topic 6 Flashcards
exons
coding sections of DNA
introns
these are no coding sections of DNA
short tandem repeats
short repeated sequences of bases
there can be several hundred copies of the same STR at a single locus on both chromosomes of a homologous pair
people vary in regard to the number of these repeats at a locus of a chromosome
STRs can be used to build up a unique pattern DNA profile
as it is highly unlikely to have 2 individuals with the same combination of STRs
how is the DNA obtained
A DNA sample can be obtained from almost all biological tissue, animal or plant.
check swab, blood cells in a blood smear at scene, bone marrow in a skeleton, sperm left after a sexual assault.
The tissue sample is the physically broken down in a buffer solution containing salt and a detergent to disrupt cell membrane.
The small suspended particles, including DNA is separated from cell derbies by filtering or centrifuging.
protease enzyme are incubated with suspension to remove proteins
and the cold ethanol is added to precipitate out the DNA. then the DNA is washed in buffer solution
how do you create the DNA fragments for PCR
restriction enzymes. found naturally in bacteria to cut up invading viral DNA. the enzymes only cut DNA at specific base sequences.
if the restriction sites are either side of a short tandem repeat sequence that fragment of DNA will remain intact, but it will be cut away from the rest of the genome
if the same restriction enzyme is used to cut 2 identical DNA samples, identical STR fragments are produced
what does PCR stand for
polymerase chain reaction
what are the stages in PCR
- 95oc for 30 seconds. the DNA separates into 2 strands as hydrogen bonds between bases break.
- 55oc for 20 seconds. the primers attach at the start of the STR repeat sequence
- 70oc for 1 minute. DNA polymerase attach nucleotides are attached, extending the DNA from the primer. The STR repeated sequence and DNA adjacent is replicated.
this sequence is repeated many times in order to create millions of these STR fragments.
what needs to go into the reaction tube in PCR
- DNA polymerase. to replicate DNA
- DNA primers with fluorescent markers. attaches to the DNA and allows DNA polymerase to attach and is view-able after gel electrophoresis
- nucleotides. In order to create the New DNA.
- a sample of DNA
DNA primers
short DNA sequences complemntary to the DNA adjacent to the STR.
the DNA primers are marked with fluorescent tags.
what is gel electrophoresis
the process of separating the DNA produced by PCR according to size
stages in gel electrophoresis
- DNA fragments placed in well of agarose submerged in buffer solution in gel tank
- negative charged DNA moves towards the positive electrode. Fragments separates into invisible bands according to their size.
- transferred to nylon or nitrocellulose membrane. DNA probe binds with fragment/
- X ray film or UV light used to view DNA profile
what is DNA probe.
labelled DNA probe is a short section of DNA with a base sequence complementary to the target DNA sequence that needs to be located. the probe binds to any complementary sequences.
if probe is radioactive. X ray film is used.
if probe is fluorescent, its position on the membrane can be visualised under ultraviolet light.
one way of viewing a DNA profile produced is gel electrophoresis is by actually viewing either as an X ray film or under UV light. however there is another way to view the DNA profile…
during PCR DNA primers attach to DNA these DNA primers have fluorescent tags attached to them.
as DNA fragments with their attached fluorescent tags move through the gel they pass a laser, the dye in the tag fluoresces and the coloured light is detected.
this gives a time that it has taken for the fragment to pass through the gel. passing a separate set of fragments of a known length through gel allows the length of time for passage through the gel to be calibrated with fragment size.
the size of fragment is determined by the number of base pairs it contains.
several STR loci can be analysed at once using tags that fluoresce at different wavelengths giving different colours.
produces a digital DNA profile with a series of numbers corresponding to the number of repeated in each fragment
ways of determining time of death (4)
- body temperature
- rigour mortis
- decomposition
- forensic entomology
how does body temperature change after death
body starts to cool as soon as death occurs
it follows a sigmoid curve
initial temperature plateau for 30 minutes then falls rapidly until the same temperature as the environment
temperature decline per hour can give an estimate of time of death
factors affecting post mortem cooling (7)
- body size
- body position
- clothing
- air movement
- humidity
- hypothermia or fever
- temperature of surroundings
what is rigor mortis
after death the muscles usually total relax and then stiffen.
joints become fixed during rigor mortis and their position will depend on the body position at time of death. after a further period of time rigor mortis will pass and the muscles will relax again.
stages in rigor mortis (6)
- after death, muscle cells become starved of oxygen and oxygen dependent reactions stop.
