## Materials and Methods
### Genetic Polymorphisms  
Genetic polymorphisms of the three enzymes involved in different phases of the DNA repair process were investigated: _XRCC1_, that has a central role in the base excision repair system (BER), for which three different polymorphisms were studied; _XRCC3_, implicated in the repair of double-strand DNA breaks by homologous recombination (HRR), and _XPD_ that pertains to the nucleotides excision oxidative damage repair system. In addition, the polymorphisms of three enzymes that act on the detoxification of reactive intermediates that can be produced following cell-IR interactions were studied: _GSTT1_ and _GSTM1_ of the _GST_ family, and the protease BLHX, also involved in the inactivation of BLM.  
To determine genetic polymorphisms, genomic DNA was isolated from a second aliquot of peripheral blood samples collected in EDTA. In brief, DNA isolated from peripheral lymphocytes was purified twice by the addition of 500 \(\upmu\)L of 70% ethanol, centrifuged at +4 \({}^{\circ}\)C, dried at room temperature for 1 h, and resuspended in 100 \(\upmu\)L of TE buffer (10 mM Tris-HCl, pH 7.4; 1 mM EDTA, pH 8.0). The DNA samples were diluted, divided in 10 ng/\(\upmu\)L aliquots and conserved at \(-20\)\({}^{\circ}\)C for subsequent amplification by standard polymerase chain reaction (PCR)-based methods. PCR was performed on 10 ng of DNA using HotMaster Taq DNA polymerase (5 PRIME Inc., Gaithersburg, MD, USA) according to the manufacturer's instructions using the following protocol: initial denaturation for 5 min at 95 \({}^{\circ}\)C followed by 35 cycles lasting 30 s at 95 \({}^{\circ}\)C, 30 s at Tm \({}^{\circ}\)C, and 30 s at 65 \({}^{\circ}\)C, and a final extension at 65 \({}^{\circ}\)C for 5 min. Negative and positive controls were used in the PCR reaction for the different genotypes.  
Specific primer pairs were designed to selectively amplify _XRCC_ exons 6, 9, and 10, _XRCC3_ gene exon 7, _XPD_ gene exon 23, and BLMH gene exon 12. The primers sequences and amplification conditions are listed in Table 1. The amplified fragments were digested with the RFLP (restriction fragment length polymorphism) technique using specific Thermo Scientific FastDigest enzymes for each of the polymorphisms studied, according to the manufacturer's instructions (Thermo Scientific, Waltham, MA, USA). After enzymatic digestion, the samples were subjected to electrophoresis in 2% agarose gel (containing Ethidium Bromide) to allow separation of the bands if the DNA fragment under study had been digested, thus allowing the identification of the presence of polymorphisms.  
The following genes were investigated:  
1. Gene _XRCC1_: Determining the polymorphisms Arg194 Trp, Arg280 His and Arg399 Gln, localized in exons 6, 9, 10, respectively using the respective restriction enzymes PvuII, MspI and RasI [12; 19];
2. Gene _XRCC3_: Determining the polymorphism Thr241 Met, localized on exon 7, using the restriction enzyme NlaIII [12];
3. Gene _XPD_: Determining the polymorphism Lys751 Gln, localized on exon 23, using PstI as restriction enzyme [4];
4. Gene _BLMH_: Determining the nucleotide substitution (A1450G), using Munl as restriction enzyme.  
In regards to analysis of the genotypes _GSTM1_ and _GSTTI_, multiplex PCR with the GAPDH fragment was used as internal control, as described by [PERSON] et al. [30]. The products of multiplex PCR were visualized under UV light after electrophoresis in 2% agarose gel containing ethidium bromide.