Characterization of human lymphoid cell lines GM9947 and GM9948 as intra- and interlaboratory reference standards for DNA typing.
The incorporation of reference DNA is crucial to the validation of any DNA typing protocol. Currently, reference DNA standards are restricted to molecular size DNA ladders and/or tumor cell line DNA. Either of these, however, presents some limitations. We have rigorously characterized two Epstein-Barr virus (EBV)-immortalized human lymphoid cell lines--GM9947 (female) and GM9948 (male)--to determine their suitability as alternative in-line standards for three widely employed allele profiling strategies. Twenty-one highly polymorphic VNTR-based allelic systems (7 RFLPs, 2 AmpFLPs, and 12 STRs) distributed over 12 chromosomes were scrutinized along with 3 gender-based discriminatory systems. The genetic stability of each locus was confirmed over a period of 225 in vitro population doublings. Allele size estimates and degree of informativeness for each of the 21 VNTR systems were compiled. The reproducibility of allele scoring by traditional RFLP analyses, using both cell lines as reference standards, was also verified by an interlaboratory validation study involving 13 analysts from two geographically distinct forensic laboratories. Taken together, our data indicate that GM9947 and GM9948 genomic DNAs could be adopted as reliable reference standards for DNA typing.
RCMP Central Forensic Laboratory, Biology Research and Development Section, Ottawa, Ontario, Canada.
C J
CJ
Frégeau
R A
RA
Aubin
J C
JC
Elliott
S S
SS
Gill
R M
RM
Fourney
D001483:Base Sequence; D002461:Cell Line, Transformed; D004247:DNA / Q000592:standards*; D004591:Electrophoresis, Polyacrylamide Gel; D005069:Evaluation Studies as Topic; D005260:Female; D005554:Forensic Medicine / Q000592:standards; D005821:Genetic Techniques / Q000592:standards; D004854:Herpesvirus 4, Human; D006801:Humans; D008221:Lymphoid Tissue / Q000166:cytology*; D008297:Male; D018895:Microsatellite Repeats; D018598:Minisatellite Repeats; D008969:Molecular Sequence Data; D012150:Polymorphism, Restriction Fragment Length; D012015:Reference Standards; D015203:Reproducibility of Results; D012727:Sex Characteristics; D012732:Sex Determination Analysis / Q000379:methods
D004247:DNA