Visual detection of lead(II) using a label-free DNA-based sensor and its immobilization within a monolithic hydrogel

Lead is highly toxic and its detection has attracted a lot of research interests. In recent years, DNA has been used for Pb2+ recognition and many fluorescent sensors with low to sub-nM detection limits have been reported. These figures of merit were typically measured using a spectrophotometer that can detect nM DNA with a high signal-to-noise ratio. For visual detection, however, μM DNA or dye was required, making it difficult to detect low nM targets. We recently achieved a visual sensitivity of 10 nM Hg2+ by immobilizing a DNA probe in a hydrogel. This was made possible because the gel was able to actively adsorb Hg2+. In this work, we aim to test whether this method can be extended to the detection of Pb2+. First, a new Pb2+ sensor was designed based on a guanine-rich DNA and DNA binding dyes such as thiazole orange and SYBR Green I. The free DNA showed a detection limit of 8 nM Pb2+ using 40 nM DNA. For visual detection in solution with 1 μM of the DNA probe, however, ∼300 nM Pb2+ was required. After immobilization in a monolithic polyacrylamide hydrogel, even 20 nM Pb2+ could be visually detected with a sample volume of 50 mL. Therefore, sensitive detection without signal amplification was achieved. Finally, we demonstrated simultaneous detection of both Hg2+ and Pb2+ in the same water sample with shape encoded hydrogel sensors.University of Waterloo || Canada Foundation for Innovation || Natural Sciences and Engineering Research Council || Ontario Ministry of Research and Innovation |

Metal-Induced Specific and Nonspecific Oligonucleotide Folding Studied by FRET and Related Biophysical and Bioanalytical Implications

This is the peer reviewed version of the following article: Kiy, M. M., Jacobi, Z. E., & Liu, J. (2012). Metal-Induced Specific and Nonspecific Oligonucleotide Folding Studied by FRET and Related Biophysical and Bioanalytical Implications. Chemistry - A European Journal, 18(4), 1202–1208, which has been published in final form at http://dx.doi.org/10.1002/chem.201102515 This article may be used for non-commercial purposes in accordance with Wiley Terms and Conditions for Self-Archiving.Metal induced nucleic acid folding has been extensively studied with ribozymes, DNAzymes, tRNA and riboswitches. These RNA/DNA molecules usually have a high content of double-stranded regions to support a rigid scaffold. On the other hand, such rigid structural features are not available for many in vitro selected or rationally designed DNA aptamers; they adopt flexible random coil structures in the absence of target molecules. Upon target binding, these aptamers adaptively fold into a compact structure with a reduced end-to-end distance, making fluorescence resonance energy transfer (FRET) a popular signaling mechanism. However, nonspecific folding induced by mono- or divalent metal ions can also reduce the end-to-end distance and thus lead to false positive results. In this study we used a FRET pair labeled HgII binding DNA and monitored metal-induced folding in the presence of various cations. While nonspecific electrostatically mediated folding can be very significant, at each tested salt condition, HgII induced folding was still observed with a similar sensitivity. We also studied the biophysical meaning of the acceptor/donor fluorescence ratio that allowed us to explain the experimental observations. Potential solutions for this ionic strength problem have been discussed. For example, probes designed to signal the formation of double-stranded DNA showed a lower dependency on ionic strength.University of Waterloo || Canadian Foundation for Innovation Ontario ministry of Research and Innovation || Natural Sciences and Engineering Research Council |

Heterogeneity of Breast Cancer Associations with Five Susceptibility Loci by Clinical and Pathological Characteristics

A three-stage genome-wide association study recently identified single nucleotide polymorphisms (SNPs) in five loci (fibroblast growth receptor 2 (FGFR2), trinucleotide repeat containing 9 (TNRC9), mitogen-activated protein kinase 3 K1 (MAP3K1), 8q24, and lymphocyte-specific protein 1 (LSP1)) associated with breast cancer risk. We investigated whether the associations between these SNPs and breast cancer risk varied by clinically important tumor characteristics in up to 23,039 invasive breast cancer cases and 26,273 controls from 20 studies. We also evaluated their influence on overall survival in 13,527 cases from 13 studies. All participants were of European or Asian origin. rs2981582 in FGFR2 was more strongly related to ER-positive (per-allele OR (95%CI) = 1.31 (1.27–1.36)) than ER-negative (1.08 (1.03–1.14)) disease (P for heterogeneity = 10−13). This SNP was also more strongly related to PR-positive, low grade and node positive tumors (P = 10−5, 10−8, 0.013, respectively). The association for rs13281615 in 8q24 was stronger for ER-positive, PR-positive, and low grade tumors (P = 0.001, 0.011 and 10−4, respectively). The differences in the associations between SNPs in FGFR2 and 8q24 and risk by ER and grade remained significant after permutation adjustment for multiple comparisons and after adjustment for other tumor characteristics. Three SNPs (rs2981582, rs3803662, and rs889312) showed weak but significant associations with ER-negative disease, the strongest association being for rs3803662 in TNRC9 (1.14 (1.09–1.21)). rs13281615 in 8q24 was associated with an improvement in survival after diagnosis (per-allele HR = 0.90 (0.83–0.97). The association was attenuated and non-significant after adjusting for known prognostic factors. Our findings show that common genetic variants influence the pathological subtype of breast cancer and provide further support for the hypothesis that ER-positive and ER-negative disease are biologically distinct. Understanding the etiologic heterogeneity of breast cancer may ultimately result in improvements in prevention, early detection, and treatment