The sea anemones that were collected for the "Clone Specific Segregation in
the Sea Anemone Anthopleura elegantissima" experiment were collected by

Lisbeth Francis in Pacific Grove, California (Biological Bulletin 1973, 144;

64-72). The topic of Francisís report is the particularity of the constant
anemone-free areas dividing contiguous accumulations of these anemones and the
connection of these areas to the dispersion and manner of these anemones. In her
report Francis describes how she did her experiment and the result of each step.

Francis also includes a discussion section where she discusses advantages versus
disadvantages of segregated aggregations and organisms that are similar to these
sea anemones. Francis first explains the materials and methods. One of the first
steps in this section is collecting the anemones. Slowly sliding a spatula under
the sea anemones, Francis dislodged them from the immense rocks to which they
were attached. At the laboratory they were kept in glass bowls containing water
from the sea and were fed periodically, exclusive of experimentation time. In
case of any impairments from the collection process, the anemones were kept in
these bowls for a few weeks before any of the experiments started. Only the most
healthy anemones were used in the experiment. To free the anemones, Francis hit
the bowl against a solid surface. To determine the sex of the anemones, they
were severed and inspected for sex organs. When their sex organs are fully grown
the femaleís are brownish-pink and they maleís are yellowish-white. The
anemones that contain one or more sex organs including oocytes or spermatocytes
were recorded as having developed sex organs. The anemones were then placed in a
drying oven for approximately 18 to 24 hours, so they could be dried to a
constant weight. The anemones living in clusters isolated from other clusters
were inspected to ascertain whether or not the anemones from each cluster were
different. The anemones living in the same cluster, Francis noticed, had
identical color patterns. There were other color patterns observed, but they
always occurred when the cluster was separated by an anemone-free area. In each
of the aggregations observed, Francis noticed that the sex of the anemones was
the same. There were either all males living together or all females. None of
the aggregations were integrated. Francisís conclusion from this is that since
they "reproduce asexually by longitudinal fission" (403), each cluster is a
clone and the anemone-free areas divide contiguous clones. From studying how
size is related to sexual maturity in sea anemones, Francis drew another
conclusion. She states that the more the anemones weigh, the more likely they
are to be sexually developed. Francisís next experiment was to try to figure
out if the anemones could place themselves into segregated groups. She collected
anemones from two clones living beside each other and attached them to a plastic
ball with a lead weight inside an aquarium. They were crammed together in five
horizontal lines with four animals in each line. The anemones were arranged so
that they were all mixed together heterogeneously instead of separated into
their two separate groups. Three days later the anemones looked as if they were
fairly attached to the ball so Francis removed the pins to let them move around
so she could observe what kind of groups they formed. Thirteen days later, four
of them had fallen off the ball and the other sixteen had organized themselves
back into their segregated groups. They moved around some more after thirteen
days, but there was not any connection between the two groups. Francis concluded
from this experiment that segregation between clones can be established by the
anemones themselves. The next experiment Francis conducted was to discover if
the anemones would create anemone-free zones in-between clonal groups if there
are no other species of animals and no waves. In this experiment the anemones
were collected from two different clones but they were not side-by-side. Francis
also kept them in the laboratory for an extended period of time (up to a year).

A baking dish was lined with foam plastic and on one side, one clone of anemones
was attached with insect pins and the other clone was attached to the other
side. Microscope slides were lined up and taped together to prevent any contact
between the two clones. Sea water was running into one side of the dish. The
flow of the water was changed periodically because the anemones are inclined to
move upstream and Francis didnít want the flow of the water to affect her
experiment. The insect pins and the microscope slides were removed. Pictures
were taken once a day