The identification of biological fluids is an important task in forensic science, as distinguishing between blood, saliva, and semen can help interpret biological evidence collected from crime scenes. Although chemical, enzymatic, and immunological tests are used to identify these fluids, some methods may have limitations in sensitivity or specificity, particularly when samples are degraded or mixed. Therefore, molecular techniques based on RNA analysis, especially messenger RNA (mRNA) and microRNA (miRNA), have emerged as promising methods for differentiating biological fluids. These methods are based on the fact that different cells and tissues exhibit distinct patterns of gene expression. For example, blood cells contain markers associated with hemoglobin and red blood cell membrane components. Important markers investigated for blood identification include HBA and HBB, which are associated with alpha and beta hemoglobin chains, respectively, and GYPA, which encodes glycophorin A, a protein found on the surface of red blood cells. Analysis of RNA transcripts associated with these genes may help identify blood in a forensic sample. However, results must be interpreted according to the specificity of the selected marker and the possibility of other biological components being present. Saliva can be identified using markers associated with salivary gland cells and their secretions. These include HTN3, which is associated with histatin 3, and STATH, which is associated with statherin. Both are proteins found in saliva. Analysis of gene expression associated with these markers may help identify the origin of a biological sample. However, the sensitivity and specificity of individual saliva markers may vary depending on the person and sample conditions. Therefore, using a combination of markers is generally preferable to relying on a single marker. Regarding semen identification, PRM1 and PRM2 are important genes associated with protamines found in sperm cells, while SEMG1, which encodes semenogelin 1, is associated with seminal fluid. Analysis of these markers may help differentiate semen from other biological fluids. However, semen identification does not always depend on the presence of sperm cells, because some semen samples may contain no detectable spermatozoa. Therefore, appropriate markers should be selected according to the purpose of the examination. Reverse transcription polymerase chain reaction (RT-PCR) is used to convert RNA into complementary DNA (cDNA), which is then amplified to detect specific target sequences. Real-time reverse transcription PCR (RT-qPCR) can also be used to measure the relative expression of molecular markers. Multiplex PCR techniques allow several markers to be analyzed in a single test, potentially helping differentiate multiple biological fluids within the same sample. In addition to mRNA, microRNA molecules are being investigated because of their relative resistance to degradation under certain environmental conditions, making them promising targets for the analysis of old or degraded biological stains. In conclusion, RNA markers and RT-PCR techniques represent promising tools for identifying the origin of biological fluids in forensic investigations. Important markers include HBA, HBB, and GYPA for blood; HTN3 and STATH for saliva; and PRM1, PRM2, and SEMG1 for semen. However, these markers should not all be considered independently conclusive tests. Their forensic application requires validation of specificity, sensitivity, and stability, while accounting for possible mixtures of different biological fluids. Therefore, molecular results should be combined with validated forensic tests, while STR analysis should be performed separately when individual identification is required.