As Wimbledon champion, Serena is one of the favourites for Singles gold, and the fourth seed moved through to the last 16 with a 6-2 6-3 win over Urszula Radwanska.
Radwanska is the sister of world number two Agnieszka, who was beaten by Serena in the final of Wimbledon three weeks ago, but the Pole could not take revenge.
By contrast, Venus has struggled for form and good health over the past year as she battles Sjogren's Syndrome and had pushed herself through matches just to qualify for London 2012.
The Games are very important to Venus, who already has three gold medals to her name, two in Doubles with Serena and one in Singles from Sydney in 2000.
Venus said: 'It was really great. Grass is my surface and I felt great out there. It's never easy to draw a top seed. I'm unseeded here and now I feel like I'm the number nine seed, and hopefully I'll beat another seed and become their number.
'I fought so much to be here. I know there's a lot of people happy to be here, but I'm not only happy to be here, I want to do something about it too and play well for my country.'
Serena is looking for her first Singles gold medal and, were she to achieve it, she would be only the second woman after Steffi Graf to have won all four grand slam titles and an Olympic gold in her career.
Meanwhile, the sisters teamed up for their first-round Doubles match against Romanians Sorana Cirstea and Simona Halep, which they won 6-3 6-2.
World number one Victoria Azarenka opened her Singles campaign with a 6-1 3-6 6-1 win against Romania's Irina-Camelia Begu.
Two former world number ones will meet in the third round, with Serbia's Ana Ivanovic facing Kim Clijsters of Belgium.
Ivanovic, seeded 11th, beat Elena Baltacha of Great Britain 6-4 7-6 (7/5), while Clijsters saw off Spain's Carla Suarez Navarro 6-3 6-3.
Last year's Wimbledon champion Petra Kvitova, the sixth seed, dropped a set for the second consecutive round but came through 7-5 2-6 6-1 against Peng Shuai, and she next meets Italy's Flavia Pennetta.
Eighth seed Caroline Wozniacki was also pushed before seeing off Belgian Yanina Wickmayer 6-4 3-6 6-3 to set up a meeting with Daniela Hantuchova, while there were first-round wins for Wimbledon semi-finalist Angelique Kerber, Sabine Lisicki and Maria Kirilenko.