The use of microbes to clean up polluted environments, bioremediation, is a rapidly changing and expanding area of environmental biotechnology. Although bioremediation is a promising approach to improve environmental conditions, our limited understanding of biological contribution to the effect of bioremediation and its impact on the ecosystem has been an obstacle to make the technology more reliable and safer. Providing fundamental data to resolve these issues, i.e., the behavior of the target bacteria directly related to the degradation of contaminants and the changes in microbial communities during bioremediation, has been a challenge for microbiologists since many environmental bacteria cannot yet be cultivated by conventional laboratory techniques. The application of culture-independent molecular biological techniques offers new opportunities to better understand the dynamics of microbial communities. Fluorescence in situ hybridization (FISH), in situ PCR, and quantitative PCR are expected to be powerful tools for bioremediation to detect and enumerate the target bacteria that are directly related to the degradation of contaminants. Nucleic acid based molecular techniques for fingerprinting the 16S ribosomal DNA (rDNA) of bacterial cells, i.e., denaturing gradient gel electrophoresis (DGGE) and terminal restriction fragment length polymorphism (T-RFLP), enable us to monitor the changes in bacterial community in detail. Such advanced molecular microbiological techniques will provide new insights into bioremediation in terms of process optimization, validation, and the impact on the ecosystem, which are indispensable data to make the technology reliable and safe.