Using Water Clover to Demonstrate Sexual Reproduction in Ferns

Using Water Clover to Demonstrate Sexual Reproductionin Ferns

WilburJ. Settle Downloaded from http://online.ucpress.edu/abt/article-pdf/42/5/295/38413/4446943.pdf by guest on 23 September 2021 INMOST HIGHERPLANTS, including ferns, sexual reproduction from the time of spore formation to the ap- TABLE 1. Directionsfor "Germination"of Sporocarps. pearance of a young sporophyte requires several weeks. Using the usual fern spores, such as those from Maiden 1. Cut off the tip of one end of two sporocarps (use two in case one fails to "germinate") Hair Fern (Adiantum) is generally satisfactory, but planning and luck are required to have gametophytes 2. Place the cut sporocarps in a petri dish containing tap water. at the desired stage for the study of sexual reproduction 3. Cover the petri dish with a lid and mark the date and time in a particularlaboratory session. on the top. For the past few years, I have used Water Clover 4. Leave the petri dish in the light at room temperature. (Marsilea),an aquatic or marsh fern with an unfernlike appearance, to demonstrate sexual reproduction in living specimens in a laboratory course focusing on vas- sporangium is a transparent sac containing several micro - cular plants. spores. Soon after the sori emerge from the sporocarp, Water Clover produces spores within its small, hard, megaspores and microspores are released from the bean-shaped structure known as a sporocarp (fig. 1), sporangia, and they begin to separate and float away (fig. which is considered a modified leaf or leaflet (Bierhorst 3). Both microspores and megaspores are large enough 1971). To begin the process, scarifythe sporocarp by cut- to be seen with the naked eye. ting or breaking to admit water. The easiest and most ef- Each sporocarp contains two types of spores: (1) the fective method is to cut off the very tip (approximately smaller microspore that develops into a male gameto- 1/2 mm) of the rounded end of the sporocarp with a sharp phyte (microgametophyte);and (2) the larger megaspore razor blade. Then place the sporocarp in tap water in a receptacle such as a petri dish and keep it at room temperature. The steps required to "germinate"sporocarps are listed in table 1. Shortly after the sporocarp is cut and placed in water, usually within one-half to one hour, it breaks open ("ger- minates") and a gelatinous ring with white cigar-shaped clusters of sporangia (sori)emerges but often remains at-- tached to the sporocarp (fig. 2). Each megasporagium consists of a single, large megaspore, and each micro-

WilburJ. Settle (photonot available)is assistantprofessor in the depart- ment of biology,State UniversityCollege, Oneonta, New York 13820, a positionhe has heldsince 1970. He teachescourses in generalbotany, planttaxonomy, cytology, vascular plants, and economicbotany. Settle receivedhis A.B.degree in biologyfrom Centre College (Kentucky)in 1962, and his M.S.and Ph.D.degrees in botanyfrom Ohio State Uni- versityin 1965 and 1969 respectively.In the summerof 1971, Settle held a facultyresearch fellowship from the ResearchFoundation of the State Universityof New York.He has publishedarticles on Silphium integrifoliumin the Ohio Journal of Science, and his current research centerson a biosystematicstudy of the genus Blephila.Settle holds membershipsin AAAS,the BotanicalSociety of America,and Sigma FIGURE1. Unopenedsporocarp of WaterClover. The distancebe- Xi. tweenthe black lines at the bottom is 1mm.

WATER CLOVER 295 FIGURE 2. Sporocarp one hour after being scarified and placed in water. The dark structure at the right is the opened sporocarp, showing the two halves. Downloaded from http://online.ucpress.edu/abt/article-pdf/42/5/295/38413/4446943.pdf by guest on 23 September 2021

