Partitioning soil respiration into three components (root respiration, rhizomicrobial respiration, and basal respiration) is called "three-source partitioning of soil respiration". These components are three different biological processes, and their responses to environment change are different. Therefore, it is important and significant to partition soil respiration into autotrophic and heterotrophic respiration to evaluate terrestrial ecosystem carbon balance quantitatively. Although there has been a general understanding of autotrophic and heterotrophic respiration affected by environment factors, the mechanisms underlying their role in the rhizosphere and their ecological significance are still not fully comprehended. To determine the mechanisms of the rhizosphere system response to environmental change, rhizosphere respiration should be partitioned into root respiration and rhizomicrobial respiration. In the present study, we discussed the significance, methods, and applications of three-source partitioning of soil respiration, and summarized the results under different conditions. Methods of three-source partitioning of soil respiration include physical excised method (such as the root excised method), isotopic tracer continuous labeling (such as ¹³C natural abundance), and isotopic tracer pulse labeling (such as ¹⁴C pulse labeling). The isotopic tracer pulse labeling method is more accurate than the other methods; however, it cannot be utilized in field studies and is expensive to undertake. The physical excised method is cheaper and requires no complex calculation; however, it greatly disturbs the soil and roots, which casts doubts on the relevance of the results. The isotopic tracer continuous labeling method has advantages for in situ measurements, accurate tracing, and causes almost no disturbance. However, this method can be utilized only under special conditions (such as when the plant organics are from C3 and the basal soil organic matter are from C4) and further study is required to improve its applicability. Therefore, it is difficult to partition soil respiration into the three components under field conditions. The excised roots and trenching method and ¹³C natural abundance method can be utilized in field studies, especially in forest ecosystems. The results showed that the proportion of root respiration and the proportion of rhizomicrobial respiration to rhizosphere respiration was approximately 45% and 55%, respectively, under laboratory conditions, and approximately 60% and 40%, respectively, under field conditions. Root respiration, rhizomicrobial respiration, and basal respiration varied with climate, depth, soil nutrition, and other environmental factors. Decomposition of rhizoorganisms, which causes rhizomicrobial respiration, has the potential to influence greatly basal respiration. The variation of the three components reflect the turnover rate of soil carbon, and affect the acquisition of competition and symbiosis by plants and microorganisms, thus maintaining nutrient balance among the various components of an ecosystem. The conventional three-source partitioning methods might produce great uncertainty by ignoring the root-microbial system processes in the soil. These processes include the adaptation of soil microbes, rhizosphere priming effects, nutrient partitioning, etc. Rhizomicrobial respiration is an important part of soil respiration in plantations and cannot be ignored. Three-source partitioning of soil respiration is a useful approach to quantitatively evaluate forest underground CO₂ flux as the global climate changes. The outlooks of the present study on the three-source partitioning of soil respiration were also discussed and clarified.