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She and Cosimo enjoyed a contented marriage. Together they had eight children. Cosimo II died in 1621, leaving their ten-year-old son Ferdinando as grand duke. Maria Maddalena and her mother-in-law, Christina of Lorraine, acted as regents until the boy came of age. Their collective regency is known as the Turtici. Maria Maddalena's temperament was analogous to Christina's. Together, they aligned Tuscany with the Papacy; re-doubled the Tuscan clergy; and allowed the trial of Galileo Galilei to occur. Upon the death of the last Duke of Urbino, instead of claiming the duchy for Ferdinando, who was married to the Duke's granddaughter, and heiress, Vittoria della Rovere, they permitted it to be annexed by Pope Urban VIII. In 1626, they banned any Tuscan subject from being educated outside the Grand Duchy, a law later resurrected by Maria Maddalena's grandson, Cosimo III.Harold Acton ascribes the decline of Tuscany to their regency. The Dowager Grand Duchesses sent Ferdinando on a tour of Europe in 1627.
The Grand Duchess died aged 42 after a visit to her brother Leopold in Innsbruck on the way back to Passau, Germany. Her son had been in power for a year.