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The low molecular weight fraction of compounds released from immature wheat pistils supports barley pollen embryogenesis.

Lippmann R, Friedel S, Mock HP, Kumlehn J - Front Plant Sci (2015)

Bottom Line: Here, a micro-culture system was established to enable the culturing of populations of barley pollen at a density too low to allow unaided embryogenesis to occur, and this was then exploited to assess the effect of using various parts of the pistil as nurse tissue.The differential effect of various size classes of compounds present in the pistil-conditioned medium showed that the relevant molecule(s) was of molecular weight below 3 kDa.This work narrows down possible feeder molecules to lower molecular weight compounds and showed that the cellular origin of the active compound(s) is not specific to any tested part of the pistil.

View Article: PubMed Central - PubMed

Affiliation: Leibniz Institute of Plant Genetics and Crop Plant Research (IPK) Gatersleben, Germany.

ABSTRACT
Pollen embryogenesis provides a useful means of generating haploid plants for plant breeding and basic research. Although it is well-established that the efficacy of the process can be enhanced by the provision of immature pistils as a nurse tissue, the origin and compound class of the signal molecule(s) involved is still elusive. Here, a micro-culture system was established to enable the culturing of populations of barley pollen at a density too low to allow unaided embryogenesis to occur, and this was then exploited to assess the effect of using various parts of the pistil as nurse tissue. A five-fold increase in the number of embryogenic calli formed was obtained by simply cutting the pistils in half. The effectiveness of the pistil-conditioned medium was transitory, since it needed replacement at least every 4 days to measurably ensure embryogenic development. The differential effect of various size classes of compounds present in the pistil-conditioned medium showed that the relevant molecule(s) was of molecular weight below 3 kDa. This work narrows down possible feeder molecules to lower molecular weight compounds and showed that the cellular origin of the active compound(s) is not specific to any tested part of the pistil. Furthermore, the increased recovery of calli during treatment with cut pistils may provide a useful tool for plant breeders and researchers using haploid technology in barley and other plant species.

No MeSH data available.


Related in: MedlinePlus

The effect of varying the number of co-cultivated, longitudinally bisected pistils on embryogenic pollen development. Isolated microspores at an initial density of 75 per mL were co-cultivated with various numbers of bisected wheat pistils. The microspores were kept within Millicell inserts to facilitate the observation of their response at low culture density and without being obscured by nurse tissue. The inserts were positioned in 3.5 cm Petri dishes containing a total volume of 2 mL medium, while the nurse tissue was kept in the medium portion outside the inserts. The diagrams show the formation of embryogenic calli of diameter (A) ≥100 μm and (B) ≥500 μm after 4 weeks of culture. EPC, embryogenic pollen culture of cv. “Igri” precultured for 1–2 weeks and used as nurse tissue at a density of 50,000 pollen grains per mL; lP, longitudinally bisected pistil (numbers of bisected pistils co-cultivated per mL medium are given). The red horizontal lines represent the responses of respective cultures in the absence of nurse tissue.
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Figure 3: The effect of varying the number of co-cultivated, longitudinally bisected pistils on embryogenic pollen development. Isolated microspores at an initial density of 75 per mL were co-cultivated with various numbers of bisected wheat pistils. The microspores were kept within Millicell inserts to facilitate the observation of their response at low culture density and without being obscured by nurse tissue. The inserts were positioned in 3.5 cm Petri dishes containing a total volume of 2 mL medium, while the nurse tissue was kept in the medium portion outside the inserts. The diagrams show the formation of embryogenic calli of diameter (A) ≥100 μm and (B) ≥500 μm after 4 weeks of culture. EPC, embryogenic pollen culture of cv. “Igri” precultured for 1–2 weeks and used as nurse tissue at a density of 50,000 pollen grains per mL; lP, longitudinally bisected pistil (numbers of bisected pistils co-cultivated per mL medium are given). The red horizontal lines represent the responses of respective cultures in the absence of nurse tissue.

Mentions: Since the presence as a nurse tissue of longitudinally bisected pistils gave the strongest stimulatory effect on embryogenic pollen development, an attempt was then made to optimize the number of pistils to be used. To this end, low-density cultures (75 isolated microspores of cv. “Igri” per mL medium) involving the co-cultivation of 0.5–6 bisected pistils and EPCs at standard density as positive control were compared. The inclusion of between one and six longitudinally bisected pistils per mL medium was the most effective treatment (Figure 3; raw data given in Supplemental Table S2). The median values of callus formation ≥500 μm in size increased step-wise with the number of co-cultivated pistil halves, while the use of only half a pistil per mL resulted in a significantly reduced pollen-derived callus formation as compared with all treatments involving one to six bisected pistils. The response across the tested range of numbers of pistils added to the cultures indicated that just one bisected pistil is capable of almost entirely satisfying the demand for stimulating signal molecules under the given circumstances.