- Respiration in the cells becomes anaerobic and produces lactic acid
- the pH of the cell falls, inhibiting enzymes and thus inhibiting anaerobic respiration.
- The ATP needed for muscle contraction is no longer produced. as a result, bonds between the muscle proteins become fixed.
- the proteins can no loner move over one another to shorten the muscle, fixing the muscle and joints.
- after a whole muscle tissue starts to break down and rigor mortis passes
Rigor mortis starts with smalled muscles stiffening before larger ones.
stages of decomposition
- autolysis
- putrefaction
- bloated
- dry body
autolysis
the first stage of decomposition
when the body own enzymes, from the digestive tract and lysosomes breaks down cells
bacteria from the gut and gaseous exchange system rapidly invade the tissues after death, releasing enzymes that result in decomposition. the loss of oxygen in the tissues favours the growth of anaerobic bacteria
putrefaction
second stage of decomposition
greenish discoloration of the skin of lower abdomen
due to formation of sulphaemoglobin in the blood
as it spreads across the body skin will darken to reddish-green and then turn a purple black in colour.
gas or liquid blisters may appear on the skin.
bloated
the 3rd stage of decomposition bloating of body due to gas production increased action of bacteria produce gases including hydrogen sulphide, methane, Carbon dioxide, ammonia and hydrogen. these form in the intestines and tissues. body will smell and become bloated. with further decomposing will deflate
dry body
the 4th and final stage in decomposition
the body deflates and fluids associated with putrefaction drains away.
soft tissues dry out and shrink
how does environmental temperature affect rate of decomposition
- low temperature slows down decomposition
- warm temperatures speeds it up
- intense heat denatures enzymes involved in autolysis delaying decay
ways of identifying a body (3)
- finger prints
- dental records
- DNA profile
forensic entomology
the study of organisms on a decaying body to estimate time since death
succession in corpses first wave
organisms present - bluebottles - green bottles - house fly - cow face fly state of body: fresh
succession in corpses second wave
state of body
bloated by gases.
species present
- flesh flies
succession in corpses third wave
state of body active decomposition (fatty acids have turned to a waxy substance) species present - beetle larvae - tabby moth maggot
succession in corpses fourth wave
state of body active decomposition (fermentation) species present - cheese skipper - lesser house fly
what is succession in corpses
as one organism feeds on a decomposing body, conditions change in such a way that it becomes attractive to another group of organisms.
as corpse continues to change, it attracts different organisms that feed on it until only the skeleton remains
forensic entomologists use the predictable sequence of organisms that feed on a decomposing body to determine time of death
decomposition role in the carbon cycle
a corpse is a great source of energy for decomposes
carbon dioxide is released into the atmosphere by respiring decomposes
this recycles the carbon back into a form that can be used in photosynthesis to synthesis more organic molecules.
what are the differences between bacteria and viruses
bacteria contain cell surface membrane, cytoplasm, cell wall, ribosomes, plasmids and sometimes mesosomes, flagellum and pilli.
viruses contain no cell wall, cell surface membrane, cytoplasm or organelles. nucleic acid enclosed in protein coat
bacteria have circular strand of DNA as the genetic material.
viruses have DNA or RNA as genetic material
bacteria can live independently.
