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Thread: Heliamphora hispida

  1. #33

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    I and one other grower tried applying NA, on the hypothesis that cytokinins from multiplication had caused lingering DNA methylation changes that were causing the plants to continue dividing instead of maturing. This had no effect, though only one NA concentration was tried, and only on a small number of plants:

    http://www.terraforums.com/forums/sh...light=elongata

    Your hypothesis has the virtue of being testable, and is worth a try, but it seems like the role of GA is not that straightforward AFAIK. Taiz and Zeiger reference added gibberellins causing early maturation in conifers, but note that added gibberellins cause reversion to juvenile form in Hedera helix. Neither are closely related to Heliamphora, but Hedera is significantly closer, so that's why I question adding GA as the solution. I'm the first to admit I'm no expert on plant phys--do you any references to forced maturation in herbs from adding Gibberellins? (That's not a rhetorical question, I'm actually asking, as I'd gladly try it if there's evidence it could work.)

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    Quote Originally Posted by mikewilder View Post
    I and one other grower tried applying NA, on the hypothesis that cytokinins from multiplication had caused lingering DNA methylation changes that were causing the plants to continue dividing instead of maturing. This had no effect, though only one NA concentration was tried, and only on a small number of plants:

    http://www.terraforums.com/forums/sh...light=elongata

    Your hypothesis has the virtue of being testable, and is worth a try, but it seems like the role of GA is not that straightforward AFAIK. Taiz and Zeiger reference added gibberellins causing early maturation in conifers, but note that added gibberellins cause reversion to juvenile form in Hedera helix. Neither are closely related to Heliamphora, but Hedera is significantly closer, so that's why I question adding GA as the solution. I'm the first to admit I'm no expert on plant phys--do you any references to forced maturation in herbs from adding Gibberellins? (That's not a rhetorical question, I'm actually asking, as I'd gladly try it if there's evidence it could work.)

    In that case, I'll be the second to admit I'm no expert

    I'll have to find a specific study on that, Mike, but it's almost always the case that dwarf plants have some kind of modification to gibberellic acids, whether it's synthesis, activation, or lack of receptors, which will result in reduced cell elongation and internode length. Also, seeing as we're distinguishing between two pitcher morphologies to judge the maturity of these plants, the fundamental role that GA plays in heterophylly could be another indicator that this hormone has somehow been disrupted.

    However, so I don't add to the misinformation already pervasive on these forums, I'll try to track a few papers down.


    Edit: Here's something I dug up on the fly that may help: http://www.sciencedirect.com/science...04377082900420

    and http://onlinelibrary.wiley.com/doi/1...nticated=false
    Last edited by mato; 06-01-2014 at 11:58 AM. Reason: Added link

  3. #35

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    I looked into it a little further, on the hypothesis that these clone lines are dwarf, added GA can cause stem elongation, but only in 1 of 3 "dwarf" mutation types studied. If the mutation is in regulation (multiple possibilities) rather than biosynthesis of ga, added ga shouldn't make a difference. So even on the assumption that these are genetic dwarfs, and that GA is involved, there are multiple dwarf genotypes (from regulation abnormality) where GA addition is not expected to have effect. But it is unclear to what extent Arabidopsis is a perfect model, and also it may well be that these Heliamphora are biosynth mutants. I do think the added twist with this genus you mentioned--switching from juvenile to adult leaves--is relevant, which is why the heterophyllous Hedera results troubled me. But it could be argued that added GA causing juvenile reversion in wt Hedera doesn't have straightforward implications for a hypothesized GA biosynth mutant.

    In the end, it's hard to say, but it's certainly hard for me to watch elongatas do nothing for years. I'll try it, don't really feel like I have anything to lose

  4. #36
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    I just ordered some, actually. I'd like to see if it affects a few Nepenthes clones that have done nothing for me in ages, as well as the hispida division I got from Butch a while ago. I'll post again after I decide on an appropriate dilution ratio.

  5. #37

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    please see "A gibberellin insensitive mutant of Arabidopsis thaliana." Koornneef et al 1985 Physiologia plantarum and let me know what concentration they applied and how, I can't get the paper but the abstract suggests they applied a nonlethal GA dose that was expected to restore function in a biosynth mutant. If PSU doesn't have it physically or electronically, they'll get a scan for you. I've got GA and at least 9 elongatas to test, just need to know concentration and application method like you.

