Cytological Study on Induced Apospory in Ferns
Total Page：16
File Type：pdf, Size：1020Kb
1962 79 Cytological Study on Induced Apospory in Ferns Chihiro Takahashi Biological Laboratory, Department of General Education, Nagoya University, Mizuho-ku, Nagoya, Japan Received September 20, 1961 Introduction The life cycle of the pteridophyte is composed of two generations, the sexual generation and the asexual generation, which are separated on different individuals called respectively the gametophyte and the sporophyte which succeed each other in the regular order. In brief there can be seen the typical alternation of generations in the pteridophyte. The sporophyte is a diploid organism producing the spore, the starting cell of the gametophyte, by the meiosis and is larger in size, more complicated and advanced in form and structure. On the other hand the gametophyte is a haploid organism producing the gametes which by the syngamy give rise to the zygote, the starting cell of the sporophyte and is smaller in size, simpler and more reduced in form and structure but is autotrophic like the sporophyte. How different in form, structure and function are the two generations which develop from an initial single cell, the spore or the zygote, having the identical hereditary material apart from a set of chromosomes or two sets of those! How does the difference between the two generations come to be as it is? The alternation of generations is usually accompanied by that of the nuclear phases. Does the former result necessarily from the latter? In other words, is the haploidy caused by the meiosis responsible for the gametophytic characters and is the diploidy caused by the syngamy responsible for the sporophytic characters? It seems that they are not definitely responsible at least because it is well known that there are the short cuts in the life cycle named the apospory and the apogamy which occur spontaneously or are experimentally induced. The apospory is the phenomenon by which the gametophyte takes place from the sporophyte without the intervention of normally formed spore. The apogamy is the phenomenon by which the sporophyte takes place from the gametophyte without the intervention of a sexual process. Thus the alternation of generations by the apogamy or the apospory is not accompanied by that of the nuclear phases. It is expected that these deviations from the normal life cycle provide an important clue to studying the alternation of generations not only in the pteridophyte but also in plants in general. The pteridophyte is a good material for extending the work along this line. Its haploid, sexual generation is inconspicuous but free-living. So the culture is easily and abundantly 80 C. Takahashi Cytologia 27 carried out in the laboratory. The diploid, asexual generation also is easy to be obtained from the gametophytic culture and to be reared at the younger stage in the laboratory. The study was undertaken in order to bring about a better understanding of the life cycle of the pteridophyte. The cytological and morphological sides of such study by means of induced apospory will be reported in the present paper. Since apospory was found, many data about this interesting phenomenon have been accumulated. Nevertheless it seems that more detailed and comprehensive study is necessary. It will deal with problems as follows: what method is favourable for inducing apos pory?; what organs, tissues and cells are involved in the apospory?; by what process is the apospory induced?; how does the aposporous prothallium behave about producing the sporophyte?; how are the morphology and the physiology of the aposporous prothallium and the tetraploid sproophyte as compared with those of the normal prothallium and sporophyte? Material and methods Two species of ferns, Pteridium aquilinum (L.) Kuhn var. latiusculum (Desv.) Und. and Dryopteris erythrosora (Eat.) O. Ktze. were used in this study. By the present writer the apospory was successfully induced in both species. But the extensive study was carried out exclusively on the former, Pteridium aquilinum var. latiusculum. The reasons were as follows: the former grows more rapidly; the abundant aposporous outgrowth occurs more easily. The spore of Pteridium aquilinum var. latiusculum reaches its maturity in September and that of Dryopteris erythrosora in July in the vicinity of Nagoya. The leaves with the mature sporangia were cut in the habitat and brought into the laboratory where they were dried on the paper. The sporangia soon caused the dehiscence and released the spores. The spores were collected, sifted from the impurities and stored in the desiccator until they were taken out of it and sown on the culture medium. The spores were sown in the Petri-dishes of six centimeters in diameter on the culture medium which contained 20 per cent dilute Knop's solution and was solidified with 1 to 2 per cent agar. The pH was 5.2 