Bioinformatics Merging Molecular Biology with ... - MrFuscosClass

Techniques in Genetic
Engineering

•

PCR

has revolutionized
molecular biology and genetic
engineering.

•

RT
-
PCR (reverse
transcription PCR)

is used to
amplify RNA. Isolated RNA is
converted to double stranded
DNA by
reverse transcriptase
,
which is then amplified with a set
of primers specific for gene of
interest

Techniques in Genetic
Engineering

•

DNA microarray

analysis (gene chip) is
used to study gene
expression.

•

Gel electrophoresis
separates DNA
fragments by size in
order to create a “map”
of genes.

Blotting

•

Southern blotting

involves
the separation and blotting of
DNA molecules.

•

Northern blotting

involves
the separation and blotting of
RNA molecules.

•

Western blotting

involves
the separation and blotting of
proteins.

DNA Sequencing

DNA Sequencing

allows scientists
to:

•

Determine the exact structure of
gene

•

Deduce amino acid sequence of
the protein

•

Identify regulatory elements (such
as promoter sequences)

•

Identify differences in genes

•

Identify genetic mutations

Reading an autoradiograph

•
An
autoradiograph
is an image
produced on an X
-
ray film that
allows the visualization between the
reaction of DNA fragment and
probes used in sequencing.

•
They are “read” from the bottom to
the top as individual nucleotides.

•
The sequence determined is
complementary to the sequence on
the template strand.

Bioinformatics

Merging Molecular Biology with Information Technology

Bioinformatics

is an
interdisciplinary science
that involves developing
and applying information
technology (computer
hardware & software) for
analyzing biological data,
such as DNA & protein
sequences.

Bioinformatics

•

Includes the analysis
of molecular
structures

•

Creates databases
for storing and sharing
biological data

Databases

•

Essential for archiving and sharing data with
researchers and public

•

Many DNA sequences become available through the
Internet

•

When scientists clone a gene and enter the sequence,
databases search and align others with similar
sequences (determines if new or already cloned)

•

Used to predict sequence of amino acids & provide
info on gene function

GenBank

•

Largest publicly available
database of DNA sequences
(contains NIH collection)

•

Shares and acquires data
from Japan & Europe

•

Maintained by National Center
for Biotechnology Information
(NCBI)

GenBank

•

Contains more than 17
billion bases from over
100,000 species

•

Access and analyze
nucleotide sequences,
protein sequences,
molecular structures,
genome data, and even
scientific literature

Basic Local Alignment
Search Tool (BLAST)

•

Used to search GenBank for
sequence matches between cloned
genes

•

Each entry provided with
accession number that scientists
use to refer back to cloned
sequence

•

Provides original journal
reference, single letter amino acid
code for protein, nucleotide
sequence (cDNA) for gene

Basic Local Alignment
Search Tool (BLAST)

Basic Local Alignment
Search Tool (BLAST)

•

Human Genome
Nomenclature Committee
establishes rules for
assigning names and
symbols to newly cloned
human genes.

“Gene names should be brief
and specific and should convey
the character or function of the
gene, but should not attempt to
describe everything known
about it.”
-
HGNC

Bioinformatics

•

GenBank is only one example!

•

Many other databases are
specialized


* SNPs


* BAC and YAC databases


* Protein databases