OBJECTIVE: To establish optimal amplification conditions with the application of 0.2 mL test tube in single cell separation and inspection. METHODS: Oral epithelium cell suspension was prepared. Five or ten cells were collected with 0.2 mL test tube. Then DNA were amplified with Identifiler Plus kit in three different conditions in which the proteinase K addition, gold enzyme concentration in PCR reaction, and PCR reaction cycles were adjusted separately. Finally the detection rate, allelic dropout rate and artificial alleles were compared among the groups. RESULTS: In these 3 different conditions: addition of proteinase K, addition of 0.4 microL gold enzyme in PCR reaction, and use of 32 cycles, the detection rate was higher and allelic dropout rate was lower than the other conditions. CONCLUSION In single cell separation and inspection, the usage of 0.2 mL test tube could be an effective supplement to chip-low volume amplification.
Alleles ; Cell Separation/methods* ; DNA/genetics* ; DNA Fingerprinting/methods* ; Endopeptidase K/administration & dosage* ; Enzymes/administration & dosage* ; Epithelial Cells/cytology* ; Feasibility Studies ; Forensic Genetics ; Genotype ; Humans ; Mouth Mucosa/cytology* ; Nucleic Acid Amplification Techniques ; Polymerase Chain Reaction/methods* ; Sensitivity and Specificity