S. C. Maheshwari

Haploids from Pollen Grains -- Retrospect and Prospect

The production of large numbers of haploid plants from pollen grains in aseptic culture of anthers or isolated pollen grains has now been demonstrated in many angiosperms. It is a technique which holds much promise for creating desired variability by mutations for biochemical and applied genetics and for producing pure lines for crop improvement. This paper reviews the research on the production of haploids from pollen grains. The response is dependent on a variety of factors such as media formulation (especially the hormone component), genotype and physiological status of the donor plant, developmental stage of the pollen grain, anther culture environment and anther wall itself-all of which influence greatly the successful production of haploid plants. Although still in its infancy, the culture of isolated pollen is seen as a promising line of further research as it has the potential of offering several advantages over the culture of whole anthers.

Induction of haploidy from pollen grains in angiosperms - the current status.

SummarySince the successful induction of haploids from anthers cultured in vitro in 1964, a great deal of attention has been given to this problem by those interested in obtaining pure lines and mutants for crop improvement and biochemical genetics. In the last 16 years the anther culture technique has been refined and extended to over one hundred and fifty different species. More recently, isolated pollen culture — which is a refinement of the original anther culture technique — has also been developed. In this review we have made an effort to critically examine existing reports with the objective of analysing the effects of various factors — e.g. culture medium, the cultural conditions, and the effect of genotype and physiological state of the parent plant on pollen induction — and to speculate on the mechanism of action of different factors in order to throw some light on the process of haploid induction.

Effect of l-proline and l-tryptophan on somatic embryogenesis and plantlet regeneration of rice (Oryza sativa L. cv. Pusa 169)

Here we report our assessment of the promotive effect of proline and tryptophan on the frequency of callusing and regeneration as also on the absolute number of plantlets regenerated per mature seed used for callus initiation. Finally, scanning electron microscopic evidence for regeneration via somatic embryogenesis in seed cultures is also given

In vitro differentiation of plantlets from tissue cultures of Albizzia lebbeck L.

Attempts at inducing differentiation in various explants of Albizzia lebbeck resulted in the production of abundant shoot buds from the hypocotyl, root, cotyledon and leaflet explants, both directly and indirectly (i.e. without and with the intervention of callus formation). Rooting was achieved on transfer of the shoots to BM +2 mg/1 IAA after some growth. The plants could be successfully transferred to soil, providing a method for mass propagation of this important leguminous tree species.

Sodium chloride resistant cell line from haploidDatura innoxia mill

SummaryA cell line resistant to sodium chloride was selected from callus cultures of haploidDatura innoxia by cloning under selective pressure. Cells of the resistant cell line retained their resistance even after subculture in absence of NaCl. Plantlets could be regenerated from resistant cells in the presence as well as absence of NaCl. In contrast, regeneration of plantlets was not possible from normal cells in the presence of NaCl, although regeneration readily occurred in the absence of NaCl.To examine the stability of the resistance in the long-term, callus cultures were initiated in presence of NaCl from stem expiants of the differentiated plantlets. All expiants of plantlets derived from resistant cells showed callus formation. This callus, derived from resistant explants, retained the trait of resistance upon subculture.

High frequency production of embryos inDatura innoxia from isolated pollen grains by combined cold treatment and serial culture of anthers in liquid medium

SummaryThis study concerns the development of pollen embryos as affected by various physical conditions of culture in media devoid of hormones. Freshly isolated pollen, from anthers ofDatura, failed to form embryos regardless of whether they were cultured on liquid or solid medium. In contrast, pollen isolated from anthers precultured on solid medium did form embryos and the response could be increased by prior cold treatment of anthers at 4 °C for 4 days. However, the best results were obtained when anthers were cultured from the very beginning in liquid medium and transferred serially to fresh medium. Under such conditions, the anthers dehisced, allowing spontaneous shedding of pollen grains. It was thus possible to have several fractions of shed pollen continuing their development into embryos. When serial culture was started with anthers from cold-treated buds not only were embryos formed in all the fractions of shed pollen but the frequency was also considerably higher than in any mode of culturing.

Studies on the Growth and Flowering of a Short-Day Plant, Wolffia microscopica II. Role of Metal Ions and Chelates*

SummaryAs found earlier, supply of EDTA was obligatory for both flowering and satisfactory vegetative growth in Wolffia microscopica. It is now shown that the metal affecting growth and flowering is most probably iron. Omission of Fe but not of Cu, Zn, Mn and B from the medium markedly affects vegetative growth. There exists also a strong interaction between EDTA and Fe, one being largely inactive in the absence of the other. When Fe-EDDHA is substituted for Fe-citrate and EDTA in the medium, no great effect is seen in vegetative growth, but flowering takes place even under continuous light. Studies with 59Fe show that, in the medium containing Fe-EDDHA, Fe uptake is stimulated several-fold; this is apparently associated with the flowering condition.

Isolation and culture of protoplasts of Capsicum annuum L. and their regeneration into plants flowering in vitro

SummaryAxenic shoot cultures ofCapsicum annuum cv.California Wonder were used as the source for isolation of protoplasts from mesophyll cells. Protoplasts underwent sustained mitotic activity and proliferated to form callus masses on NT or DPD medium enriched with 2,4-D, NAA and BAP each at 1 mg/l level. The callus could be differentiated into whole plants on the differentiation media and plants floweredin vitro under long day conditions.

Differentiation in explants from mature leguminous trees

Stem and petiole explants, obtained from mature trees, ofAlbizzia lebbeck,Cassia fistula andC.siamea callused and differentiated shoot-buds and later shoots on B5 medium supplemented with either 0.5 mg/l IAA + 1 mg/l BAP or BM + 2 mg/l NAA + 0.5 mg/l BAP. The stem explants were more responsive than the petiole explants. InA.lebbeck, the IAA substituted medium favoured differentiation from both types of explants. However, inC.fistula, the type of explants rather than the medium composition had an overriding influence on shoot differentiation since those from petiole hardly responded in either medium. It has been possible to obtain plantlets from bothA.lebbeck andC.fistula under conditions conducive to rooting. Plantlets ofA.lebbeck have also been successfully transferred to the field.

Transient expression of gus gene in intact seed embryos of Indica rice after electroporation-mediated gene delivery

SummaryTwo-day-old germinating intact seed embryos of Oryza sativa variety Basmati 370 were electroporated with a view to examine suitability of this system for gene delivery. The experiments were done with a plasmid having gus gene under the control of CaMV 35S promoter. Spectrofluorophotometric GUS assay revealed high activity of the introduced gene when embryos were given three electrical pulses at 1600 V cm-1 and 100 μF capacitance with a pulse length of 75 ms. Additionally, histochemical localization of GUS activity in seedlings and various organs such as leaves, coleoptiles and roots was also done. Expression of GUS activity was studied up to 15 days and found to be organ-specific, thereby showing that embryos can indeed serve as efficient recipient system. Use of cycloheximide revealed that GUS activity appears as a result of early protein synthesis after electroporation and is substantially stable in vivo.