DNA and RNA

Definitions:

  • Heredity is the passing on of characteristics/traits from one generation to the next
  • A gene is a short region of a chromosome that contains a code for the production of a protein
  • Gene expression is the process by which the code in DNA is used to make a protein

Genetic Code

  • The code for a particular protein can be thousands of bases long
  • Only approx 3% of DNA is thought to actually code for proteins (coding DNA)
  • The rest (97%)  is called non-coding DNA – does not code for any proteins

Chromosome Structure

  • The genes are contained within a much longer piece of DNA called a chromosome
  • The genes are spread out along the length of the chromosome
  • There is coding DNA (DNA that codes for a specific protein) and non-coding DNA (DNA whose function is generally unknown)
  • The non-coding regions of the chromosomes used to be called ‘junk DNA’
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  • Chromosomes are composed of 40% DNA: 60% protein
  • The protein (histones) makes the DNA very stable and enable it to be supercoiled into a very small space (i.e. the nucleus) •DNA wraps around proteins called histones, which then supercoil into a chromosome
  • NOTE: chromosome only exist during mitosis
  • At all other times the DNA is in the form of chromatin
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DNA

  • DNA stands for deoxyribonucleic acid
  • It is a polymer and has a helical shape.
  • DNA consists of two strands (double-stranded) – made up of alternating sugar (deoxyribose) and phosphate molecules
  • The two strands are attached to each other by nitrogenous bases.
  • DNA contains 4 bases:

1. Adenine (A)
2. Guanine (G)
3. Thymine (T)
4. Cytosine (C)
Adenine and guanine are purines
Thymine and cytosine are pyrimidines

  • The strands are twisted on themselves creating a spiral ladder
  • The spiral ladder shape is called a double helix
  • The bases attach the two strands together in pairs (complementary base pairing)
  • The bases always attach to the sugar molecules

Complementary base pairing

  • Complementary base pairing occurs between the bases in DNA:
  • Adenine can only pair with thymine (A = T)
  • Cytosine can only pair with guanine (C ≡ G)
  • A = T: double hydrogen bond
  • C ≡ G: triple hydrogen bond
  • Individual hydrogen bonds are very weak but as there are so many hydrogen bonds they are collectively very strong – holding the two strands of DNA together and making DNA very stable
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Nucleotide

  • A nucleotide is a 3 molecule unit composed of a phosphate, sugar (deoxyribose), and base (A, T, C or G)
  • It is the basic unit of the structure of DNA

DNA replication

  • DNA replication occurs towards the end of interphase
  • An enzyme unwinds and unzips the DNA
  • Free nucleotides diffuse in from the cytosol and are placed into their complementary position by the enzyme DNA polymerase
  • Once the DNA has been replicated the DNA coils and supercoils into chromosomes in preparation for mitosis/meiosis
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DNA Profiling

  • DNA profiling is a method of making a unique pattern of bands from a sample of DNA for identification purposes

DNA Profiling Method

1. DNA isolation:

DNA isolation: extraction/release of DNA from cells.

  • DNA is released from cells by using a type of detergent that splits open cell membranes
  • Even if the sample to be tested is very small (such as a hair follicle/blood smear) the amount of DNA can be increased (by DNA replication) several million-fold in a few hours!
2. Cutting:

Cutting: DNA is cut into fragments

  • Restriction enzymes cut DNA at specific base sequences
  • Products of this process are fragments of DNA (restriction fragments) that are different sizes
  • Everyone’s DNA is different which means that restriction enzymes will cut everyone’s DNA in slightly different places
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3. Separation:

Separation: restriction fragments of DNA are grouped based on size and moved apart

  • Because everyone has their own unique DNA they also have their own unique set of restriction fragments after the cutting stage
  • The mixture of restriction fragments can be separated out into a unique pattern of bands
  • The process of separating out the different fragments is carried out by gel electrophoresis•Agarose gel is poured into specialised shallow tray and allowed to set
  • The mixture of DNA is loaded into ‘wells’ at the top end (negative end) of the gel and an electric current is passed through the gel
  • DNA is a negatively charged molecule and will be attracted towards the positive end
  • The large restriction fragments will move more slowly than the short fragments – this creates a unique pattern of bands of fragments
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4. Pattern analysis:
  • An invisible pattern has been produced by the gel electrophoresis
  • To make the pattern visible the whole gel is stained (e.g. ethidium bromide) and then viewed under UV light
  • The patterns produced can then be compared and analysed for identification purposes

Applications of DNA Profiling

  • Species identification
  • Criminology: placing suspect at a crime scene
  • Medical: used often in paternity testing

RNA

  • RNA – ribonucleic acid
  • RNA is single stranded
  • RNA contains nitrogenous bases:

– Adenine (A)
– Uracil (U)
– Cytosine (C)
– Guanine (G)

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  • RNA contains the sugar ribose
  • Nucleotides in RNA are composed of a phosphate, sugar (ribose) and a base (A, U, C, or G)
  • RNA is made by the enzyme RNA polymerase using one DNA strand as a template (see picture below).

DNA versus RNA

DNA:RNA:
> Double-stranded
> Sugar: deoxyribose
> Nucleotides:
– Adenine
– Thymine
– Guanine
– Cytosine
> Single-stranded
> Sugar: ribose
> Nucleotides:
– Adenine
– Uracil
– Guanine
– Cytosine

Experiment: to isolate DNA from onion

  • Add a finely chopped small onion to 3 g salt and 100 ml distilled water
  • Heat (60˚C) and stir mixture gently for 15 minutes (heat denatures enzymes that breaks DNA down)
  • Cool in ice bath for 5 minutes (to prevent the DNA itself being broken down)
  • Blend mixture for 3 seconds (this step breaks the cell walls)
  • Filter blended mixture through coffee filter paper (not lab filter paper)
  • Take 3 ml of filtrate in test tube and add 3 drops of freshly squeezed kiwi fruit juice – slowly swirl the test tube to mix (kiwi fruit juice contains proteases that digest the histones)
  • Slowly add 10 ml ice cold ethanol down the side of the test tube (DNA is insoluble in ice cold ethanol)
  • DNA becomes visible at the junction between the filtrate and ethanol
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Chapter 14 notes page

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