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1.  SLC6A3 and body mass index in the Prostate, Lung, Colorectal and Ovarian Cancer Screening Trial 
BMC Medical Genetics  2009;10:9.
To investigate the contribution of the dopamine transporter to dopaminergic reward-related behaviors and anthropometry, we evaluated associations between polymorphisms at the dopamine transporter gene(SLC6A3) and body mass index (BMI), among participants in the Prostate, Lung, Colorectal and Ovarian (PLCO) Cancer Screening Trial.
Four polymorphisms (rs6350, rs6413429, rs6347 and the 3' variable number of tandem repeat (3' VNTR) polymorphism) at the SLC6A3 gene were genotyped in 2,364 participants selected from the screening arm of PLCO randomly within strata of sex, age and smoking history. Height and weight at ages 20 and 50 years and baseline were assessed by questionnaire. BMI was calculated and categorized as underweight, normal, overweight and obese (<18.5, 18.5–24.9, 25.0–29.9, or ≥ 30 kg/m2, respectively). Odds ratios (ORs) and 95% confidence intervals (CIs) of SLC6A3 genotypes and haplotypes were computed using conditional logistic regression.
Compared with individuals having a normal BMI, obese individuals at the time of the baseline study questionnaire were less likely to possess the 3' VNTR variant allele with 9 copies of the repeated sequence in a dose-dependent model (** is referent; OR*9 = 0.80, OR99 = 0.47, ptrend = 0.005). Compared with individuals having a normal BMI at age 50, overweight individuals (A-C-G-* is referent; ORA-C-G-9 = 0.80, 95% CI 0.65–0.99, p = 0.04) and obese individuals (A-C-G-* is referent; ORA-C-G-9 = 0.70, 95% CI 0.49–0.99, p = 0.04) were less likely to possess the haplotype with the 3'variant allele (A-C-G-9).
Our results support a role of genetic variation at the dopamine transporter gene, SLC6A3, as a modifier of BMI.
PMCID: PMC2640369  PMID: 19183461
2.  cis sequence effects on gene expression 
BMC Genomics  2007;8:296.
Sequence and transcriptional variability within and between individuals are typically studied independently. The joint analysis of sequence and gene expression variation (genetical genomics) provides insight into the role of linked sequence variation in the regulation of gene expression. We investigated the role of sequence variation in cis on gene expression (cis sequence effects) in a group of genes commonly studied in cancer research in lymphoblastoid cell lines. We estimated the proportion of genes exhibiting cis sequence effects and the proportion of gene expression variation explained by cis sequence effects using three different analytical approaches, and compared our results to the literature.
We generated gene expression profiling data at N = 697 candidate genes from N = 30 lymphoblastoid cell lines for this study and used available candidate gene resequencing data at N = 552 candidate genes to identify N = 30 candidate genes with sufficient variance in both datasets for the investigation of cis sequence effects. We used two additive models and the haplotype phylogeny scanning approach of Templeton (Tree Scanning) to evaluate association between individual SNPs, all SNPs at a gene, and diplotypes, with log-transformed gene expression. SNPs and diplotypes at eight candidate genes exhibited statistically significant (p < 0.05) association with gene expression. Using the literature as a "gold standard" to compare 14 genes with data from both this study and the literature, we observed 80% and 85% concordance for genes exhibiting and not exhibiting significant cis sequence effects in our study, respectively.
Based on analysis of our results and the extant literature, one in four genes exhibits significant cis sequence effects, and for these genes, about 30% of gene expression variation is accounted for by cis sequence variation. Despite diverse experimental approaches, the presence or absence of significant cis sequence effects is largely supported by previously published studies.
PMCID: PMC2077339  PMID: 17727713
3.  Effects of DNA mass on multiple displacement whole genome amplification and genotyping performance 
BMC Biotechnology  2005;5:24.
