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In fall 1775, the United States Navy was established to intercept incoming British ships carrying war supplies to the British troops in the colonies. To aid in this, the Second Continental Congress authorized the mustering of five companies of Marines to accompany the Navy on their first mission. The first Marines that enlisted were from Philadelphia and they carried drums painted yellow, depicting a coiled rattlesnake with thirteen rattles, and the motto "Don't Tread On Me." This is the first recorded mention of the future Gadsden flag's symbolism.
http://en.wikipedia.org/wiki/Gadsden_flag
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