Virginia Galilei, the first of Galileo's younger sisters, was born in Pisa in 1573. No news survives about her childhood, except in relation to the family's ramblings between Pisa and Florence. When hardly out of her adolescence she married Benedetto Landucci (1569-?), probably in 1591, the year Vincenzo Galilei (c.1520-1591) died. So the task of paying her dowery fell squarely on the shoulders of the firstborn Galileo. Despite the fact that Galileo's new post at the University of Padua in 1592 had meant a considerable rise in his income, he was constantly behind in his dowery payments, to the point that his mother wrote warning him not to return to Tuscany without the money, as his brother-in-law had threatened to have him imprisoned. His desperate search for money to pay off the debts from his sisters Virginia and Livia's marriages was a constant subject in the correspondence between Galielo and his family.

He was on good terms with his sister and in the end, owing to his growing fame and prestige, even Benedetto Landucci made peace with him. It was also thanks to Galileo's intercession with Cristina di Lorena (1565-1636) that in 1609 Landucci was hired as an assay weigher. The job, which consisted of weighing silver when it was sold, earned him an income of 60 florins, and if it had not been for Galileo's recommendation probably someone else would have been chosen.

Virginia had four children: two boys, Vincenzo and Luca, and two girls, Isabella and Virginia, who became nuns, taking the names Sister Arcangela and Sister Chiara. The latter lived at the convent of San Matteo in Arcetri with Galileo's daughters.