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Chemical
Chemical formula
Relative molecular mass (g)
Large molecules (such as starch, glycogen and enzymes) have indeterminate molecular masses and are best made up as percentage solutions. A 1% solution of an enzyme seems to work effectively in most cases. Techniques for Making Solutions
Making solutions can be time-consuming, especially if the solute is not very soluble (eg starch). The solubility of solutes increases with temperature, and therefore it is a good idea to warm the water when making many solutions. The solution needs to be stirred to ensure the solute disperses in the water, and to prevent ‘hot spots’ building up in certain parts of the beaker or flask. Magnetic stirrers are very useful in ensuring that a solution is properly mixed, and are strongly recommended for making starch solutions. Magnetic stirrers can be obtained from suppliers such as Philip Harris, (see page 118 for details). The glassware used to make solutions should be clean and of good quality. Cracked glassware should be disposed of in the usual way (see page 15). It is sometimes a good idea to add the solute to a smaller volume of water and then add the rest of the water once the bulk of the solute has dissolved. This prevents the solid solute displacing the water from the beaker. When making up a sucrose solution, for example, we recommend adding the solute to 500cm3 of water, then adding the remaining 500cm3 of water once the solution has dissolved. To make a 1% Starch Solution
To make a 1% starch solution, make a smooth paste with 10g of soluble (potato) starch and 10cm3 of distilled or deionised water. Pour the starch paste into 80cm3 of boiling water while stirring continuously. Make up to 100cm3 when cold. To make an Albumin Solution
When a percentage solution of albumin is to be prepared using albumen powder, add the powder a little at a time with gentle stirring, preferably with the aid of a magnetic stirrer. This can take the best part of a working day or may even have to be left stirring overnight. (If the powder is added en masse to the liquid with brisk stirring, the powder will float on the surface and very little will dissolve immediately. Also, foaming will occur and the solution will need filtration.) Albumen powder is preferable to egg white, as it will give an easy and accurate method for the calculation of solution strength. Dealing with egg white is messy! To make a Sodium Alginate Solution for Immobilised Enzymes
Sodium alginate may be needed for an immobilised enzyme practical. The powder should be added slowly, with stirring and GENTLE heating – a process that will take Daphnia can be fed on microscopic algae, suspensions of bacteria, yeast or Horlicks. It is important not to add too much food to the water. Enough should be added to enable the Daphnia to eat it in a short period of time so that the water becomes clear again. Daphnia are useful in laboratory experiments because their heart can be seen beating through the transparent carapace. They can be viewed using a cavity slide under low power of a microscope. Sometimes small strands of cotton wool can be used to stop the Daphnia moving around too much. Coverslips are not used. 0.5% caffeine solution can be added to the cavity slide to study the effect of caffeine on the heart rate. Plastic pipettes can be modified by having their ends cut off to transport Daphnia from the stock containers to the microscope slides. Drosophila
Drosophila are useful animals in advanced biology classes, but they require careful
preparation and maintenance. Recognising male and female flies requires expertise
and the timing of the maintenance can be critical at certain stages of a genetic
breeding experiment.
Detailed instructions on the care and maintenance of Drosophila are beyond the scope
of this chapter, but what follows is an overview of the key points. Further information on
keeping Drosophila can be found at: http://tinyurl.com/3cus4g.
Equipment and culture media for keeping Drosophila are readily obtained from
biological suppliers, such as
Philip Harris or Blades
Biological Ltd. The latter
company’s website
(www.blades-bio.co.uk/)
contains useful advice on
Drosophila.
