Abstract
The mitochondrial locus 16519T/C was used as a model for the evaluation of the benefits of ion-pair reversed-phase high-performance liquid chromatography on-line hyphenated to electrospray ionization time-of-flight mass spectrometry (ICEMS assay) for the determination of allelic frequencies of single nucleotide polymorphisms. This marker has gained interest in forensic science owing to its ability to increase the discrimination power of mitochondrial DNA testing as a consequence of its high variability across various populations. In a first set of experiments, artificial mitochondrial DNA mixtures prepared from all four theoretically possible 16519 alleles served as samples. Any allele occurring at a frequency of as low as 1–5% was unequivocally detectable irrespective of the kind of allelic mixture. Measured and expected allelic frequencies correlated well following correction of observed experimental bias, which was most probably attributable to differential PCR amplification and/or preferential ionization. For thirteen different T/C mixtures with C contents in the range 1.0–99.0%, an average error of 1.2% and a maximum error of 2.2% were observed. Furthermore, ICEMS was applied to the quantitative genotyping of eight selected individuals of which four were heteroplasmic with C contents in the range 1.9–34.1%. To check the reliability of these results, allelic proportions were additionally determined by a cloning assay. The results of the two assays correlated well (R 2=0.9971). In all cases, deviations were obtained that were smaller than 5.4%. The overall observed assay performance suggests that the described mass spectrometric technique represents one of the most powerful assays for the determination of allelic frequencies available today.
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Anderson S, Bankier AT, Barrell BG, de Bruijn MH, Coulson AR, Drouin J, Eperon IC, Nierlich DP, Roe BA, Sanger F, Schreier PH, Smith AJ, Staden R, Young IG (1981) Nature 290:457–465
Chinnery PF, Turnbull DM (1999) Lancet 354(Suppl 1):SI17–SI21
DiMauro S, Schon EA (2001) Am J Med Genet 106:18–26
Chinnery PF, Howell N, Lightowlers RN, Turnbull DM (1997) Brain 120:1713–1721
Holland MM, Parsons TJ (1999) Forensic Sci Rev 11:21–50
Budowle B, Allard MW, Wilson MR, Chakraborty R (2003) Annu Rev Genomics Hum Genet 4:119–141
Jobling MA, Gill P (2004) Nat Rev Genet 5:739–751
Jazin EE, Cavelier L, Eriksson I, Oreland L, Gyllensten U (1996) Proc Natl Acad Sci USA 93:12382–12387
Chinnery PF, Zwijnenburg PJ, Walker M, Howell N, Taylor RW, Lightowlers RN, Bindoff L, Turnbull DM (1999) Am J Med Genet 85:498–501
Huhne J, Pfeiffer H, Waterkamp K, Brinkmann K (1999) Int J Legal Med 112:172–175
Bendall KE, Macaulay VA, Baker JR, Sykes BC (1996) Am J Hum Genet 59:1276–1287
Bendall KE, Macaulay VA, Sykes BC (1997) Am J Hum Genet 61:1303–1308
Gocke CD, Benko FA, Rogan PK (1998) Hum Genet 102:182–186
Parsons TJ, Muniec DS, Sullivan K, Woodyatt N, Alliston–Greiner R, Wilson MR, Berry DL, Holland KA, Weedn VW, Gill P, Holland MM (1997) Nat Genet 15:363–368
Salas A, Lareu MV, Carracedo A (2001) Int J Legal Med 114:186–190
Ivanov PL, Wadhams MJ, Roby RK, Holland MM, Weedn VW, Parsons TJ (1996) Nat Genet 12:417–420
Lutz S, Weisser HJ, Heizmann J, Pollak S (2000) Int J Legal Med 113:155–161
Brandstatter A, Niederstatter H, Parson W (2004) Int J Legal Med 118:47–54
Schollen E, Vandenberk P, Cassiman JJ, Matthijs G (1997) Clin Chem 43:18–23
Liang MH, Wong LJ (1998) Am J Med Genet 77:395–400
Calloway CD, Reynolds RL, Herrin GL, Anderson WW (2000) Am J Hum Genet 66:1384–1397
Morley JM, Bark JE, Evans CE, Perry JG, Hewitt CA, Tully G (1999) Int J Legal Med 112:241–248
Lagerström–Fermér M, Olsson C, Forsgren L, Syvänen A (2001) Am J Hum Genet 68:1299–1301
Sternberg D, Danan C, Lombes A, Laforet P, Girodon E, Goossens M, Amselem S (1998) Hum Mol Genet 7:33–42
Tully LA, Parsons TJ, Steighner RJ, Holland MM, Marino MA, Prenger VL (2000) Am J Hum Genet 67:432–443
Boles RG, Chaudhari D, Soderkvist J, Podberezin M, Ito M (2003) Clin Chem 49:198–200