Whole genome amplification (WGA) promises to eliminate practical molecular genetic analysis limitations associated with genomic DNA (gDNA) quantity. We evaluated the performance of multiple displacement amplification (MDA) WGA using gDNA extracted from lymphoblastoid cell lines (N = 27) with a range of starting gDNA input of 1–200 ng into the WGA reaction. Yield and composition analysis of whole genome amplified DNA (wgaDNA) was performed using three DNA quantification methods (OD, PicoGreen® and RT-PCR). Two panels of N = 15 STR (using the AmpFlSTR® Identifiler® panel) and N = 49 SNP (TaqMan®) genotyping assays were performed on each gDNA and wgaDNA sample in duplicate. gDNA and wgaDNA masses of 1, 4 and 20 ng were used in the SNP assays to evaluate the effects of DNA mass on SNP genotyping assay performance. A total of N = 6,880 STR and N = 56,448 SNP genotype attempts provided adequate power to detect differences in STR and SNP genotyping performance between gDNA and wgaDNA, and among wgaDNA produced from a range of gDNA templates inputs.
The proportion of double-stranded wgaDNA and human-specific PCR amplifiable wgaDNA increased with increased gDNA input into the WGA reaction. Increased amounts of gDNA input into the WGA reaction improved wgaDNA genotyping performance. Genotype completion or genotype concordance rates of wgaDNA produced from all gDNA input levels were observed to be reduced compared to gDNA, although the reduction was not always statistically significant. Reduced wgaDNA genotyping performance was primarily due to the increased variance of allelic amplification, resulting in loss of heterozygosity or increased undetermined genotypes. MDA WGA produces wgaDNA from no template control samples; such samples exhibited substantial false-positive genotyping rates.
The amount of gDNA input into the MDA WGA reaction is a critical determinant of genotyping performance of wgaDNA. At least 10 ng of lymphoblastoid gDNA input into MDA WGA is required to obtain wgaDNA TaqMan® SNP assay genotyping performance equivalent to that of gDNA. Over 100 ng of lymphoblastoid gDNA input into MDA WGA is required to obtain optimal STR genotyping performance using the AmpFlSTR® Identifiler® panel from wgaDNA equivalent to that of gDNA.
PMCID: PMC1249558  PMID: 16168060
4.  Performance of high-throughput DNA quantification methods 
BMC Biotechnology  2003;3:20.
The accuracy and precision of estimates of DNA concentration are critical factors for efficient use of DNA samples in high-throughput genotype and sequence analyses. We evaluated the performance of spectrophotometric (OD) DNA quantification, and compared it to two fluorometric quantification methods, the PicoGreen® assay (PG), and a novel real-time quantitative genomic PCR assay (QG) specific to a region at the human BRCA1 locus. Twenty-Two lymphoblastoid cell line DNA samples with an initial concentration of ~350 ng/uL were diluted to 20 ng/uL. DNA concentration was estimated by OD and further diluted to 5 ng/uL. The concentrations of multiple aliquots of the final dilution were measured by the OD, QG and PG methods. The effects of manual and robotic laboratory sample handling procedures on the estimates of DNA concentration were assessed using variance components analyses.
The OD method was the DNA quantification method most concordant with the reference sample among the three methods evaluated. A large fraction of the total variance for all three methods (36.0–95.7%) was explained by sample-to-sample variation, whereas the amount of variance attributable to sample handling was small (0.8–17.5%). Residual error (3.2–59.4%), corresponding to un-modelled factors, contributed a greater extent to the total variation than the sample handling procedures.
The application of a specific DNA quantification method to a particular molecular genetic laboratory protocol must take into account the accuracy and precision of the specific method, as well as the requirements of the experimental workflow with respect to sample volumes and throughput. While OD was the most concordant and precise DNA quantification method in this study, the information provided by the quantitative PCR assay regarding the suitability of DNA samples for PCR may be an essential factor for some protocols, despite the decreased concordance and precision of this method.
PMCID: PMC280658  PMID: 14583097

Results 1-4 (4)