Traditionally, stocks of Drosophila have been kept in small, 1/3 pint, milk bottles, and laboratory crosses (set bottom tubes. The diagram below shows a culture vessel. The plastic ladder helps to hold the medium in place. It is probably not a good idea to give students access to the master cultures of the flies. Sub-culture these bottles into smaller tubes for class use. The food medium often contains anhydrous copper sulphate which turns blue when water is added. Copper sulphate acts as a fungicide. A few grains of dried yeast are added to the medium to enable fermentation of the sugars in the medium to occur. Basic stocks can be kept at normal temperature 19-21°C and relative humidity 60-70% and should be sub-cultured every eight weeks or so. Cultures for experimental work should be kept at 26°C to obtain a fourteen day life-cycle. Female Drosophila mate only once, and when setting up crosses (and cultures) it is important to start with virgin female flies. Females do not mate until they are between 10-12 hours old. When collecting virgin female flies, clear all of the adult flies out of the culture bottles 8 hours before you collect the virgin females. Separate the male and virgin female flies into two separate bottles. Do NOT include any flies in the virgin female bottle unless you are certain of its gender. Male flies are generally smaller than the female flies, with a broader (and less pointed) abdomen. The bands on the abdomen tend to be thicker and often run together to form a solid band of black colour. Flies that have just emerged from pupae are fragile and light in colour and their wings are not fully expanded. These flies darken in a few hours and take on the normal appearance of the adult fly. Males have sex combs, which are groups of black bristles on the uppermost joint of the forelegs, whereas the females do not. One important distinguishing characteristic is the presence of the genital arch on the male flies. The tan coloured genital arch can be seen when the flies are placed on their backs. The genital arch is a reliable characteristic for distinguishing males in newly-emerged flies. It is necessary to anaesthetise Drosophila before handling them. A 1:1 mixture of ethoxyethane with ethanol is used as this reduces mortality due to prolonged exposure to the ether. A commercial anaesthetic, Flynap, is available from Blades Biological Ltd. An etheriser (as in the diagrams below) can be made by attaching some cotton wool to the underside of a cork bung with a hole drilled into it. A plastic funnel can be inserted into the hole. A few drops of the ethoxyethane/ethanol mixture can be added to the cotton wool. The cork is placed into a flat bottom tube and allowed to rest for a few moments. Flies can be gently tapped from the culture bottle into the etheriser. It is important not to over-etherise the flies, especially if they are to become the parents in a breeding experiment. An emergency etheriser can be made from a glass Petri dish lid with cotton wool taped to the underside. This can be used to re-anaesthetise any flies that become active before they have been examined. There are a variety of different mutant flies that can be obtained from biological suppliers (eg Blades Biological Ltd and Philip Section 8: Chemical Recipes
Azure A Stain for Nucleic Acids
100.0ml of 20% ethanol (20ml ethanol (or IDS, formerly known as IMS) made up to 100.0ml with distilled water) Dissolve Azure A stain in the prepared ethanol. Benedict’s Solution (Benedict’s Reagent)
Heat 800ml distilled water and add sodium citrate and sodium carbonate. Make up the volume to 850ml with distilled water. Separately dissolve the copper sulphate in 100ml distilled water. Slowly pour the second solution into the first with constant stirring. Make up the resulting deep blue mixture up to 1000ml with distilled water and bottle. Caffeine Solution
Dissolve caffeine powder in distilled water with stirring. Calcium Chloride Solution (for Immobilised Enzyme Practical)
30.0g calcium chloride (fused, granular) Dissolve calcium chloride (fused, granular, calcium chloride) in distilled water. It may take some time. Store in a bottle. It will keep for several months. Cobalt (II) Chloride Papers
2 or 3 discs of Whatman number 1 filter paper Make a 15% aqueous solution of cobalt (II) chloride by adding 15g cobalt (II) chloride to 100ml distilled water. Take filter paper and place in a container of distilled water for 1 minute until thoroughly wet. Carefully take out of the water and remove excess water by gently squeezing between blotting paper. The damp discs should then be placed in the 15% cobalt (II) chloride solution for 1 minute, taken out carefully and blotted as before then dried at 80oC. Store in a desiccator over silica gel.

Source: http://www.sfe.co.uk/products/examples/Biotech_examples.pdf