Szuhai K, Ouweland J, Dirks R, Lemaitre M, Truffert J, Janssen G, Tanke H, Holme E, Maassen J, Raap A (2001) Nucleic Acids Res 29:E13
van den Bosch BJ, de Coo RF, Scholte HR, Nijland JG, van Den Bogaard R, de Visser M, de Die–Smulders CE, Smeets HJ (2000) Nucleic Acids Res 28:E89
Sham P, Bader JS, Craig I, O’Donovan M, Owen M (2002) Nature Rev Gent 3:862–871
Kwok PY (2000) Pharmacogenomics 1:231–235
Shifman S, Pisante–Shalom A, Yakir B, Darvasi A (2002) Mol Cell Probes 16:429–434
Oberacher H, Huber CG, Oefner PJ (2003) Hum Mutat 21:86–95
Whitehouse CM, Dreyer RN, Yamashita M, Fenn JB (1985) Anal Chem 57:675–679
Karas M, Hillenkamp F (1988) Anal Chem 60:2299–2301
Muddiman DC, Null AP, Hannis JC (1999) Rapid Commun Mass Spectrom 13:1201–1204
Oberacher H, Oefner PJ, Parson W, Huber CG (2001) Angew Chem Int Ed 40:3828–3830
Walcher W, Oberacher H, Troiani S, Hölzl G, Oefner PJ, Zolla L, Huber CG (2002) J Chromatogr B 782:111–125
Fei Z, Ono T, Smith LM (1998) Nucleic Acids Res 26:2827–2828
Shahgholi M, Garcia BA, Chiu NH, Heaney PJ, Tang K (2001) Nucleic Acids Res 29:E91
Tost J, Gut IG (2002) Mass Spectrom Rev 21:388–418
Gut IG (2004) Hum Mutat 23:437–441
Buetow KH, Edmonson M, MacDonald R, Clifford R, Yip P, Kelley J, Little DP, Strausberg R, Koester H, Cantor CR, Braun A (2001) Proc Natl Acad Sci USA 98:581–584
Jurinke C, van den Boom D, Cantor CR, Koster H (2002) Adv Biochem Eng Biotechnol 77:57–74
Krahmer MT, Walters JJ, Fox KF, Fox A, Creek KE, Pirisi L, Wunschel DS, Smith RD, Tabb DL, Yates JR (2000) Anal Chem 72:4033–4040
Null AP, Hannis JC, Muddiman DC (2001) Anal Chem 73:4514–4521
Hawkridge AM, Zhou L, Lee ML, Muddiman DC (2004) Anal Chem 76:4118–4122
Van Ert MN, Hofstadler SA, Jiang Y, Busch JD, Wagner DM, Drader JJ, Ecker DJ, Hannis JC, Huynh LY, Schupp JM, Simonson TS, Keim P (2004) BioTechniques 37:642–648
Hofstadler SA, Sannes–Lowery KA, Hannis JC (2005) Mass Spectrom Rev 24:265–285
Null AP, Nepomuceno AI, Muddiman DC (2003) Anal Chem 75:1331–1339
Oberacher H, Parson W, Holzl G, Oefner PJ, Huber CG (2004) J Am Soc Mass Spectrom 15:1897–1906
Hall TA, Budowle B, Jiang Y, Blyn L, Eshoo M, Sannes–Lowery KA, Sampath R, Drader JJ, Hannis JC, Harrell P, Samant V, White N, Ecker DJ, Hofstadler SA (2005) Anal Biochem 344:53–69
Huber CG, Oberacher H (2001) Mass Spectrom Rev 20:310–343
Oberacher H, Huber CG (2003) Anal Bioanal Chem 376:292–294
Andreasson H, Gyllensten U, Allen M (2002) BioTechniques 33:402–4, 407–11
Oberacher H, Niederstätter H, Casetta B, Parson W (2005) Anal Chem 77:4999–5008
Premstaller A, Oberacher H, Huber CG (2000) Anal Chem 72:4386–4393
Huber CG, Buchmeiser MR (1998) Anal Chem 70:5288–5295
Lutz S, Weisser HJ, Heizmann J, Pollak S (1998) Int J Legal Med 111:67–77
Coble MD, Just RS, O’Callaghan JE, Letmanyi IH, Peterson CT, Irwin JA, Parsons TJ (2004) Int J Legal Med 118:137–146
Berger B, Hölzl G, Oberacher H, Niederstätter H, Huber CG, Parson W (2002) J Chromatogr B 782:89–97
Oberacher H, Niederstatter H, Parson W (2005) J Mass Spectrom 40:932–945
Barnard R, Futo V, Pecheniuk N, Slattery M, Walsh T (1998) BioTechniques 25:684–691
Hoogendoorn B, Norton N, Kirov G, Williams N, Hamshere ML, Spurlock G, Austin J, Stephens MK, Buckland PR, Owen MJ, O’Donovan MC (2000) Hum Genet 107:488–493
Wasson J, Skolnick G, Love–Gregory L, Permutt MA (2002) BioTechniques 32:1144–1150
Lutz–Bonengel S, Sanger T, Pollak S, Szibor R (2004) Int J Legal Med 118:274–281
Bini C, Pappalardo G (2005) Forensic Sci Int 152:35–38
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The authors wish to thank Applied Biosystems for the generous provision of the mass spectrometer and the associated equipment.
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Oberacher, H., Niederstätter, H., Huber, C.G. et al. Accurate determination of allelic frequencies in mitochondrial DNA mixtures by electrospray ionization time-of-flight mass spectrometry. Anal Bioanal Chem 384, 1155–1163 (2006). https://doi.org/10.1007/s00216-005-0269-1
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DOI: https://doi.org/10.1007/s00216-005-0269-1