Viruses must have a living organisms as host
bacteria’s average diameter is 0.5 - 5 um.
viruses 20- 40 nm wide range of sizes and shapes
bacteria often have mucus based outer capsule.
virus may have outer membrane of host cell surface membrane, but containing glycoproteins from the virus
features of virus
10 -100 times smaller than bacteria have no cell wall or ribosomes not a living cell nucleic acid DNA or RNA present infects plants, animals and bacteria can reproduce with host cell a sexual reproduction
what do viruses consist of
a strand of nucleic acid (RNA /DNA) enclosed within a protein coat
the DNA can be single or double stranded
some have outer envelope taken from the host cell’s surface membrane. the envelope contains lipids, proteins and glycoproteins from virus itself and act as antigens
how do viruses invade cells and reproduce (6 steps)
- virus attaches to host cell
- virus inserts nucleic acid
- viral DNA is incorporated into the human DNA
- viral DNA is replicated
- viral protein coat is synthesised
- new virus particles are formed
- virus particles released due to cell lysis
characteristics of gram positive bacteria
- peptidoglycan cell wall
- takes up violet stain but not safranin
- deep purple colour after staining
- eukayotic cell
- nucleus surrounded by envelope
- mitochondria
- large ribosomes
- contains cytoskeleton
- several long strands of DNA associated with histones
- sometimes contain cillia and flagella.
characteristics of gram negative bacteria
- liposaccharid layer outside peptidoglycan cell wall
- does not take up crystal violet. takes up safarin
- pink colour
- prokayrotic cell
- does not contain nucleus, mitochondria or cytoskeleton small 20nm ribosomes
- circular DNA with no histones
- some have flagella.
symptoms of HIV / AIDS
viral disease
HIV - human immunodeficiency virus
initial symptoms are fevers, headaches, tiredness, swollen glands or no symptoms at all
after 12 weeks HIV antibodies appear in the blood
all symptoms disappear for years but eventually patients will suffer weight loss, fatigue, diarrhoea, night sweats and infections such as thrush. this is full blown AIDs
dementia, cancer and opportunistic infections such as TB take hold
what is AIDs
acquired immune deficiency syndrome
is caused by infection of HIV
symptoms of AIDs are those of opportunistic infections such as TB
HIV is a envelope virus. the lipid envelope is formed from the host cell membrane sticking through the envelope are viral glycoproteins
how is HIV transferred
- HIV is found in the blood, vaginal secretions and semen
transmitted by direct blood / body fluid transfer through cuts
needle sharing, unprotected sex
maternal transmission from mother to unbron child or in breast milk.
what are the phases of HIV infection / AIDs
- acute phase
- chronic phase
- Disease phase
acute phase of HIV infection
HIV antibodies appear in blood 3-12 weeks after initial infection
sweats, fever, headache, sore throat, swollen lymph nodes or no symptoms at all.
there is rapid replication of virus and loss of T helper cells
after a few weeks infected T helper cells are recognised by T killer cells and destroyed reducing rate of viral replication
chronic phase of HIV infection / AIDs
second phase
also called latent stage
virus continues to reproduce rapidly but numbers are kept in check by the immune system
no symptoms but increasing tendency to suffer colds or other infections slow to rid of
dormant diseases like TB or shingles can reactivate
this phase can last for a long time with correct lifestyle and drug treatment
disease phase of HIV infection / AIDs
increased viral load and declining T helper cells indicates the onset of AIDs
low number of T helper cells leaves immune system vulnerable
high risk of infection by opportunistic such as pneumonia and TB
significant weight lost, dementia and tumours such as kaposisarcoma
antigens
any molecule the body recognises as not being of its own self.
what is the difference between non specific immune response and specific immune response
non specific immune response helps to destroy any invading pathogen whereas specific immunity is always directed at a specific pathogen.
what are the no specific responses to infection
- lysozyme
- inflammation
- phagocytosis
- lymph system
- interferon
lysozyme
is an enzyme that kills bacteria by breaking down their cell wall.
it is found in tears, saliva and nasal secretions
protects the body from harmful bacteria in the air we breath in or the food that we eat.
inflammation
inflammatory response
damaged white blood cells and mast cells release histamine which causes arterioles to dilate increasing blood flow and capillary permeability causing leakage
plasma fluid, white blood cells and antibodies leaks into tissues causing oedema
infecting bacteria are now attacked by intact white blood cells.
