Molecular Biology

PCR, RT-PCR, Gene sequencing, Genotyping, Gene expression.
Contact: Dr. Huanliang Liu (HLiu@med.miami.edu)

Isolation of DNA and RNA:  Clinical specimen processing, isolation of DNA and RNA, quality assessment and quantification, storing isolated DNA and RNA, maintenance of specimen records and repository collection.

PCR and RT-PCR:  experimental design of PCR and RT-PCR assay, cDNA synthesis, primer design and PCR condition optimization.

Candidate gene sequencing for genetic variation identification, genotyping and function analyses:  Consultation for candidate gene (s) selection, sample processing, primer and PCR condition optimization, RT-PCR, PCR, sequencing, sequence analyses, design of appropriate methods for genotyping, expression and function analyses of interesting genetic variations.

Gene expression quantification:  cDNA synthesis, design of absolute and relative quantitative qPCR experiments, and TaqMan qPCR quantification of gene expression.

Special partnership with Miami Institute for Human Genomics (MIHG): to provide completed services to identify, develop, and maintain cutting edge technologies and apply state-of-the-art methods in genomics, proteomics to the identification and characterization of human genetics HIV pathogenesis.  http://mihg.med.miami.edu - (Center for Genetic Epidemiology and Statistical Genetics, Center for Genome Technology, Center for Human Molecular Genomics, Center for Genomic Medicine, Center for Models of Human Disease, and the forthcoming Department of Human Genetics).

Maintenance and storage of cell lines, plasmids and transformed bacteria; manipulation of DNA, mutagenesis and plasmid propagation. 
Contact: Dr. Walter Scott (WScott@med.miami.edu)

These are standard procedures that most molecular biology laboratories carry out for themselves.  These procedures could be provided as a service if there is enough interest to provide salary support for the work.  In addition, we are willing to provide trouble-shooting advice.

Lentivirus construction, lentivirus stock preparation and titration. 
Contact: Dr. Walter Scott (WScott@med.miami.edu)
Use of lentivirus vectors has recently become widely used for a variety of experiments involving the introduction of genes into differentiated mammalian cells and for studying modifications in individual genes of HIV-1. The procedure basically involves the introduction of plasmid DNAs containing various portions of the HIV genome and the envelope gene from vesicular stomatitis virus to provide the packaging materials for forming virus particles that can infect a wide variety of cells. The gene to be delivered by the lentivirus vector is cloned into a plasmid that contains packaging signals so that lentivirus particles are made that contain almost exclusively the gene to be delivered.  Virus stocks are titered and then used to infect cells or tissues in culture or, in some cases, are injected into animals.

Note:  Preparation and use of lentivirus vectors falls under section III-D of the NIH Guidelines for Recombinant DNA Research, which requires prior submission to and approval by the Institutional Biosafety Committee (IBC).  Most lentivirus vector experiments can be carried out under BL2 containment or BL2-plus (using a BL2 facility with the laboratory procedures specified for BL3 experiments).  We will be happy to provide advice on submission of documents to the IBC and evaluation of biosafety plans.

siRNA and dominant negative mutant gene transfer
Contact: Dr. Walter Scott (WScott@med.miami.edu)

These are procedures often used to shut off the activity of specific genes in eukaryotic cells in order to study their functions.  Gene transfer may be accomplished by various methods including transfection and retrovirus medicated transduction.  Collaborative interactions are welcomed on such studies.