Biomolecular Technology - Teoria (modulo 2) (2007/2008)

Course partially running (all years except the first)

Course code
4S00175
Name of lecturer
Barbara Molesini
Number of ECTS credits allocated
1
Academic sector
BIO/11 - MOLECULAR BIOLOGY
Language of instruction
Italian
Period
1° Sem dal Oct 1, 2007 al Jan 23, 2008.

To show the organization of the course that includes this module, follow this link * Course organization

Lesson timetable

1° Sem
Day Time Type Place Note
Thursday 8:30 AM - 12:30 PM lesson Lecture Hall I from Nov 22, 2007  to Jan 9, 2008
Friday 8:30 AM - 12:30 PM lesson Lecture Hall I from Nov 22, 2007  to Jan 9, 2008
Friday 2:30 PM - 6:30 PM lesson Lecture Hall I from Nov 22, 2007  to Jan 9, 2008

Syllabus

>NUCLEIC ACID HYBRIDIZATION

1.INTRODUCTION TO NUCLEIC ACID HYBRIDIZATION: FORCES STABILIZING DNA STRUCTURE, STABILITY OF NUCLEIC ACIDS, EFFECTS OF pH, EFFECTS OF TEMPERATURE, MELTING TEMPERATURE Tm, FACTOR AFECTING Tm
2.TYPES OF HYBRIDIZATIONS
3.SOLUTION HYBRIDIZATION: REASSOCIATION OF DNA, FACTORS AFFECTING REASSOCIATION/HYBRIDIZATION (FACTORS AFFECTING RATE OF HYBRIDS FORMATION AND STABILITY OF HYBRIDS)
4.FILTER HYBRIDIZATION: TYPES AND PROPERTIES OF SUPPORT, TRANSFER OF NUCLEIC ACID TO FILTER, FIXING THE DNA TO THE MEMBRANE, SCREENING OF RECOMBINANT LIBRERIES, DOT/SLOT BLOT AND REVERSE DOT/BLOT, SOUTHERN BLOT, NORTHERN BLOT (RNA EXTRACTION AND GEL ELECTROPHORESIS OF RNA SAMPLES)
5.CHOICE OF PROBES: DNA OR RNA ? RADIOACTIVE AND NONRADIOACTIVE PROBES, DIRECT AND INDIRECT LABELING, LABELING, UNIFORM LABELING OF THE PROBE (NICK-TRANSLATION, RANDOM PRIMED LABELING, PCR-MEDIATED LABELING, RUN-OFF TRANSCRIPTION, CHEMICAL METHODS), END LABELING (T4 POLYNUCLEOTIDE KINASE, FILL-IN BY KLENOW, DNA TAILING BY TERMINAL DEOXYNUCLEOTIDYL TRANSFERASE), DETECTION OF RADIOCTIVE HYBRIDS (AUTORADIOGRAPHY), DETECTION OF NONRADIOCTIVE HYBRIDS (ALKALINE PHOSPHATASE, HORSERRADISH PEROXIDASE, CHROMOGENIC SUBSTRATES, CHEMIOLUMINESCENT SUBSTARTES)
6.SPECIFIC APPLICATIONS OF FILTER HYBRIDIZATION: DNA MAPPING, DETECTION OF RELATED SEQUENCES, DNA FINGERPRINTING BY HYBRIDIZATION, DETECTION OF MUTATIONS, DETECTION OF METHYLATIONS


>PCR

1.WHAT IS PCR?
2.REAGENTS: EFFICIENCY, SPECIFICITY, FIDELITY
3.PCR CYCLE. FINAL NUMBER OF COPIES OF THE TARGET SEQUENCE
4.AMPLIFYING THE CORRECT PRODUCT: DETECTION AND ANALYSIS OF PCR PRODUCTS, HOW TO AVOID CONTAMINATION (URACIL N-GLYCOSYLASE, UV, ENZYMATIC TREATMENT), HOT START, NESTED PCR
5.TECHNIQUES AND APPLICATIONS: 5’RACE-PCR AND 3’RACE-PCR, RT-PCR,PCR MUTAGENESIS (DELETION OF SEQUENCES, BASE SUBSTITUTIONS, INSERTION MUTAGENESIS), MODIFICATION OF PCR PRODUCTS(INTRODUCTION OF RESTRICTION SITES, ADDING PROMOTERS AND RIBOSOME-BINDING SITES), JOINING OVERLAPPING PCR PRODUCTS, QUANTITATIVE PCR

Studying