Optimisation of postmortem tissue preservation and alternative protocol for serotonin transporter gene polymorphisms amplification in SIDS and SIUD cases.
The major obstacle to genetic research in SIUD (sudden intrauterine unexplained death) and SIDS (sudden infant death syndrome) cases is the complex characteristics of the human anatomic samples available. In fact, in Italy autopsies are performed at least 24 h post-mortem and tissues can be left in formalin for long fixation times (>4/5 days), thus compromising nucleic acids integrity. In this study we compared the quality of DNA and RNA extracted from tissues differently preserved. As expected, the DNA and RNA from formalin-fixed and paraffin-embedded tissues, formalin-acetic acid-alcohol tissues and ethanol tissues were of poor quality and not adequate for subsequent molecular analysis. The best results were obtained with RNAlater preserved tissues: this buffer was equivalent, if not superior, to freezing method for preservation of postmortem DNA and RNA. In addition, we introduce a new protocol for the amplification of the serotonin transporter gene promoter region (5-HTT) ideal to obtain the increase of specific product, avoiding artifacts formation.
Casale
Valentina
V
Lino Rossi Research Center for the study and prevention of unexpected perinatal death and SIDS, University of Milan, via Commenda 19, 20122 Milan, Italy.
Oneda
Roberta
R
Lavezzi
Anna Maria
AM
Matturri
Luigi
L
D001344:Autopsy / Q000379:methods* / Q000592:standards; D004587:Electrophoresis, Agar Gel; D005313:Fetal Death / Q000175:diagnosis / Q000235:genetics*; D020022:Genetic Predisposition to Disease; D005865:Gestational Age; D006801:Humans; D007223:Infant; D007231:Infant, Newborn; D011110:Polymorphism, Genetic; D011180:Postmortem Changes; D050486:Serotonin Plasma Membrane Transport Proteins / Q000235:genetics*; D013398:Sudden Infant Death / Q000175:diagnosis / Q000235:genetics*; D016707:Tissue Fixation / Q000379:methods; D014021:Tissue Preservation / Q000379:methods* / Q000592:standards
D050486:Serotonin Plasma Membrane Transport Proteins