No. 4465 May 28, 1955 NATURE 953

A Modified Method of place ; but even where this happens, loss of substance Electrophoresis through migration off the pa.per rarely occurs. The the strip prevents the re• work on the separation of 'Cellophane' covering IN . the co~se of our on the filter paper. Protein fractions from wheat gluten by meana of depositing of moisture protem by this method a.re sharp and as well p_aper el~ctrophoresis, it was found that good resolu• spots obtained spots obtained in electrophoresis between tion of hitherto unresolved fractions could be achieved defined as glass plates. paper electrophoresis using long pa.per strips and· bf full results of this investigation will be voltages over an extended period of time. This The high ed elsewhere. does not require an organic cooling liquid, publish method H. ZENTNER 1• It may be useful for such as monochlorobenzene Institute of Australia, of enzyme mixtures, where contact Bread Research the separation 111 Pacific Highway, organic liquid may inactivate the enzymes. with North Sydney, N.S.W. Jan. 6. !n this :i_nethod, the filter paper strip is wrapped, 1069 (1962). sp1:al fash10n, a~ound a glass tube, 22 in. long and 'Consden, R., Nruurt, 170, ¾ m. external diameter, through which cold water is nmning (Fig. 1). To minimize evaporation from the strip, it is covered with a st.cip of thin 'Cellophane' Isolation of Fungi from Hyphae present of tlie same width. Hwnid conditions are maintained in Soil glass tube in a wooden by enclosing a portion of the it is known that fungi may exist in soil with filter paper saturated with WHILE box, which is lined both as active mycelium avd as dormant spores', a of the filter paper, covered by buffer. The ends major problem in the study of soil fungi has been and out of the hwnid chamber 'Cellophane', pass in discover which fungi i:n a soil are present as slits at the bottom, ¾ in. to by way of two narrow lium. Plating methods by which fungal colonies sleeves, attached t o myce from each end. 'Cellophane' are isolated from soil• or from soil suspensions• on terminating below the level of the chamber and agar media suffer from the disadvantage that the prevent the drying out of these the buffer solutions, unit of origin, spore or hypha, of the resulting colonies strips. portions of the is not known. Direct microscopic examination, either of Rossi-Cholodny slides or soil films', has the com• plementary deficiency that although fungal hyphre are seen, most are sterile and their identity cannot be investigated. I have devised a simple technique for the isolation of fungi directly from hyphre present in soil. Essentially the method depends on the observation that when a soil suspension is prepared, many of the fungal hyphre remain with the heavier soil particles of the residue. Removal of the fine suspended material from the residue also permit s Flg. 1 visual examination of the latter for the presence of which may then Before use, the glass tube is treated with silicone individual hyphro or hyphal masses, media. grease. The filter paper strip is then passed through be removed and grown on agar ·5 gm. is placed in a one sleeve, to which it is clamped with a paper clip, A soil crumb of about l ·0-1 and left to become and wrapped tightly around the glass tube, passing beaker part-filled with water the crumb is broken apart out through the sleeve at the other end and clamped saturated. After 4--5 min. a r apid jet of tap water. fast. To permit wrapping at an angle, the 'Cellophane' by filling the beaker with are allowed to sediment sleeves-through which the paper strips pass-are The heavier soil particles most of the suspension is slit along the outside edge leaving a short uncut por• for l ·0-1 ·5 min. and then is _added, the heavier adjoining the box. At one end, together with poured off. Further water tion and the suspension the filter paper strip, a 'Cellophane' strip of the same particles allowed to sediment procedure is continued until widt.h as the filter paper is clamped in. The point of again removed. This standing for l ·0-1 ·5 min. application for the protein mixture is then marked on the liquid remains clear after residue are then distributed the paper and the buffer painted on with a brush. The soil particles of the water among three sterile This procedure prevents tearing and bruising of the in a small quantity of for the presence of fungal paper, which may occur when a buffer-moist, blotted Petri plates and searched dissecting strip is wrapped tightly around the glass tube. The hyphre using a binocular Individual hyphre or portions substance is then applied to the marked line and the ( x 12 ·5 and x 25). removed with fine forceps from 'Cellophane' strip wrapped tightly so as to cover the of hyphal masses are and a.re placed in a drop of whole length of the strip and to pass out through among the soil pa rticles sterile isolation plate. second sleeve. The 'Cellophane' sleeves, filter sterile water on a dean the isolat ed from soil are and 'Cellophane' strip are then pierced and Since many fungal hyphre paper or to hwnus particles on with a piece of bent and the paper attached to mineral grains weighted fungi and other micro• clips removed. The buffer vessels and the carbon or in which contaminating be present, it is desirable to remove electrodes are then placed in position below the organisms may of these attached particles. Many chamber. as many as possible by gently tapping the hypha In our experiments, strips of Whatman 3MM may be removed of the plate ; others cannot be paper, 60 cm. long and 2 ·5 cm. wide, were used. against the bottom killing the hypha . Hyphal masses For a run of 24 hr. with a potential gradient of 9 V./ removed without to be teased out, s ince they usually cm., 2 x 2 litres of l\follvaine's buffer pH 2 ·07 and may also need than one species of fungus. When ionic strength of 0 ·025 were used. Unless high consist of more (usually 20-50) are on the plate, it voltages are used (17 V./cm.), no condensation of sufficient hyphre elted but cooled agar anrl the hyphro water on the exposed surface of the glass tube takes is poured with m

© 1955 Nature Publishing Group