Microalgae are promising microorganisms used to produce value-added products or to develop sustainable approaches for environmental remediation. The ATP-binding cassette proteins (ABCs) of Chlamydomonas reinhardtii have been characterized as indispensable transporters for CO2 concentrating mechanism, lipid biosynthesis, and heavy metal sequestration. However, few microalgal ABC proteins have been studied compared with higher plants or non-photosynthetic microorganisms. This study performed a genome-wide, evolutionary, and transcriptomic survey of C. reinhardtii ABC proteins (CrABCs). A total of 75 CrABCs were identified and classed into eight ABC subfamilies, from ABCA to ABCI. We found that no whole or partial genome duplication events occurred in C. reinhardtii after the ancient endosymbiosis events, but gene duplications occurred in a small range of chromosomal regions, which forced ABC family expansion. Abundant light, abscisic acid, and jasmonic acid response cis-elements were mapped in the CrABC promoters, coinciding with the evolutionary history of hormone signaling in Chlorophyta. The expression survey under light/dark rhythms revealed a close bond of CrABCs with cell division and development. A broad study of CrABCs supported their expected roles in heavy metal detoxification, lipid metabolism, and environmental adaptation. Moreover, the evolutionary and expression survey predicted the functions of unknown CrABCs, which are elaborated in the text. Two half-size CrABCGs—CrABCG3 and CrABCG26—were described as plasma-membrane transporters that might participate in lipidic compound secretion. This study provides fundamental and exhaustive information about CrABCs, which are indispensable for the functional elucidation of ABC proteins in microalgae.