Mary Singleton was born in 1936 in Fort Lewis, Washington. She received her B.S. (1958) in Chemistry from Wheaton College (Wheaton, Illinois), graduating summa cum laude. She received her M.S. (1960) in Chemistry at the University of California, Berkeley. From 1960-1962 she worked with Melvin Calvin, who won the Nobel Prize in Chemistry in 1961, at Lawrence Berkeley Laboratory at the University of California, Berkeley. In 1962 she left Lawrence Berkeley Laboratory and for the next twelve years she raised her children and moved to Europe, Wisconsin and California as her husband's job required.
Singleton began working again in 1974 at Lawrence Livermore National Laboratory (LLNL) in California, where she worked for twenty-two years. Most of her career was in research, including tritium-getter materials, oil shale processing, and growth of nonlinear optical crystals for the LLNL laser project. When she retired at the end of 1996 she was deputy plutonium facility manager. Soon after retirement, she enrolled as a student at the University of California, Berkeley in the Division of the History of Science and Technology and attended Oxford University during the summer of 2000. Her studies focused on the history of women in science, including the Manhattan Project and Dorothy Hodgkin's students.
In 1998 Singleton was among five other female employees of the Lawrence Livermore National Laboratory who filed a lawsuit against the Lab on December 23, 1998. They charged that the Lab paid and promoted female lab employees, in a variety of job categories, less than their male counterparts with comparable education and experience. The situation was facilitated by a system of annual salary adjustments based on a subjective method which ranked an employee based on their relative "value" to other employees, called the "Relative Value Rank." The settlement in November of 2003 required the lab to make significant reforms and benefited over 3000 female employees who worked at Lawrence Livermore National Laboratory at the time. Singleton had been removed as a plaintiff in the original case since when she retired she had signed an agreement to surrender her right to sue the University of California. She negotiated a separate agreement.
Singleton has been an active member in the American Chemical Society (ACS), serving on the Women Chemists Committee of the ACS and as chair of the California Section.