A number of factors are found to favour apogamous response in ferns. Some of the factors are: 1. Carbohydrate Nutrition 2. Mineral Nutrition 3. Gene Dosage 4. Light Quality 5. Hormones.
Factor # 1. Carbohydrate Nutrition:
Dark grown cultures of P. aquilinum do not form apogamous sporophytes even in the presence of sucrose.
The increased light intensity inhibited the response, and did not promote apogamy at low levels of sucrose (0.25 per cent). The optimal response was obtained at 4 per cent glucose and 25 ft-C (Fig. 12.3) Changes of medium at set intervals facilitated the separation of apogamous plant development into gametophytic no detectable response., initiative, and developmental phases (Fig. 12.4).
The production of maximal number of apogamous structures required four per cent sucrose during gametophytic and initiative phases, but sugar concentration could be dropped to 0.1 per cent during developmental phase. Gibberellic acid and naphthalene acetic acid promoted apogamous response when supplied up to the initiation phase.
Induction of apogamy on a carbohydrate-enriched medium points towards the possibility of enhanced substrate contributing towards additional energy which facilitates the development of sporophytic phase from the gametophyte.
However it falls short of an explaination as to how additional carbohydrate and additional energy supports apogamous response? As for lower pteridophytes apogamy has been recorded in cultures of Lycopodium and Equisetum. Sugar does not favour apogamy in Equisetum.
To begin with, osmotic potential of the medium was described to be without an effect on apogamy. This was despite the fact that high negative osmotic potential reduced apogamous development. Subsequent studies have shown that once minimal sugar requirement is met, an increase in osmotic concentration of culture enhances apogamous response.
Factor # 2. Mineral Nutrition:
Poor mineral nutrition is not conducive to apogamy. Elimination of any one of the elements from the nutrient medium did not promote apogamy in Pteridium aquilinum, and high levels of phosphorous rather increased the response.
Factor # 3. Gene Dosage:
A correlation between apogamy and increase in chromosome number is observed in P. aquilinum. Under similar cultural and nutritive conditions the diploid gametophytes, raised through apospory, produced more apogamous sporophytes than did the haploid prothalli (Fig. 12.5).
Since the level of ploidy affects apogamous response, there may be a relationship between the level of ploidy and normal life cycle of ferns. Haploid gametophytes conform to sexual cycle and polyploids are prone towards apogamy.
Factor # 4. Light Quality:
Sugar-induced apogamy is always preceded by enhancement of prothallial growth. On medium containing four per cent sucrose, under different qualities of light, the prothalli of Pteridium exhibited differential developmental response. Under far-red light apogamy occurred without enhanced gametophytic growth. Enhanced gametophytic growth, therefore, could be separated from the response to produce apogamous outgrowth.
In dark there is little growth of gametophytes even on medium enriched with 4% sucrose and there is practically no apogamous response as compared to light-grown gametophytes.
Factor # 5. Hormones:
For apogamous development the fern gametophytes require an increased quantity of respiratory substrate which is met with increased supply of sugar in aseptic cultures. In soil cultures it is, possibly, met by high light intensity. However, besides carbohydrate nutrition hormonal influence seems to be operative in the induction of apogamy.
In Ampelopteris prolifera the response at low level of sucrose (0.5 per cent) in the presence of an auxin (1-2 ppm, 2, 4-D) was of the same order as that of higher concentration (4.0 per cent) of sucrose.
Similarly, tryptophan markedly promoted the differentiation of sporophytes in dark grown gametophytic calli of P vittata on a medium enriched with sucrose. Of interest in this context are the findings on the role of ethylene in the induction of apogamy in Pteridium gametophytes.
The apogamous response increased with an increase in the number of gametophytic colonies in a culture vessel, and also with the reduction in the volume of culture vessel. This indicated the involvement of a gaseous substance.
The production of ethylene by gametophytes was demonstrated by gas chromatography, and ethylene supplied in a continuous flow system promoted the formation of apogamous plants over that of air control (Fig. 12.6). Placing of mercuric perchlorate, an effective absorber of ethylene, reduced the response to a very low level (Fig. 12.7).
The role of ethylene was substantiated in experiments employing carbohydrate and carbon dioxide, a competitive inhibitor of ethylene. Ethylene in absence of carbohydrate was ineffective in promotion of apogamy. Frequent evacuation of cultures to reduce level of ethylene reduced apogamous response.
In another experiment apogamy was inhibited, without an affect on gametophytic growth, on flushing the cultures with 7% CO2, However, when a low level (1 ml/L) of ethylene was given there was some response.
These results revealed that along with carbohydrate, ethylene has a role in apogamy and a certain ‘threshold’ level of ethylene is required for the response. Removal of endogenous ethylene by evacuation, absorption by mercuric perchlorate, or its competitive inhibition by CO2 results in reduction of response.
The interactive role of carbohydrate and ethylene became clear in experiments which separated the response in terms of (a) induction, (b) development. For induction as well as development both ethylene and carbohydrate are required but development is possible in absence of ethylene.
Ethylene was required for induction but was effective only when supplied along with sucrose. For a significant increase in frequency of response a minimal exposure of eight days of ethylene is essential. This further increases up to 24 days.
Further experiments highlighted the correlative role of ethylene and carbohydrate in apogamous response. Neither ethylene alone nor carbohydrate alone at a low level is effective in induction of apogamy. Hence, all is not clear about the role of either ethylene or carbohydrate.