that develops into a female gametophyte(megagameto- and easily obtained, these sperm are excellent material phyte). Each sporocarpcontains many megasporesand for demonstratingthe morphology of male gametes. even moremicrospores. After fertilizationoccurs, a young sporophyte (embryo) An unusualfeature of this fern is the rapid develop- is large enough to be seen macroscopically in only a few ment of the spores into gametophytes.This rapiddevel- days (fig. 4). With a dissecting microscope or hand lens opmentis the resultof the formationof a much reduced a small green leaf, a small white root, and rhizoids can be gametophyteconsisting of only a few cells. Spermare re- seen three days after the sporocarp was opened. leased from male gametophytesof Marsileacrenata in The development from small sporophyte (embryo) to five hours and from Marsileaquadrifolia in eight or nine a plant a few inches high requires several weeks. If the hours (Bierhorst1971). Femalegametophytes have ma- plants are to be kept to maturity, they should be trans- ture archegoniaformed in fifteenhours in two species of ferred to soil that is kept constantly moist; a terrarium Marsilea(Bierhorst 1971). makes an ideal place for growing this fern. We maintain a Each microsporerapidly undergoes a few mitoticdi- large terrarium with wet soil in our greenhouse where visions resultingin the formationof sixteen corkscrew- Water Clover, small club moss, and ferns thrive. Though shaped, multiflagellatesperm (Bierhorst1971). Each it is an interesting and unusual type of fern, instructors megasporeundergoes a few mitoticdivisions forming a may not want to allow the plants to reach this point be- small archegoniumwhose neck at maturityconsists of cause of time and/or space limitations. only five cells (Bierhorst1971). An elaborategelatinous Any number of different ages of "germinated"sporo- structureforms around the megaspores. Althoughthe carps could be used to demonstrate the various stages of gelatinousstructure is easilyvisible, accurate details of its sexual reproduction, but at least the following three ages structurewere describedonly recently by Machlisand should be observed: (1) approximately one week after Rawitscher-Kunkel(1967). This structureappears to at- "germination"of sporocarp-a young sporophyte with tract sperm that collect in it. The sperm are unusually green leaf should be found growing from one end of each largeand can easilybe seen witha dissectingmicroscope. megaspore; (2) approximately three days after "germina- Sperm could be transferredto a compound microscope tion" of sporocarp-a very small sporophyte should be for more detailed observations.Because they are large barely visible at one end of each megaspore; (3) between

_ | 1 * 1110 | | Wg W FIGURE3. A megaspore (left) and microspore (right)of Water Clover.

~~~~~~~~~g B#E. EXN * The material is quick and easy to prepare. It is 'in- stant;"add water to the cut sporocarps. * The sporocarps, which are the only biological sup- plies required are readily available from biological supply companies and quite inexpensive; they cur- rently cost less than $5.00 per dozen. * The sporocarps can be stored for long periods. Bierhorst (1971) has confirmed that they remain alive and viable for at least 35 years. * No special growth medium is required to germinate the sporocarps;tap water works well. * The entire demonstration can be performed without the use of compound microscopes. Only a dissecting microscope or hand lens is needed. * Swimming sperm can be easily observed without a microscope. compound Downloaded from http://online.ucpress.edu/abt/article-pdf/42/5/295/38413/4446943.pdf by guest on 23 September 2021 * Young sporophytes can be kept and cultivated, serving as an example of an unusual fern.

References BIERHORST,D.W. 1971. Morphology of vascular plants. New York:Macmillan Company. MACHLIS,L., and RAWITSCHER-KUNKEL,E. 1967. The hy- drated megaspore of Marsilea vestita. Amer. J. Bot. 54(6): 689.

Temperature Control ... from p. 289 FIGURE4. Young sporophyte one week after "germination"of sporo- Those who use this game will probably want to compile carp.The top structureis the youngestleaf, the bottomaxis is the young root,and the ellipticalstructure to whichthey are attachedis a mega- their own lists of species, and invent their own situations, spore.The smallround structures in the vicinityof the sporophyteare when the cards are being made. The selection of species microspores. and preparation of Situation Cards is a useful class exer- cise and something students could profitably do outside eight and twelve hours after "germination"of sporocarp class. Tables 1 and 2 show the species and conditions we -swimming sperm should be plentiful at this stage. have used. These are offered only as examples. This demonstration provides students with an oppor- tunity to analyze the reproductive strategy employed by Evaluation this plant. Students should be able to infer what advan- This game is one of our most exciting laboratory ex- tages the rapid sexual reproduction gives to this plant and ercises. Students especially enjoy the water phase of the make inferences about the habitatwhere it could thrive. game that is played if both participants survive the first The demonstration could also be effectively used in round; it provides an entertainingchange of pace. introductory college courses such as general botany or The game also does much to supplement and drama- the plant section of general biology to illustrate sexual tize lecture material. The information a student must reproduction in plants in general and in ferns in particu- acquire about certain animals can hardly be conveyed in lar. It would be especially appropriate for the fern por- a more challenging format. Because students enjoy com- tion of advanced courses in plant morphology or taxon- peting with one another, they are more enthusiastic about omy. High school biology instructorsmight find it useful searching references to learn about the animals. Students to demonstrate some of the general principles of sexual tested on the information presented in the game retain reproduction in plants, perhaps to illustrateand to explain an estimated 75% of the material. This is well above the the concept of alternation of phases (gametophyte and overall score for all laboratoryexaminations. We consider sporophyte) in plants. the game an excellent learning experience. The steps I have described could be carried out by students either individuallyor in small groups as a laboratory Acknowledgment-Illustrationsfor this articleand the photo of on fern reproduction. However, timing of observations the authorwere drawnby ChristineMendyk. would be crucial. Some of the advantages I have identified of using Reference Water Clover to demonstrate sexual reproduction in ferns HELLER,H.C., CRAWSHAW, L.I., and HAMMEL,H.T. 1978. are the following: The thermostatof vertebrateanimals. Scien. Am. 239(2):102.

WATER CLOVER 297