@article {10457, title = {Avoiding Ethanol Presence in DNA Samples Enhances the Performance of Ultraviolet Resonance Raman Spectroscopy Analysis.}, journal = {Appl Spectrosc}, volume = {71}, year = {2017}, month = {2017 Jan}, pages = {152-155}, abstract = {

Ethanol is an essential chemical reagent in DNA preparation as its use increases the yield of extraction. All methodologies for DNA isolation involve the use of ethanol in order to prevent DNA dissolution in water and to optimize the binding of DNA to chromatographic membranes. In this note, we show how the presence of ethanol traces in DNA aqueous solution affects ultraviolet Raman spectra, leading to possible misinterpretations. We report a simple method to remove the ethanol Raman features from the spectra, based on heating the DNA sample at 80 {\textcelsius}, followed by a slow cooling procedure.

}, keywords = {DNA, Ethanol, Female, Humans, Placenta, Pregnancy, Spectrophotometry, Ultraviolet, Spectrum Analysis, Raman}, issn = {1943-3530}, doi = {10.1177/0003702816654152}, author = {Cammisuli, Francesca and Pascolo, Lorella and Morgutti, Marcello and Gessini, Alessandro and Masciovecchio, Claudio and D{\textquoteright}Amico, Francesco} } @article {1877, title = {A real-time polymerase chain reaction-based protocol for low/medium-throughput Y-chromosome microdeletions analysis.}, journal = {Genet Test Mol Biomarkers}, volume = {16}, year = {2012}, month = {2012 Dec}, pages = {1349-55}, abstract = {

PURPOSE: We describe a real-time polymerase chain reaction (PCR) protocol based on the fluorescent molecule SYBR Green chemistry, for a low- to medium-throughput analysis of Y-chromosome microdeletions, optimized according to the European guidelines and aimed at making the protocol faster, avoiding post-PCR processing, and simplifying the results interpretation.

METHODS: We screened 156 men from the Assisted Reproduction Unit, Department of Obstetrics and Gynecology, Institute for Maternal and Child Health IRCCS Burlo Garofolo (Trieste, Italy), 150 not presenting Y-chromosome microdeletion, and 6 with microdeletions in different azoospermic factor (AZF) regions. For each sample, the Zinc finger Y-chromosomal protein (ZFY), sex-determining region Y (SRY), sY84, sY86, sY127, sY134, sY254, and sY255 loci were analyzed by performing one reaction for each locus.

RESULTS: AZF microdeletions were successfully detected in six individuals, confirming the results obtained with commercial kits.

CONCLUSION: Our real-time PCR protocol proved to be a rapid, safe, and relatively cheap method that was suitable for a low- to medium-throughput diagnosis of Y-chromosome microdeletion, which allows an analysis of approximately 10 samples (with the addition of positive and negative controls) in a 96-well plate format, or approximately 46 samples in a 384-well plate for all markers simultaneously, in less than 2 h without the need of post-PCR manipulation.

}, keywords = {Azoospermia, Chromosome Deletion, Chromosomes, Human, Y, Female, Humans, Kruppel-Like Transcription Factors, Male, Real-Time Polymerase Chain Reaction, Sex Chromosome Aberrations, Sex Chromosome Disorders of Sex Development}, issn = {1945-0257}, doi = {10.1089/gtmb.2012.0220}, author = {Segat, Ludovica and Padovan, Lara and Doc, Darja and Petix, Vincenzo and Morgutti, Marcello and Crovella, Sergio and Ricci, Giuseppe} } @article {1661, title = {HLA-G 14 bp deletion/insertion polymorphism in celiac disease.}, journal = {Am J Gastroenterol}, volume = {106}, year = {2011}, month = {2011 Jan}, pages = {139-44}, abstract = {

OBJECTIVES: Nonclassical major histocompatibility class I HLA-G antigen is a tolerogenic molecule that inhibits lytic activity of natural killer (NK) cells and cytotoxic T lymphocytes. Because of its immunomodulatory and tolerogenic properties, HLA-G molecules may have a role in celiac disease (CD). We analyzed the HLA-G 14 bp deletion/insertion polymorphism, known to have a functional effect on mRNA stability, in a group of 522 CD patients, stratified for the presence of HLA-DQ2 genotype, and 400 healthy individuals to evaluate the possible effect of the polymorphism on the risk to develop the disease.

