SOD2 T-9C polymorphism was typed using RFLP-PCR protocol. Briefly, a 20 μl reaction mixture containing 5 ng of template DNA, 0.75 units of GoTaq polymerase (Promega, Madison, USA), and a final concentration of 200 mM each deoxynucleotide and 1 reaction buffer, and 1 mM of forward (5'-ACC AGC AGG CAG CTG GCG CCG G-3') and reverse (5'-GCG TTG ATGThe balance between oxidants and antioxidants is shifted towards an oxidized state. In animals, oxidative stress increases with aging due to the high production of free radicals by aged mitochondria and decreased cellular antioxidant capacity. The mitochondrial magnesium superoxide dismutase (SOD2) is considered the first line of defense against reactive oxygen species [4]. The gene for SOD2 has a common T to C polymorphism, resulting in a valine to alanine change at position 16 of its mitochondrial targeting sequence (Ala16Val), which affects the structure of the protein [6]. It reduces its entrance into the mitochondria [29], leading to increased oxidative stress. The male-to-female ratio was 1.3:1 in the elderly group and 1:1 in the control group (P = 0.346). The number of relatives who exceeded 85 years old was higher in the elderly group compared to the young control group (70.3% versus 59.8%, respectively, P = 0.013). The higher number of relatives who exceeded 85 years old (>25%) in the elderly group indicates a genetic component to longevity in the Jordanian population. Table 1 shows the frequency of homozygous and heterozygous genotypes for SOD2 T-9C and MTHFR C677T SNPs in our sample of elderly and young controls. The genotype frequencies of the SOD2 T-9C SNP of elderly and control groups were not statistically different (Chi-square test, P = 0.576). Accordingly, the frequency of SNP -9 T to C was not significantly different between elderly and controls (Chi-square test, P = 0.355). Similar results were observed with the MTHFR C677T SNP (Chi-square test: for genotype frequencies, P = 0.944 and for allele frequencies, P = 0.727). The MTHFR gene also affects oxidative stress status in the human body. The gene codes for an enzyme that plays a key role in folate metabolism [30]. Nucleotide transition (C to T) at nucleotide 677 of MTHFR causes alanine to valine substitution in the N-terminal catalytic domain, leading to 30% and 65% reduction in activity for heterozygotes and homozygotes of the variant allele, respectively [31]. Reduced activity of MTHFR leads to high levels of blood homocysteine, which is rapidly auto-oxidized, leading to the production of cytotoxic reactive oxygen species and endothelial damage [32]. Several studies indicated that gender was a main variable in the genetics of longevity and suggested that men and women might have different genetic determinants of aging [19,21]. Table 2 shows the frequencies of SOD2 and MTHFR alleles and genotypes in elderly and control male subjects. Among the limitations of this study are the sample size and age of recruitment of elderly subjects (>85 years). One hundred and thirty subjects with a mean age of 90.01 years were included in the present study. The population of Jordan was 5.7 million in 2007, and only 4.1% of the total population were individuals of 65 years or older [26]. In addition, the mortality rate starts inclining exponentially at fifty years old, getting to a maximum level in the late seventies indicating that reaching above 85 years old is a rarity (Khoury et al., 1999). Therefore, oldest old people are considered exceptional individuals in Jordan. Moreover, the lack of elderly centers in Jordan makes it very hard to recruit elderly subjects that fit sampling criteria. Despite all these obstacles, the sample size of the current research falls within the range of longevity studies reviewed by Glatt et al., [23] and previous studies that asked the same question in other populations (see discussion above). Future studies with a bigger sample size might be more appropriate for this kind of research. The results of this study indicate that SOD -9T/C and MTHFR 677C/T are not important determinants of lifespan in the Jordanian population. Longevity is a complex trait, which likely results from a blessed combination of genetic and non-genetic factors [38]. It is possible that the excess of environmental factors exerts stronger influence on longevity than the genetic traits [39]. For example, in institutionalized or home-bound elderly, oxidative stress was reported to increase significantly [40,41], while in free-living elderly, it is not always elevated [42]. In addition, MTHFR C677T polymorphism effect on homocysteine level can be minimized by folate intake. Studies attempting to assess the overall genetic influence on variations in human lifespan indicated that approximately a quarter of the variation in adult lifespans could be attributed to genetic variation among individuals [43]. Thus, the strong influence of environmental factorsRavenscroft J, Malik S, Gill MS, Walker DW, Clayton Wallace DC, Malfroy B, Doctrow SR, Lithgow GJ: Extension of span with superoxide dismutase/catalase mimetics. Free Radic Biol Med. Zelko IN, Mariani TJ, Folz RJ: Superoxide dismutase family: a comparison of the Cu/Zn-SOD (SOD1), Mn-SOD (SOD2), and EC-SOD (SOD3) gene structures and expression. Free Radic Biol Med. Shimoda-Matsubayashi S, Matsumine H, Kobayashi T, Torii Y, Shimizu Y, Mizuno Y: Structural dimorphism in the mitochondrial targeting sequence in the human superoxide dismutase gene. A predictive evidence for formational change to influence mitochondrial and a study of allelic association in Parkinson's disease. Chem Biophys Res Commun. Martin RC, Li Y, Liu Q, Jensen NS, Barker DF, Doll MA, Hein Manganese Superoxide Dismutase V A Polymorphism in the Mitochondrial Targeting Sequence Associated with Reduced Enzymatic Activity in served Human Hepatocytes. DNA Cell Biol. Sutton A, Imbert A, Igoudjil A, Descatoire V, Cazanave S, Pessayre Degoul F: The manganese superoxide dismutase dimorphism