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<methtype>Lab</methtype>
<methdesc>Golden king crab for this experiment were caught in commercial pots along
the Aleutian Islands, Alaska, in the falls of 2005 and 2006 and transported to the
ASFC Kodiak Laboratory via air cargo. Crab were held in 2,000 l tanks with flow-through
seawater chilled to 3-4 degrees Celsius, and fed chopped frozen fish and squid to
excess. Tanks were covered with opaque sheets of foam to both provide insulation
and to keep the females in mostly-dark conditions. Six females, 4 captured in 2005
and 2 in 2006, were used in this study. Females were either late-stage ovigerous,
as evinced by eyed embryos, or hatched out, as evinced by empty egg cases when collected
and were between 120 and 141 mm in carapace length (CL). Six mature males, CL 128-132
mm, were present with the females in a pre-mating holding tank. After the females
molted, they mated and extruded a new clutch of eggs. At this point they were placed
into another tank where all the post-mating females were held together with no males.</methdesc>
</method>
<method>
<methtype>Lab</methtype>
<methdesc>Embryos were collected regularly throughout development from a random location
within the clutch. Time between samples varied with the developmental stage and averaged
once every 9 days or a mean of 48 times (range 42-54) throughout development for each
female. At sampling, embryos were examined and photographed under a stereo microscope.
Uneyed embryos were stained for 5 min in Bouin's solution to enable staging. The
median stage of development (see results for descriptions) was determined for each
female on each sampling day. In addition, up to 10 unstained embryos were photographed
at 90° to the sagittal plane under a stereo microscope for image analysis. The
images were calibrated each day with a micrometer because the scope had an adjustable
zoom and this ensured accurate measurements. Measurements were made using Image-Pro
Plus V. 7.0 (Media Cybernetics, Inc. , Rockville, MD). In this data, the term egg;
refers to the entire embryo (i.e. the entire contents of the fertilized egg) and the
term embryo; refers to the differentiated part of the egg as distinct from the yolk,
and yolk; refers to the undifferentiated deutoplasm. For each egg, the area and the
mean, minimum, and maximum diameter of the egg was measured. The mean, minimum, and
maximum diameters were determined from 180 measurements of the diameter at 2 degree
intervals around the entire egg. When the embryo became visible, the yolk area was
also measured by tracing the yolk in Image-Pro and the percent area yolk (yolk area
x 100/egg area; prior to the embryo becoming the yolk area = egg area) and the embryo
area (egg area minus the yolk area) were calculated. Finally, when the eyes became
visible, the eye area and the mean, minimum, and maximum diameter of the eyes were
measured using the same techniques as above.</methdesc>
</method>
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