and non-photosynthetic growth. 80 mutants with photosynthetic growth defects were identified and are the subject of current study. Finally, a fluorescence-activated cell sorting method compatible with ChlaMmeSeq was developed to enable screens for mutants with high lipid content.
- Genetic Engineering and Molecular Biology