BIO 2133 Study Guide - Quiz Guide: Restriction Fragment Length Polymorphism, Amplified Fragment Length Polymorphism, Restriction Enzyme
27 Apr 2018
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MODULE 9: MOLECULAR MARKERS
LEARNING OBJECTIVES
•Explain what is meant by a single-nucleotide polymorphism (SNP) and how SNPs can be used as
genetic markers even if they do not cause phenotypic changes.
A single nucleotide polymorphism is a variation in a single nucleotide that occurs at a specific position
on the genome where each variation is present to some appreciable degree within the population
A gene is polymorphic if more than one allele occupies the gene’s locus within a population.
Protein polymorphisms exist for example, human blood groups, Rh factor and histocompatibility
complex, sickle cell anemia, cystic fibrosis
Enzymes are mostly polymorphic because they contain more than one variant of an enzyme
Enzymes encoded by the same locus slightly differ in amino acid sequences
These can be used as genetic markers because they are large in quantity and are usually stable
inheritance over generations.
Likewise, it is very easy to observe / identify changes through sequencing.
•Describe molecular genetic approaches used for studying genes, gene functions (knock-outs and
gene targeting technologies), and genomes to determine inheritance patterns and identities of genes
that can mutates
QUESTIONS
Molecular Techniques for Analyzing DNA (Molecular Markers)
•What are restriction enzymes and how do they work?
In the laboratory, restriction enzymes (or restriction endonucleases) are used to cut DNA into smaller
fragments. The cuts are always made at specific nucleotide sequences. Different restriction enzymes
recognise and cut different DNA sequences.
Like all enzymes, a restriction enzyme works by shape-to-shape matching. When it comes into contact
with a DNA sequence with a shape that matches a part of the enzyme, called the recognition site, it wraps
around the DNA and causes a break in both strands of the DNA molecule.
Each restriction enzyme recognises a different and specific recognition site, or DNA sequence.
Recognition sites are usually only short - 4-8 nucleotides.
•What type of molecular markers do they help us detect?
Restriction enzymes help us to detect restriction fragment length polymorphism (RFLP), single
nucleotide polymorphism (SNP), amplified fragment length polymorphism (AFLP).
•What are PCRs, how are they performed and what type of molecular markers do they !
help us detect?
PCRs help us to detect random amplified polymorphic DNA (RAPD), amplified fragment length
polymorphism (AFLP), simple sequence repeats (SSR), variable number tandem repeats (VNTR),
sequence characterized amplification region (SCAR).
•What are nucleotide hybridizations techniques and how do they help us detect !
molecular markers?
Nucleotide hybridization techniques are the techniques in which single-stranded nucleic acids (DNA or
RNA) are allowed to interact, and the complexes called hybrids are formed by molecules with similar,
complementary sequences.
•How is DNA sequencing performed and how can it help us detect molecular markers?
Single nucleotide polymorphism or SNPs are single base substitutions. SNP is the most abundant type
of molecular marker in all organisms. SNP causes variation in the restriction site of an enzyme. In such a
case, difference in the recognition of template DNA of different individuals, by the restriction enzyme, may
lead to different patterns of bands when DNA is digested, resulting in polymorphism. DNA sequencing
technique is useful to sequence the DNA of these individual to perform SNP.
MODULE 10: REVERSE GENETICS
LEARNING OBJECTIVES
•Explain why information on functions of human genes can often be acquired through studies of
simple organisms such as yeast, nematode worms, and fruit flies.
You can transform specific code into an animal and study the proteins and mRNA that result from that
gene.
•Explain how a gene that mutates to cause a disease can be molecularly identified
QUESTIONS
•What is meant by gene knockout and how can it be achieved?
a potent and irreversible means to inactivate a gene, can be achieved using conventional homologous
recombination or with engineered endonuclease
•What is a chimera and how are they useful for reverse genetics analysis
people that have two different sets of DNA, could happen when a woman is pregnant with fraternal
twins and one embryo dies early on - the other embryo can absorb its twin’s
cells
could also happen after a bone marrow transplant during normal pregnancy
•How do you create a transgenic organism?
genes of one species can be modified, or genes can be transplanted from
one species to another - made possible by recombinant DNA technology
most transgenic organisms are generated in labs for research, and example
are knockout mice
Take a bacterial plasmid. Cut out the portion of gene that you are interested
to incorporate (into the plasmid, from another organism), using a restriction enzyme. The same restriction
enzyme is used to cut the bacterial plasmid and using T4 DNA Ligase, incorporate the desired portion of
DNA into the plasmid. The incorporated portion is called a transgene.
Now, using a vector (bacteria/virus) which has a high multiplication rate; transport the transgene into
the desired organism.
The transgene replicates within the host cell or incorporates itself into the host DNA, and commands for
the production of required protein synthesis.
•What is gene silencing and how does it allow us to study the role of genes?
Document Summary
Learning objectives: explain what is meant by a single-nucleotide polymorphism (snp) and how snps can be used as genetic markers even if they do not cause phenotypic changes. A single nucleotide polymorphism is a variation in a single nucleotide that occurs at a speci c position on the genome where each variation is present to some appreciable degree within the population. A gene is polymorphic if more than one allele occupies the gene"s locus within a population. Protein polymorphisms exist for example, human blood groups, rh factor and histocompatibility complex, sickle cell anemia, cystic brosis. Enzymes are mostly polymorphic because they contain more than one variant of an enzyme. Enzymes encoded by the same locus slightly differ in amino acid sequences. These can be used as genetic markers because they are large in quantity and are usually stable inheritance over generations. In the laboratory, restriction enzymes (or restriction endonucleases) are used to cut dna into smaller fragments.