The low molecular weight fraction of compounds released from immature wheat pistils supports barley pollen embryogenesis.

Lippmann R, Friedel S, Mock HP, Kumlehn J - Front Plant Sci (2015)

The effect of varying the number of co-cultivated, longitudinally bisected pistils on embryogenic pollen development. Isolated microspores at an initial density of 75 per mL were co-cultivated with various numbers of bisected wheat pistils. The microspores were kept within Millicell inserts to facilitate the observation of their response at low culture density and without being obscured by nurse tissue. The inserts were positioned in 3.5 cm Petri dishes containing a total volume of 2 mL medium, while the nurse tissue was kept in the medium portion outside the inserts. The diagrams show the formation of embryogenic calli of diameter (A) ≥100 μm and (B) ≥500 μm after 4 weeks of culture. EPC, embryogenic pollen culture of cv. “Igri” precultured for 1–2 weeks and used as nurse tissue at a density of 50,000 pollen grains per mL; lP, longitudinally bisected pistil (numbers of bisected pistils co-cultivated per mL medium are given). The red horizontal lines represent the responses of respective cultures in the absence of nurse tissue.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4493395&req=5

Figure 3: The effect of varying the number of co-cultivated, longitudinally bisected pistils on embryogenic pollen development. Isolated microspores at an initial density of 75 per mL were co-cultivated with various numbers of bisected wheat pistils. The microspores were kept within Millicell inserts to facilitate the observation of their response at low culture density and without being obscured by nurse tissue. The inserts were positioned in 3.5 cm Petri dishes containing a total volume of 2 mL medium, while the nurse tissue was kept in the medium portion outside the inserts. The diagrams show the formation of embryogenic calli of diameter (A) ≥100 μm and (B) ≥500 μm after 4 weeks of culture. EPC, embryogenic pollen culture of cv. “Igri” precultured for 1–2 weeks and used as nurse tissue at a density of 50,000 pollen grains per mL; lP, longitudinally bisected pistil (numbers of bisected pistils co-cultivated per mL medium are given). The red horizontal lines represent the responses of respective cultures in the absence of nurse tissue.
Mentions: Since the presence as a nurse tissue of longitudinally bisected pistils gave the strongest stimulatory effect on embryogenic pollen development, an attempt was then made to optimize the number of pistils to be used. To this end, low-density cultures (75 isolated microspores of cv. “Igri” per mL medium) involving the co-cultivation of 0.5–6 bisected pistils and EPCs at standard density as positive control were compared. The inclusion of between one and six longitudinally bisected pistils per mL medium was the most effective treatment (Figure 3; raw data given in Supplemental Table S2). The median values of callus formation ≥500 μm in size increased step-wise with the number of co-cultivated pistil halves, while the use of only half a pistil per mL resulted in a significantly reduced pollen-derived callus formation as compared with all treatments involving one to six bisected pistils. The response across the tested range of numbers of pistils added to the cultures indicated that just one bisected pistil is capable of almost entirely satisfying the demand for stimulating signal molecules under the given circumstances.

Bottom Line: Here, a micro-culture system was established to enable the culturing of populations of barley pollen at a density too low to allow unaided embryogenesis to occur, and this was then exploited to assess the effect of using various parts of the pistil as nurse tissue.The differential effect of various size classes of compounds present in the pistil-conditioned medium showed that the relevant molecule(s) was of molecular weight below 3 kDa.This work narrows down possible feeder molecules to lower molecular weight compounds and showed that the cellular origin of the active compound(s) is not specific to any tested part of the pistil.

View Article: PubMed Central - PubMed

Affiliation: Leibniz Institute of Plant Genetics and Crop Plant Research (IPK) Gatersleben, Germany.

ABSTRACT
Pollen embryogenesis provides a useful means of generating haploid plants for plant breeding and basic research. Although it is well-established that the efficacy of the process can be enhanced by the provision of immature pistils as a nurse tissue, the origin and compound class of the signal molecule(s) involved is still elusive. Here, a micro-culture system was established to enable the culturing of populations of barley pollen at a density too low to allow unaided embryogenesis to occur, and this was then exploited to assess the effect of using various parts of the pistil as nurse tissue. A five-fold increase in the number of embryogenic calli formed was obtained by simply cutting the pistils in half. The effectiveness of the pistil-conditioned medium was transitory, since it needed replacement at least every 4 days to measurably ensure embryogenic development. The differential effect of various size classes of compounds present in the pistil-conditioned medium showed that the relevant molecule(s) was of molecular weight below 3 kDa. This work narrows down possible feeder molecules to lower molecular weight compounds and showed that the cellular origin of the active compound(s) is not specific to any tested part of the pistil. Furthermore, the increased recovery of calli during treatment with cut pistils may provide a useful tool for plant breeders and researchers using haploid technology in barley and other plant species.

No MeSH data available.


Related in: MedlinePlus