Molecular Genetic

Agustini Utari

Outline
Modes of Inheritance Gene Structure Mutation type Methods for Detecting Mutation Some of Chromosomal abnormality

MODES OF INHERITANCE
Mendelian Inheritance Autosomal dominant Autosomal resessive X-linked

Autosomal Dominant

MODY, Achondroplasia, Marfan syndrome

Autosomal Recessive

CAH due to CYP21 gene mutation, 5-reductase deficiency, Pendred syndrome, Laron dwarfism

X-linked inheritance

Androgen Insensitivity Syndrome

MODES OF INHERITANCE
Non Mendelian Inheritance Anticipation Genomic Imprinting Mitochondrial Inheritance Multifactorial and Polygenic inheritance

Anticipation
Mostly seen in an autosomal dominant or Xlinked Not all affected individuals are affected to the same degree Severity of the disease increases with each generation Trinucleotide Repeat Expansion (Fragile X (CGG), Huntington disease (CAG)

Genomic Imprinting
Parent of origin effect Not all genes are expressed equally from both chromosome the differential expression of genetic material depending on whether it was inherited from the male or female parent

Uniparental Disomy (UPD)

UPD refers to inheritance of both copies of a chromosome from the same parent Prader Willi Syndrome (PWS) and Angelman Syndrome

Mitochondrial Inheritance
DNA is not only located in the nucleus, but also in mitochondria After fertilization, the mitochondria in the zygote are only derived from the oocyte, not from the sperm cell mutation only inherited via the mother Two characteristic features : - only if certain percentage of mtDNA is mutated, a phenotypic effect will be seen ( Heteroplasmy --the presence of more than one mitochondrial DNA (mtDNA) variant within a cell ) - the degree of heteroplasmy may differ from tissue to tissue

Mitochondrial Inheritance
Ex : m. 3242 A>G mtDNA mutation Cause MELAS (Mitochondrial myopathy, Encephalopathy, Lactic Acidosis and Stroke Like Episodes) if heteroplasmy is high in brain and or muscle Cause Mitochondrial Diabetes if it is present at high level in the pancreas

Multifactorial and poligenic inheritance

Many factors play a role in causing this disorders Genetic, environmental Ex : - Diabetes Melitus - Cleft lip/cleft palate - Bipolar disorders

HUMAN GENETIC

HUMAN GENETIC
46 chromosome : 23 pair, including 2 sex chromosome Of all chromosome, one copy is from father (paternal) , the other from mother (maternal) Chromosome consist of DNA, the real genetic material

DNA
Four different nucleotides : A, C, G, T A is complementary to T C is complementary to G

Transcription : DNA RNA DNA : A, C, G, T double stranded RNA : A, C, G, U single stranded

The Genetic Code

GENE STRUCTURE

Mutation Type
Point Mutation
Missense Mutation Nonsense Mutation Silent Mutation

Deletions and insertions Splice Mutations

A silent mutation alters DNA sequence, but has no apparent detectable effect on a phenotype or a function.

An insertion changes the number of DNA bases in a gene by adding a piece of DNA. As a result, the protein made by the gene may not function properly.

A deletion changes the number of DNA bases by removing a piece of DNA. Small deletions may remove one or a few base pairs within a gene, while larger deletions can remove an entire gene or several neighboring genes. The deleted DNA may alter the function of the resulting protein(s).

 A duplication consists of a piece of DNA that is abnormally copied one or more times. This type of mutation may alter the function of the resulting protein

SPLICE MUTATION A mutation that alters or abolishes the specific sequence denoting the site at which the splicing of an intron takes place. Such mutations result in one or more introns remaining in the mature messenger RNA and can disrupt the generation of the protein product

Techniques for Detecting and Characterizing Mutation

Sample Preparation
DNA analysis 5 ml EDTA treated blood in unfrozen form DNA can be extracted from blood spot on a guthrie card

PRIMER
A short synthetic oligonucleotide Primers are designed to have a sequence which is the reverse complement of a region of the target DNA thus it can anneal to the target sequence

Polymerase Chain Reaction (PCR)

A piece of DNA is amplified by in vitro enzymatic replication PCR can amplify a single piece or a very small number of pieces of DNA generating million of copies of the original DNA Widely used in molecular biology

DNA Sequencing
The most reliable method to confirm a genetic defect Provide the exact location and nature of the disorders

Restriction Fragment Length Polymorphism (RFLP)

The simplest and earliest method to detect Single Nucleotide Polymorphisms (SNPs) SNP- RFLP is based on availability of several different restriction enzymes and their specifity towards unique restriction site

RLFP for UCP2, using acrylamide gel

UCP3, agarose gel

The molecular Y detection: by identification of SRY gene

Marker
1000bp

XY XX XX

XY

Negative SRY gene =Female

TERIMA KASIH