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The three daughters of Charles V
On 1519, Charles V would marry Isabella of Portugal as he had seen that Beatrice, her younger sister had married Henry VIII to further his alliance with the English, however, the marriage would take sometime to produce a surviving child until a daughter named Maria of Spain in June 25, 1525 was born, followed by a son named Philip on May 21, 1527, followed by two daughters, Catherine(December 2, 1528) and Joanna(June 26, 1535).
Catherine is intended for Afonso of Portugal the first son of the King of Portugal to further the matrimonial link between the Portuguese and the Spanish in the long run and Maria is to marry the eldest son of the Holy Roman Emperor and Joanna is intended for a marriage treaty, those were the fates of the three daughters of Charles V.