Ice box cooling a conical flask with a funnel and filter cloth ready to filter the pea leaves homogenate and collect the chloroplasts in the flask.
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After filtering, the contents of the conical flask is centrifuged to pellet both the intact and the broken chloroplasts.
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The pellet of intact and broken chloroplasts is resuspended and placed gently onto Percoll pads.
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Centrifugation causes separation by the Percoll and we see a pellet of intact chloroplasts and a less-dense interface of broken chloroplasts.
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Intact thylakoids remain after lysis of the intact chloroplasts and we wash with repeated changes of the buffer.
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Water bath set at 26 degrees centigrade and illuminated from below to energize the thylakoid membranes.
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Reaction mixtures to see if recombinant protein with a particular bacterial Tat signal will be targeted to the pea thylakoid Tat pathway.
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