METHODS: HLA-G 14 bp deletion/insertion polymorphism (rs1704) was detected by polymerase chain reaction and double-checked by direct sequencing.

RESULTS: The 14 bp inserted (I) allele and the homozygous I/I genotype were significantly more frequent in CD patients than in healthy controls. The presence of I allele was associated with an increased risk of CD (OR 1.35) and the effect of I allele was consistent with a recessive genetic model (P<0.001).

CONCLUSIONS: Our results also indicate that the effect of the HLA-G D/I polymorphism is restricted for HLA-DQ2, and not simply due to the presence of linkage disequilibrium with the major known risk factor; moreover we found that the presence of the I allele confers an increased risk of CD in addition to the risk conferred by HLA-DQ2 alone and that subjects that carry both DQ2 and HLA-G I alleles have an increased risk of CD than subjects that carry DQ2 but not the I allele.

}, keywords = {Adolescent, Adult, Aged, Aged, 80 and over, Alleles, Case-Control Studies, Celiac Disease, Child, Child, Preschool, Confidence Intervals, Female, Genetic Predisposition to Disease, Genotype, Histocompatibility Antigens Class I, HLA Antigens, HLA-DQ Antigens, HLA-G Antigens, Humans, Male, Middle Aged, Mutagenesis, Insertional, Odds Ratio, Polymerase Chain Reaction, Polymorphism, Genetic, Reference Values, RNA Stability, Sequence Deletion, Young Adult}, issn = {1572-0241}, doi = {10.1038/ajg.2010.340}, author = {Fabris, Annalisa and Segat, Ludovica and Catamo, Eulalia and Morgutti, Marcello and Vendramin, Anna and Crovella, Sergio} } @article {1644, title = {A polymorphism in the 5{\textquoteright} UTR of the DEFB1 gene is associated with the lung phenotype in F508del homozygous Italian cystic fibrosis patients.}, journal = {Clin Chem Lab Med}, volume = {49}, year = {2011}, month = {2011 Jan}, pages = {49-54}, abstract = {

BACKGROUND: The identification of cystic fibrosis (CF) patients who are at greater risk of lung damage could be clinically valuable. Thus, we attempted to replicate previous findings and verify the possible association between three single nucleotide polymorphisms (SNPs c.-52G>A, c.-44C>G and c.-20G>A) in the 5{\textquoteright} untranslated region (5{\textquoteright} UTR) of the β defensin 1 (DEFB1) gene and the CF pulmonary phenotype.

METHODS: Genomic DNA from 92 Italian CF patients enrolled in different regional CF centres was extracted from peripheral blood and genotyped for DEFB1 SNPs using TaqMan({\textregistered}) allele specific probes. In order to avoid genetic confounding causes that can account for CF phenotype variability, all patients were homozygous for the F508del CFTR mutation, and were then classified on the basis of clinical and functional data as mild lung phenotype (Mp, n=50) or severe lung phenotype patients (Sp, n=42).

RESULTS: For the c.-20G>A SNP, the frequency of the A allele, as well as the AA genotype, were significantly more frequent in Mp than in Sp patients, and thus this was associated with a protective effect against severe pulmonary disease (OR=0.48 and 0.28, respectively). The effect of the c.-20G>A A allele is consistent with a recessive model, and the protective effect against Sp is exerted only when it is present in homozygosis. For the other two SNPs, no differences were observed as allelic and genotypic frequency in the two subgroups of CF patients.

CONCLUSIONS: Our results, although necessary to be confirmed in larger and multiethnic populations, reinforce DEFB1 as a candidate modifier gene of the CF pulmonary phenotype.