  6. #38
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    Koornneef et al. 1985:

    Materials and methods

    Plant and seed material
    All mutants described derive from the pure line Lands- berg "erecta" (wild type). The dwarf mutant 37.630 was provided by Dr Dellaert who isolated this mutant in the progeny of a seed whicb was X-ray treated witb 467 Gy (10 Gy = 1 krad) wbile submerged in a 1.2% dithio- threitol solution. For mutant induction procedures see L, W. M. Deliaert (1980. Thesis, PUDOC, Wageningen, The Netherlands) and Koornneef et al. (1982a). This dwarf mutant was first thought to be a GA sensitive mu- tant because of its phenotypic resemblance to other pre- viously isolated GA sensitive dwarfs at the ga-1, ga-2, ga-3, ga-4 and ga-5 loci (Koornneef and van der Veen 1980). Like these dwarf mutants, 37.630 was also char- acterized by slightly narrower leaves with a darker leaf colour than the wild type. In addition, the number of axillary shoots was increased. However, dwarf mutant 37.630 had no reduced petals. In this respect it did not resemble dwarfs at the ga-1, ga-2 and ga-3 loci.
    Mutant NG5 used in the present experiments is a non-germinating GA-responsive mutant at the ga-1 lo- cus;, 29.423 is a dwarf mutant also allelic to tbe ga-I lo- cus, but it germinates without GA application. Its beigbt is slightly more than that of 37.630 (Fig. 1). Ap- parently, in 29.423 the ga-1 gene is still partially func- tional and therefore it was called a leaky ga-1 mutant (Koornneef and van der Veen 1980).
    In order to obtain a genotype combining the extretne GA sensitivity of these ga mutants with the characteris- tics of the 37.630 dwarf,, the mutant was crossed with line NG5. The F, was a semi-dwarf comparable to the F, wild type x 37.630, and other ga x 37.630 F,'s, which indicates non-allelistn. In the Fj population, 16 plants with the 37.530 phenotype were allowed to self, and 13 progenies segregated non-germinating seeds when sown under standard conditions (see below). About half of these seeds germinated within a week after transfer to a medium containing 10 iiM GA,+,. When transferred to the greenhouse, the seedlings developed into very small dark green bushy dwarfs with undeveloped petals, as is typical for ga-1 mutants. Spraying tbese plants twice with a 100' jiM GA4.J7 solution led to shoot elongation along with normal flower development and good seed production. Sufficient seeds of "heterozygotes were ob- tained by large scale crossing of the mutants and double mutants witb male steriles (ms-1 locus) of wild type and ga-1 mutants. Seed material of the different genotypes was grown simultaneously in the greenhouse and was harvested and stored under identical conditions. Ga-1 mutants were sprayed 4 and 5 weeks after germination with 100 (iM GA4.,, in order to promote seed set.

    Conditions of culture
    Seeds were sown in 9 cm glass petri dishes on perlite (40 ml) to which 25 ml of a standard mineral solution were added (Oostindier-Braaksma and Feenstra 1973). When ga-1 mutants were sown, 10 (iM GA^.^, was included. After 5 days at 4-^C to break seed dormancy, tbe seeds were allowed to germinate at 24C under continuous fluorescent light (Philips TL 57) at an irradiance of ap- proximately 8 W m"^. After 8 days the seedlings were transplanted into clay pots and grown in an air-condi- tioned greenhouse. From October until April additional light (Philips, HPI/T400 W lamps) was given from 24.00 h-10.00 h. Day temperature was 22-25C and night temperature 16-18C.

    Physiological characterization
    To test the GA-sensitivity, GA was either sprayed 00 the plants or applied with a Hamilton micro-syringe in a quantity of 4 |il as 2-4 small drops on just expanding leaves. GA was applied 2, 3 and 4 weeks after in- cubation of the seeds at 24C, around the start of elon- gation of the main stem. Final plant height was meas- ured approximately 8 weeks after germination, when elongation of the main stem was complete. At the same time, the number of primary axillary shoots in the roset- te was also counted. The seed germination tests were performed in triplicate, with 50 seeds per treatment ac- cording to Koornneef et al. (1982b). Germination was assessed 7 days after incubation.

  7. #39

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    Very much appreciated Mato, including their reference to "a leaky GA-1 mutant", which made me laugh. But I don't understand any of the concentrations:
    "10 (iM GA^.^ " is that supposed to be 10 uMolar, I'm unfamiliar with iM?
    "100 (iM GA4" ditto
    "GA was either sprayed 00 the plants or applied with a Hamilton micro-syringe in a quantity of 4 |il as 2-4 small drops" Huh? 4 microliters? of 100 uMolar GA4? or 10 uMolar GA4? Maybe I'm just slow, can you tell what they are saying?

  8. #40
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    Sorry, it looks like the characters did not copy over correctly. Yes, "(i" should all be micro (μ).

    This might help a little bit:

    Last edited by mato; 06-01-2014 at 03:28 PM.

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