Appendix C. Determining blade area from length/width vs. area regressions.
The length and width of haploid and diploid blades were measured for Mazzaella flaccida (60 blades each) and Mazzaella laminarioides (40 blades each). We also calculated the area of each blade from digital images using NIH Image (v. 1.62). Thus, we obtained a relationship for length/width: area. The resulting equations that estimate area from the blade's linear dimensions (with a y-intercept of zero; P values from ANOVA for each relationship) are:
|Haploids: Area(mm2) = 0.71 × length(mm) × width(mm) (P < 0.0001),|
|Diploids: Area(mm2) = 0.67 × length(mm) × width(mm) (P < 0.0001).|
|Haploids: Area(mm2) = 0.484 × length(mm) × width(mm) (P < 0.0001),|
|Diploids: Area(mm2) = 0.561 × length(mm) × width(mm) (P < 0.0001).|
We also quantified the relationship between reproductive structure size and the number of spores/structure for each phase. Structure size was determined using NIH Image software and a Hitachi CCD digital color camera mounted on a dissecting microscope. The entire size range of reproductive structures on haploid and diploid individuals was sampled (45 structures for each phase of M. flaccida, 25 for each phase for M. laminarioides). Each structure was partially dissected from surrounding tissues in a small amount of sterile artificial seawater and was gently prodded until all spores had been released. The spores and seawater were then transferred to a microcentrifuge tube, the mixture was mixed via rapid pipetting, and then subsamples of the spore solution were removed and counted on a Neubauer chamber. Six subsamples were counted per structure. Differences in spore color (and subsequent germination success) have been noted for M. laminarioides (Santelices and Martinez 1997). We did not include pale yellow or colorless spores, because they were rare and were difficult to distinguish from surrounding tissues, and because such spore types have been found to have little or no germination success.
Haploid cystocarps ranged in size from 0.2 to 1.3mm2, and spore number closely correlated with cystocarp size (M. flaccida R = 0.82, P < 0.0001, M. laminarioides R =0.73, P < 0.0001). Diploid sori also varied in size (0.04 to 0.35 mm2) but yielded weaker relationships between sorus size and spore number (M. flaccida R = 0.49, P < 0.0001; M. laminarioides R = 0.36, P = 0.12). Because the correlation of diploid sorus size and the number of spores was not significant for M. laminarioides, we calculated the mean number of spores per sorus. The resulting regressions used to estimate spore number from reproductive structure size are:
|Haploids: no. spores = 18,199 × area(mm2) + 1725,|
|Diploids: no. spores = 29,203 × area(mm2) + 2150.|
|Haploids: no. spores = 16,528 × area (mm2) + 2198,|
|Diploids: no. spores = 6,926.|
Santelices, B., and E. A. Martinez. 1997. Hierarchical analysis of reproductive potential in Mazzaella laminarioides (Gigartinaceae, Rhodophyta). Phycologia 36:195207.