7206854
Genetic Techniques
- PCR - Polymerase Chain Reaction
- Dna to be amplified, copies
identical Sequence of DNA, Start
martial could be DNA, RNA,
archival speicmen DNA, cloned
DNA, foresic samples
- 1. Denaturation of Double strand of DNA
- DsDNA - Temp -93-95'C -30 sec
- 2. Annealing
- Temp - 40-70'C -30 sec
- 3.DNA Synthesis
- Temp-70-75'C - 1 minute
- DNA polymerase extends the
primers, adding nucleotides onto
the primer in a sequential manner,
using the target DNA as a template.
- Temp-70-75'C - 1 minute
- This allows the primers to
bind (anneal) to their
complementary sequence in
the template DNA
- Temp - 40-70'C -30 sec
- This breaks the weak hydrogen bonds
that hold DNA strands together in a
helix, allowing the strands to separate
creating single stranded DNA.
- DsDNA - Temp -93-95'C -30 sec
- Product size - Length of forward
primer + Length of reverse
primer + region to be ampilfied
- Think - DNA replication outside the body
- Think - DNA replication outside the body
- There are five basic reagents, or ingredients,
necessary for PCR: template DNA, PCR primers,
nucleotides, Taq polymerase and PCR buffer.
- Template DNA
- starting DNA
that is used
- starting DNA
that is used
- PCR Primer
- is a short piece of
DNA that identifies
the region of DNA
- is a short piece of
DNA that identifies
the region of DNA
- Nucleotides
- subunit of a DNA
chain - A,T,C,G
- subunit of a DNA
chain - A,T,C,G
- Taq Polymerase
- DNA polymerase
from a bacterium
called Thermus
aquaticus
- DNA polymerase
from a bacterium
called Thermus
aquaticus
- PCR Buffer
- PCR buffer is a solution that
optimizes conditions like salt
concentration and pH to enable Taq
polymerase to work efficiently
- PCR buffer is a solution that
optimizes conditions like salt
concentration and pH to enable Taq
polymerase to work efficiently
- Template DNA
- Dna to be amplified, copies
identical Sequence of DNA, Start
martial could be DNA, RNA,
archival speicmen DNA, cloned
DNA, foresic samples
- Why is PCR used?
- Diagnosis and screening of genetic diseas
- Detection of slowly growing microoranisms
- HLA transplantation
- DNA finger prints
- Cloning/ Mapping
- Cloning/ Mapping
- DNA finger prints
- HLA transplantation
- Detection of slowly growing microoranisms
- Diagnosis and screening of genetic diseas
- Polymorphism detection
- DNA polymorphism - types of
mutations and variable number
of ministallite repeats (VNRS)
- DNA polymorphism - types of
mutations and variable number
of ministallite repeats (VNRS)
- RT PCR - 1 - 1. the RT reaction and PCR
amplification. RNA is first reverse transcribed
into cDNA using a reverse transcriptase as
described here, 2. the resulting cDNA is used as
templates for subsequent PCR amplification
using primers specific for one or more genes.
- DNA Copies for RNA targets RT PCR
- Transcription rates
- Cloning of rare gene
- Cloning of rare gene
- Transcription rates
- DNA Copies for RNA targets RT PCR
- Genetic Engineering
- Process that manipulates
Genes - normally
reproductive process
- Usually involves isolation,
manipulation and reintroduction
of DNA into cells or model
Organism
- Recombinant DNA technology
- INSULIN
- Basic procedure: 1. DNA purified from cells or tissues 2. Restriction enzymes used to generate
specific DNA fragments recognise and cut DNA at specific sequence 3. Fragments join to other DNA
molecules which act as vectors (carrier molecules), vector + DNA fragment= recombinant DNA
molecule 4. Recombinant DNA molecule transferred to host cell 5. Within host cell recombinant
replicates producing dozens of identical copies 6. As host replicates, recombinant molecule passed
to all progeny creating population of cells each carrying cloned DNA sequence 7. Cloned DNA can be
recovered from host cells, purified and analysed or 8. Cloned DNA in host cells is transcribed, its
mRNA translated, gene product isolated and used or sold.
- INSULIN
- CLoning
- Reproductive
- Is the creation of an exact genetic
replica of a small segment of DNA, a cell
or a whole organism
- Dolly the sheep
- Dolly the sheep
- Is the creation of an exact genetic
replica of a small segment of DNA, a cell
or a whole organism
- Therapeutic cloning
- Stem cells
- Stem cells
- Reproductive
- Gene therapy
- Used for correcting defective
gene, normal gene is inserted to
replace abnormal gene
- 1. Gene Augmentation Therapy – The introduction of DNA into a
cell with a view to augmenting gene function, e.g. recessivediseases.
2. Targeted Modulation of gene expression – Methods for turning
off genes using for example ribozymes and anti- sense
oligonucleotides, e.g., infectious disease and cancers
- Difficulties - only good for
LOF, targeting gene needed
for treatment, correct level of
gene expression, altering
pheontype
- Shift to cancer and HIV
- genetic disease
- Shift to cancer and HIV
- genetic disease
- Difficulties - only good for
LOF, targeting gene needed
for treatment, correct level of
gene expression, altering
pheontype
- 1. Gene Augmentation Therapy – The introduction of DNA into a
cell with a view to augmenting gene function, e.g. recessivediseases.
2. Targeted Modulation of gene expression – Methods for turning
off genes using for example ribozymes and anti- sense
oligonucleotides, e.g., infectious disease and cancers
- Used for correcting defective
gene, normal gene is inserted to
replace abnormal gene
- Process that manipulates
Genes - normally
reproductive process
- Genetic Counselling
- A child is born with several abnormalities There is a history in the family
of a medical condition A woman has a history of several miscarriages
Woman aged over 35 yrs and pregnant Anyone who has uncertainties
about genetic risks
- Genetic testing
- Pre-pregnancy- blood test
- In pregnancy - CVS -11-12 weeks or Amniocentesis - 15-19 weeks
- After pregnancy - Guthrie test, blood test
- Pre-pregnancy- blood test
- Genetic testing
- A child is born with several abnormalities There is a history in the family
of a medical condition A woman has a history of several miscarriages
Woman aged over 35 yrs and pregnant Anyone who has uncertainties
about genetic risks
Media attachments
Want to create your own Mind Maps for free with GoConqr? Learn more.