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During metaphase, chromosomes are aligned at the cell's equatorial plane, and each chromosome is composed of two sister chromatids, which are the result of DNA replication that occurred during the S phase of interphase. This duplication is evident as each chromosome appears as a distinct X-shaped structure. The sister chromatids are still attached at a region called the centromere, making it clear that chromosome duplication has taken place prior to metaphase. Thus, while the physical separation of chromatids occurs later during anaphase, the evidence of duplication is clearly observable during metaphase.

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