or so. The conditions under which prothallia were reared were either the room tem perature and the dispersed light in the laboratory or the continuous illumina tion of the fluorescent lamp and the temperature adjusted at 20•‹ to 25•Ž in the incubator. The culture brought about the same results except the growth duration under both conditions. The culture was discarded, when it was badly damaged by the contamination of algae, fungi or bacteria . The spore germination takes place several days after sowing . Then a protonema, a filamentous prothallium, is formed as a result of the one-dimen sion growth. An apical portion of the protomena begins growing two - dimensionally to turn into a spatulate prothallium where a meristematic region 1962 Cytological Study on Induced Apospory in Ferns 81 is established and forms an apical notch. Now the gametophyte takes the form of a cordate prothallium and after this, of a bow-like prothallium. They have a three-dimensionally built midrib and only there the archegonia are formed. The antheridium can be formed at any developmental stage of the gametophyte, even on the basal cell without the protonema formation just after the germination (Takahashi, unpublished). The successive form change of the prothallium with its growth is attributed to the establishment and the activity of the meristematic region by which new cells are produced mainly perpendicularly to an apico-basal axis. Accordingly the value of a ratio of the prothallium width to the prothallium length decreases which results in the form change of the prothallium. If there is no syngamy on the cordate or the bow-like prothallium, the apical notch will usually change into the multicell-layered, somewhat flattened cylindrical projection which has not the wings, or has the narrow and rippled ones and continues to grow at the tip. The projection bears a large number of archegonia and rhizoids on the ventral surface and sometimes on the dorsal surface too. This struc ture resembles that reported by Lang (1898) and Mottier (1927, 1931). They observed that the apogamous phenomena occurred in connection with the projecting growth, but the present writer did not. The embryo arises sexually on the cordate and the bow-like prothallia and on such projecting prothallia as well. The sporophyte thrives normally under the present condi tion. No apogamy was found through the course of this study. The young sporophyte with leaves and roots is established a month or two after the beginning of the spore culture. Such early leaves as the first, second and third leaves were usually used in this study, with the exception of the case for the special purpose as mentioned later. The leaves were detached near their base and cultured in the Petri-dishes with 20 per cent dilute Knop's solution. A large number of aposporous outgrowths have been got by this method. Since the aposporous prothallium was induced and grown in the Petri-dish with a lid, there was no possibility of contamina tion by fern spores of the same and other species. In order to avoid the excessive contamination of algae, fungi or bacteria it is better to use the liquid medium than to use the agar medium. It was proved that the apos porous outgrowth occurred also in the aseptic culture. But it was not necessary to culture the sporophytic organ under the sterile condition so far as the present study was concerned. The culture was kept either in the incubator adjusted at 20•‹ to 25•Ž of which the walls were made of glass through which the dispersed light streamed in, or in the laboratory under the room temperature and the dispersed light. Some months after the begin ning of the culture the aposporous outgrowth can be noticed on most of the detached leaves. The aposporous prothallium was cultured as it was attached to, or detached from, the sporophytic organ. In either case the aposporous pro- Cytologia 27, 1962 6 82 C. Takahashi Cytologia 27 thallium gave rise sexually to a new sporophyte. Such a sporophyte was planted in a sphagnum-packed pot, when the writer intended to develop it further. Results Attempts were made to induce apospory in Pteridium aquilinum var. latiusculum by methods, such as: injuring or damaging with tweezers the undeveloped leaves as they were attached to the sporophyte, which later gave rise to various types of the malformed leaves; subjecting the very young sporophyte to starvation by providing water without nutrition under the weak light, resulted in the failure of developing the adult type of leaves; culturing the detached leaves or roots in the nutrient solution. It was re vealed that only the last method was successful in the early leaves and roots. The sporophyte usually raises its leaves freely in the air, but it happened that the leaves, especially the first ones, grew diving into the agar or the solution, or in contact with the agar surface. However in these cases apos pory did not occur.