Rustriningsih was born in Kedungampel village of Kebumen, Central Java on 3 July 1967 as the eighth of ten children.

After completing her basic education, she studied in General Soedirman University's faculty of social and political sciences.

When PDI encountered internal conflict and government intervention in 1996, Rustriningsih was a Megawati loyalist, forming sub-branches for the party and being arrested (though always later released) several times.

[1]: 79  She adopted a reformist and trustworthy image, and received an "Outstanding Women in Local Government" from UNESCAP, and was featured by CNN.

[1]: 84 During her second term as regent, Rustriningsih ran as Bibit Waluyo's running mate for the governorship of Central Java and won.

In the five-year tenure, she would attempt and fail to become the head of PDI-P's provincial board in Central Java, with her brother also losing in Kebumen's 2010 regency election.

[10] In the election itself, the PDI-P backed candidate and incumbent Ganjar Pranowo lost in Kebumen, and the defeat was attributed to Rustriningsih's influence in the regency.