}, keywords = {5{\textquoteright} Untranslated Regions, Adult, beta-Defensins, Cystic Fibrosis, Cystic Fibrosis Transmembrane Conductance Regulator, Female, Genotype, Homozygote, Humans, Italy, Male, Mutation, Phenotype, Polymorphism, Genetic, Young Adult}, issn = {1437-4331}, doi = {10.1515/CCLM.2011.023}, author = {Crovella, Sergio and Segat, Ludovica and Amato, Annalisa and Athanasakis, Emmanouil and Bezzerri, Valentino and Braggion, Cesare and Casciaro, Rosaria and Castaldo, Giuseppe and Colombo, Carla and Covone, Angela Elvira and De Rose, Virginia and Gagliardini, Rolando and Lanzara, Carmen and Minicucci, Laura and Morgutti, Marcello and Nicolis, Elena and Pardo, Francesca and Quattrucci, Serena and Raia, Valeria and Ravazzolo, Roberto and Seia, Manuela and Stanzial, Valentino and Termini, Lisa and Zazzeron, Laura and Cabrini, Giulio and Gasparini, Paolo} } @article {1715, title = {Five new OTOF gene mutations and auditory neuropathy.}, journal = {Int J Pediatr Otorhinolaryngol}, volume = {74}, year = {2010}, month = {2010 May}, pages = {494-8}, abstract = {

OBJECTIVE: Purpose of this paper is to analyse OTOF gene in a series of subjects affected by auditory neuropathy.

METHODS: Four children showing mild to profound prelingual deafness, confirmed by the absence of a clear and detectable responses at auditory brainstem responses (ABR), associated with the presence of bilateral OAE, were enrolled in the study.

RESULTS AND CONCLUSIONS: Genetic analysis identified five new mutations (a nonsense, a small and a large deletion and two splicing site mutations), and one missense mutation (F1795C) previously described. These results further confirm the role of OTOF gene in auditory neuropathy. In the absence of a context of neurological syndrome, the combination of absent ABR and positive OAE responses should lead to an auditory neuropathy diagnosis and to a mutational screening in OTOF.

}, keywords = {Auditory Diseases, Central, Deafness, Evoked Potentials, Auditory, Brain Stem, Female, Humans, Male, Membrane Proteins, Mutation, Otoacoustic Emissions, Spontaneous, Polymerase Chain Reaction}, issn = {1872-8464}, doi = {10.1016/j.ijporl.2010.02.004}, author = {Zadro, Cristina and Ciorba, Andrea and Fabris, Annalisa and Morgutti, Marcello and Trevisi, Patrizia and Gasparini, Paolo and Martini, Alessandro} } @article {1700, title = {HLA-G*0105N allele is associated with augmented risk for HIV infection in white female patients.}, journal = {AIDS}, volume = {24}, year = {2010}, month = {2010 Jul 31}, pages = {1961-4}, abstract = {

We analyzed HLA-G 3777G > C, HLA-G 14 bp deletion/insertion and HLA-G*0105N polymorphisms in HIV-positive white adult participants, infected through horizontal heterosexual transmission, and unexposed uninfected individuals, all from north eastern Italy. We report a new association between the HLA-G*0105N allele and HIV infection in adult white female participants, being HLA-G*0105N null allele correlated with an augmented risk (odds ratio = 4.35, 95\% confidence interval = 1.38-18.07, P = 0.005) for HIV infection.

}, keywords = {Adolescent, Adult, Aged, Female, Genetic Predisposition to Disease, Histocompatibility Antigens Class I, HIV Infections, HIV-1, Humans, Middle Aged, Polymorphism, Genetic, Young Adult}, issn = {1473-5571}, doi = {10.1097/QAD.0b013e32833c3324}, author = {Segat, Ludovica and Catamo, Eulalia and Fabris, Annalisa and Morgutti, Marcello and D{\textquoteright}Agaro, Pierlanfranco and Campello, Cesare and Crovella, Sergio} }