Dean Madden from the National Centre for Biotechnology Education at the University of Reading, UK, describes how DNA was discovered - and how it can be simply extracted in the classroom.
In 1868, Johann Friedrich Miescher travelled from his native Switzerland to Tübingen in Germany. The 24-year-old had come to study in the laboratory of Ernst Felix Hoppe-Seyler, a pioneering biochemist who coined the modern name for the red pigment in blood: haemoglobin. After several months of toil in a laboratory in the cellar of Tübingen Castle, Miescher managed to isolate a previously unknown acidic substance from white blood cells (leucocytes) which had been washed from pus-laden bandages from a nearby hospital. Miescher called his discovery 'nuclein' because it was found in the nuclei of the cells. This substance was impure, however, and Hoppe-Seyler insisted on repeating the work himself before he would allow an account to be published in his newly formed biochemistry journal.
Upon returning to his home in Basel in 1870, Miescher refined the method and was able to extract nuclear material from the sperm of the salmon for which, in those days, the Rhine was famed. Like those in leucocytes, the nuclei in sperm cells are relatively large. From these, Miescher extracted pure nuclein for the first time. In 1889 a pupil of his, Richard Altmann, gave us the modern term for nuclein: nucleic acid. Thus, in 2003, we celebrated the golden jubilee of the double helix and not '50 years of DNA'.
To a science teacher, an account of Miescher's methods makes fascinating reading, not least because of the crude techniques available and the striking resemblance they bear to many of today's classroom protocols. Compared with Miescher, however, today's teachers have an easy time. With no refrigeration, Miescher had to start work at 5 a.m. to ensure that the reagents were cold enough to precipitate DNA, and he had to prepare his own protease enzymes from the stomachs of freshly slaughtered pigs (Judson, 1996).
The isolation of DNA from everyday materials has become a popular and widespread activity in school laboratories over the past 20 years. Although similar practical protocols had been described previously (e.g. Sands, 1970), these were not widely adopted owing to their complexity and the hazardous nature of several of the required reagents (Falconer & Hayes, 1986). Simpler methods of isolating DNA first appeared in American textbooks in the mid-1980s (e.g. Helms et al., 1986) and subsequently made their way into specialist school biotechnology projects (e.g. Rasmussen & Matheson, 1990). By the early 1990s, these methods had crossed the Atlantic, featuring in German and English publications (Bayrhuber et al., 1990; NCBE, 1991). With the arrival of simple and inexpensive methods, what was an undergraduate or post-16 practical exercise moved down the age range and even into the primary classroom (Assinder, 1998).
The most commonly used (if not, for obvious reasons, the most popular) method of extracting DNA requires little more than onions, household detergent and salty water. This method became widespread in the UK due to the practical workshops and publications of the National Centre for Biotechnology Education (NCBE, 1991; 1993). More sophisticated methods of extracting DNA from cress and dried peas were developed by Science and Plants for Schools and the Australian CSIRO education centre 'The Green Machine' (NCBE, 2001), but their inclusion of DNA gel electrophoresis restricted these protocols to use by older students.
In the search for sweeter-smelling alternatives to onions, several have suggested applying the 'onion method' to a variety of fruits, including kiwi fruit, bananas and strawberries. Although these fruits seem to yield copious amounts of DNA, the 'DNA' produced is in fact little more than pectin. This can be demonstrated simply by adding pectinase to the preparation (it's also quite easy to precipitate pectin in alcohol, an old jam-maker's trick).
The protocol described here uses frozen peas. This has several advantages over the traditional onion method. Firstly, no blender is needed to break up the plant tissue. Provided the peas have thawed, they can be squashed with the back of a spoon or a glass rod. Secondly, supplies of peas can be stored easily in the freezer and taken out in suitable amounts when required. And last, but not least, the peas don't smell! Isolating the DNA (and RNA) takes about 35 minutes, including an incubation period of 15 minutes.
The ethanol must be ice cold. Place it in a plastic bottle in a freezer at least 24 hours before you attempt this activity. Please read the safety note, below.
A hook for recovering the DNA can be made by briefly heating the tip of a Pasteur pipette in a Bunsen burner flame, then bending the tip round before allowing the glass to cool. To electrophorese the DNA extract, simply dissolve some of it in about 0.5 ml of bromophenol blue loading dye, then load about 20 µl into a well in a 1% agarose gel. Staining with 0.04% (w/v) Azure A solution after electrophoresis will reveal the nucleic acids. RNA shows up a lighter pink colour (Madden, 2000).
Variations of this extraction procedure can be used for other food items, such as fish sperm (milt or soft roe) or fish eggs (Strömberg, 2001). Several publications refer to the use of calf thymus tissue, but its use in schools is no longer recommended (see safety note, below).
Most freezers are not spark-proof. Consequently, you must ensure that ethanol is placed in the freezer in a sealed, vapour-tight container. An alternative to using a freezer is to stand the sealed bottle of ethanol in ice for several hours before use. For more information about safety in schools when working with DNA, teachers in the UK should consult Topics in Safety (Delpech & Madden, 2001).
Since the discovery of bovine spongiform encephalopathy and variant Creuzfeldt-Jakob disease in the UK, UK school safety authorities advise that calf thymus should no longer be used in schools, as there is a risk (albeit small) of accidental exposure to the infectious agent while the extract is being prepared.
Most of the items required for this procedure can be obtained from a supermarket.
Novozymes Neutrase can be bought in small volumes from the UK's National Centre for Biotechnology Education.
This article first appeared in School Science Review in March 2003.
Strömberg E (2001) DNA from 'caviar'. Bioscience Explained 1(1)