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This article considers Lyly’s formative place in the development of roles for female characters in early modern drama. It discusses the centrality and dominance of such characters in Lylian plays and explores the relationship between this playwright and his theatre companies. The first section charts the exploration of the possibilities of female identity within the contingencies of patriarchal power in Lyly’s early work, whilst the second section traces the dramaturgical investigation of female subjectivity in The Woman in the Moon. The article then considers Lyly’s own exclusion from the literary canon as itself an example of misogyny, and demonstrates the much more positive relationship with female characters in his work.