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Laboratory of Forensic Services DNA Manual Sacramento County District Attorney DNA: Contamination Control Introduction DNA laboratories require special laboratory configuration and sample handling protocols. Because of the sensitivity of PCR-based procedures, special precautions must be taken to avoid contamination of samples to be amplified. Descriptions of control measures for the following potential sources of sample contamination follow: Environmental DNA ve d environmental DNA cross-contamination PCR product carry-over. The following precautions should be taken to minimize the potential of contamination by environmental human genomic DNA. ch i Analysts should wear gloves at all times. Sample tubes should be closed when not in use. Dispersal of aerosols should be kept to a minimum. Crosscontamination The following precautions should be taken during extraction and set-up to prevent transfer of DNA from one sample to another. Ar Analysts must use a fresh pipette tip for each sample, open tubes carefully, and keep sample containers closed when not in use. Samples expected to have low concentration of DNA, such as single hairs, should be processed at a different time from those of higher concentration. Evidence and reference samples should be processed separately in time or space. Primary evidence collected from a victim and from a suspect should be processed separately in time or space. A pipettor and barrier tips should be used in setting up PCR reactions for aliquoting PCR pre-mix. Barrier tips should be used for PCR amplification set-up. Continued on next page Title: Approved by: Issue Date: DNA: Contamination Control Laboratory Director 04/2011 Revision No: Document Class: 2 LFS Policy/Procedure All printed copies are uncontrolled. This document printed on 7/22/2015 Page 1 of 3 Laboratory of Forensic Services DNA Manual Sacramento County District Attorney DNA: Contamination Control, Continued PCR product carry-over PCR product carry-over occurs when amplified DNA contaminates a sample that has not yet been amplified. Amplified PCR product must be contained to prevent it from ever coming in contact with samples before they are amplified. The DNA laboratory has four designated work areas. Evidence examination areas: These areas are used for the examination of evidence items and for the screening, identification, and collection of biological stains. DNA extraction areas: These areas are used for extraction and isolation of DNA, including sample digestion, organic extraction, concentration, and quantitation. Extraction of samples containing low DNA levels are separated in time from other DNA extractions. Additionally, the extraction of reference samples are separated in time or space from the extraction of evidence samples. Dedicated equipment and supplies located in this area are for extraction and quantitation only. PCR set-up area: This area is used to prepare extracted DNA for quantitation/amplification. Dedicated equipment and supplies located in this area are for PCR set up only. Amplification/typing room: This room is used only for those activities that involve the handling of amplified DNA. This includes quantitation (based on Real Time PCR methodology), capillary electrophoresis of amplified DNA, waste disposal of amplified DNA products, and storage of amplified DNA. Dedicated equipment and supplies located in this room are for use only with amplified DNA procedures. Ar ch i DNA laboratory work areas ve d To minimize the potential for carry-over contamination, the DNA Laboratory is organized so that the area in which amplified DNA is handled is physically isolated from the extraction and set up areas. See DNA laboratory work areas below. Continued on next page Title: Approved by: Issue Date: DNA: Contamination Control Laboratory Director 04/2011 Revision No: Document Class: 2 LFS Policy/Procedure All printed copies are uncontrolled. This document printed on 7/22/2015 Page 2 of 3 Laboratory of Forensic Services DNA Manual Sacramento County District Attorney DNA: Contamination Control, Continued The types of contamination described above are concerns during sample preparation and PCR set up. The contamination of amplified DNA with unamplified DNA does not pose a problem. However, ordinary precautions, such as changing pipette tips between samples, should be taken to prevent cross-contamination between samples of amplified DNA. Ar ch i ve d Additional precautions Title: Approved by: Issue Date: DNA: Contamination Control Laboratory Director 04/2011 Revision No: Document Class: 2 LFS Policy/Procedure All printed copies are uncontrolled. This document printed on 7/22/2015 Page 3 of 3