Nuclear transformation of Chlamydomonas reinhardtii with silicon carbide fibers
- National Renewable Energy Lab., Golden, CO (United States)
Efficient nuclear transformation of cell wall-deficient strains of the green alga Chlamydomonas reinhardtii can be accomplished by vortexing the cells in the presence of glass beads and polyethylene glycol (Kindle 1990 PNAS 87:1228). Intact (walled) cells can also be transformed using this protocol, but at very low efficiencies. Two recent reports have described the use of silicon carbide fibers to mediate DNA entry into plant suspension cells (Kaeppler et al. 1990 Plant Cell Rep. 9:414; Asano et al. 1991 Plant Sci. 79:247). The author has found that nuclear transformation efficiencies of walled cells of C. reinhardtii can be increased 3 to 10 fold by vortexing the cells in the presence of silicon carbide fibers and PEG. Using a modification of the glass bead transformation procedure, the wild-type nitrate reductase structural gene was used to complement a NR-deficient mutant of C. reinhardtii, nit-1-305. The transformation efficiency increased with longer vortexing times, although the absolute number of transformants varied between experiments, ranging from 10 to 40 transformants per 10[sup 7] cells. In contrast to vortexing with glass beads, cell viability was very high, with greater than 80% cell survival even after vortexing for 10 minutes. Neither cell death nor transformation efficiency increased when cell wall-deficient mutants (cw15 nit-1-305) were used as compared to intact cells. Experiments are in progress to test the applicability of silicon carbide-mediated transformation to other algal strains for which cell wall mutants or protoplasting procedures are unavailabile.
- OSTI ID:
- 6659608
- Report Number(s):
- CONF-9208211-; CODEN: JPYLAJ
- Journal Information:
- Journal of Phycology; (United States), Vol. 28:3; Conference: 1992 meeting of the Phycological Society of America, Honolulu, HI (United States), 9-13 Aug 1992; ISSN 0022-3646
- Country of Publication:
- United States
- Language:
- English
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