.The sterile procedures described below should be followed.
1) Wipe all inner surfaces of the inoculating hood
and working surfaces with 70 percent ethanol.
2) Place all flasks that will be used in the hood, including
flasks to be transferred from (the transfer
flask) and flasks containing sterilized media that
will be inoculated under the culture transfer hood.
3) Irradiate flasks to be inoculated with an ultraviolet
lamp for at least 20 minutes. Be sure the hood
has a dark cover over the viewing glass (UV radiation
can be damaging to the eyes).
4) Switch off the UV lamp; ignite a small Bunsen
burner; remove caps from one transfer and one
new flask; and flame the neck of each flask by
slowly rotating the neck through the flame.
5) Tilt the neck of the transfer flask toward the new
flask. In one motion, remove both stoppers and
pour an inoculum into the new flask. Transfer
approximately 50 mL for diatom species and 100
mL for flagellates. Avoid touching the necks of the
two flasks. Never touch the portion of the stopper
that is inserted into the flask. Once the inoculum
is added, replace the stopper in the transfer flask.
Slowly flame the neck of the new flask before
replacing its stopper.
6) Replace the cap over the neck of the new flask and
use a waterproof marker pen to label the new flask
with the algal species inoculated and the date of
transfer.
7) After all inoculations are completed, turn off the
burner and transfer all new flasks to an algal incubator
or a well-lit area in the algal culture facility.
The remaining inoculum in the transfer flasks can
be used to inoculate larger cultures such as 4-L
flasks or carboys.