Zygosity was initially determined by self-report questionnaire with standard questions regarding physical similarity and degree to which others could tell co-twins apart. If co-twins gave inconsistent answers, they were followed-up by telephone, and if inconsistency or uncertainty was still apparent, they were asked to send in photographs at various ages, from which a zygosity assignment was made by project staff. Zygosity assignment based on self-report and responses to standard informative questions has been shown to be approximately 97% accurate (Reed et al., 2005). Subsequently, 347 pairs were genotyped at nine independent DNA microsatellite polymorphisms plus the sex marker amelogenin using the ABI Profiler PlusT multiplex marker set (AmpFLSTRR Profiler PlusT, Applied Biosystems, Foster City, CA). The probability of dizygotic twins being concordant for both alleles at all of the polymorphic loci examined when using this kit, is reported to be less than 10−4 (i.e., resulting in 99.99% zygosity certainty) (Nyholt, 2006). Additionally, most pairs were genotyped, as part of ongoing linkage studies, at a minimum of 20 (maximum of 1369) microsatellite markers. Only 16 pairs had fewer than 20 markers
