Re: Embryo culture (was: Re: spec-x with Chinenses series species?)

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-----Original Message-----
From: "Dennis Kramb dkramb@badbear.com [iris-species]"
Sent: May 11, 2015 9:57 AM
To: iris-species@yahoogroups.com
Subject: Embryo culture (was: Re: [iris-species] spec-x with Chinenses series species?)

 

I'm going to try the simpler sterilization techniques first.  I don't own any sophisticated equipment, and I've never owned a pressure cooker.

The article by Harald Mathes was the only one in which the test tube size was mentioned.  He used 160mm tubes.  The plants in his photos were sometimes crumpling up against the tube lids, so I decided taller is better.  I ended up purchasing 200mm (7.8 inches) tubes because the 250mm were out of stock.

I found a nutrient media at PhytoLabs that includes agar & sugar mixed in.  There are two options.  One has 30g/L of sucrose.   The other has 15g/L each of sucrose and glucose.  I have no idea which one would be better for Irises.  They're cheap enough I guess I could buy both.

So yeah, imagine my surprise at how cheap the nutrient media is.  But then realizing that I need to buy a precision scale to measure out the media powder. ! And precision scales are difficult to find under $100!  Yikes!  I might be able to use a scale at work.  I need to check.

Dennis in Cincinnati

On Sun, May 10, 2015 at 9:21 AM, k*@astound.net [iris-species] <i*@yahoogroups.com> wrote:

Hi Dennis,

One thing to note when slicing non-aril seeds is that you
are looking for the point of attachment of the seed to the
pod (hilum). That is where the tip of the embryo is poised
to push out of the seed. If you remove the seed coat from
around the hilum, there should be slight bulge. On most
iris species it shouldn't be too hard to find, but on a
few it is problematic. The length that the embryo extends
back into the seed is quite species dependent. Note that
I've had more experience trying to remove embryos from the
seeds of various species than I've had getting embryos to
adult plants.

I'm away from home at the moment. I know the test tubes I
use are smaller than yours, but feel that mine are smaller
than ideal. 10" tubes sound good. Do you have a pressure
cooker deep enough for your tubes or will you try a lower
level of sterilization?
I have bought medium from PhytoTechnology Laboratories. I
found some notes about product number M508. It needs to be
used added to agar and sugar. I'm not sure that product is
the one I had success with initially. Last I tried some
cultures with old medium packets that had been
refrigerated for several years. It didn't work as well as
I had hoped. I'm not sure if it is the particular medium
(I don't recall if it was M508) or the fact that it was
old. I am thinking of researching other media.

Ken

On Sat, 09 May 2015 21:55:01 -0400
  "dkramb d*@badbear.com [iris-species]"

<i*@yahoogroups.com> wrote:
> I found it in the 2008 Yearbook.  WOW what a great set
>of photos!
> You're absolutely right that details on the embryo
>extraction process
> are what all the other articles fail to describe.  One
>of them just has
> a single sentence on it.  LOL.
>
> I just purchased a bunch of equipment (test tubes,
>forceps, scalpels,
> funnel, etc.) to get me started but I haven't purchased
>any
> agar/nutrient powders yet.  Can you give me any
>suggestions?  I'm mostly
> doing spec-x crosses involving versicolor, and others
>with tectorum.
>
> Oh by the way, if you grow yours in test tubes what size
>do you find
> works best?  I went with 1" x 10" since versicolor gets
>awfully tall
> awfully fast.
! >
> Thanks, Ken!
> Dennis in Cincinnati
>
>
> On 5/9/2015 8:58 PM, k*@astound.net [iris-species]
>wrote:
>> I wrote a small article with photos on removing embryos
>> from aril seeds back in 2009 for ASI. Because of the
>> photos, the Word document is almost 3M, but it should be
>> e-mailable if someone would like a copy.
>>
>> Ken
>>
>> On Sat, 9 May 2015 15:29:04 -0400
>>    "Dennis Kramb d*@badbear.com [iris-species]"
>> <i*@yahoogroups.com>  wrote:
>>> I'm reading my old ASI yearbooks to learn the techniques
>>> of the pros for
>>> embryo rescue.  it seems remarkably easy!  I have some>>> TB seed and Juno
>>> seed to practi! se with.  I may try the forced/cutting
>>> technique too, as
>>> there are extremely good results reported with it as
>>> well.  I tried
>>> scarifying some tridentata and tectorum seeds this
>>> winter with 100% failure
>>> rate. LOL.  My tectorum seeds were surplus SIGNA seed
>>> like you mentioned.
>>>
>>> Dennis in Cincy
>>> On May 9, 2015 2:59 PM, "Walter Pickett
>>> w*@yahoo.com
>>> [iris-species]"<i*@yahoogroups.com>  wrote:
>>>
>>>>
>>>> Yess, the kit is a luxury I couldn't, or didn't want to,
>>>> afford.
>>>> The MS medium is available from many chemical supply
>>>> ! companies.  It comes
>>>> in bulk and in pre-measured packet for 1 liter distilled
>>>> water, or
>>>> deionized water.  Some say they use tap water and I
>>>> doubt they have any
>>>> problems with it.  For $1 per gallon for distilled, I've
>>>> never tried tap
>>>> water, but actually, while scientific journal articals
>>>> always give exact
>>>> recipes, there is a lot of room for improvizing.  Think,
>>>> how exact is the
>>>> chemistry of the soil these iris grow in?
>>>> Of iris, I have onlly done I. tectorum.  The only reason
>>>> is used the
>>>> sterilized seeds and growing medium was  the it wouldn't
>>>> mold before
>>>> germination.  And I was praciticing for embryo culture
>>>>! ; with arils.  I
>>>> didn't want to risk the more ! rare arill seeds.  I.
>>>> tectorum is a fie plant,
>>>> but it seeds easily and there are always leftover I.
>>>> tectorum seeds after
>>>> the sale,  The Signa seed sale didn't even have any
>>>> raail seeds  this
>>>> year.  But of course the ASI has a seed sale every year.
>>>>
>>>>
>>>>
>>>>    On Saturday, May 9, 2015 12:05 PM, "Dennis Kramb
>>>> d*@badbear.com
>>>> [iris-species]"<i*@yahoogroups.com>  wrote:
>>>>
>>>>
>>>>
>>>>   I am super annoyed at how few of my seeds have
>>>> germinated so far this
>>>> spring.&nb! sp; I'm sick of the waiting game, so I'm gonna
>>>> investigate this embryo
>>>> rescue at home technique.  I checked out the kitchen
>>>> culture page and I had
>>>> a little bit of sticker shock at the prices of their
>>>> kits, so I'm gonna
>>>> keep shopping around.
>>>>
>>>> Dennis in Cincinnati
>>>>
>>>>
>>>> On Wed, Apr 22, 2015 at 4:33 PM, Walter Pickett
>>>> w*@yahoo.com
>>>> [iris-species]<i*@yahoogroups.com>  wrote:
>>>>
>>>>
>>>>
>>>> Sean.
>>>> Embryo rescue isn't all that hard, depending on your
>>>> eyesight.  I h! ave to
>>>> use a diesecting scope now.
>>>&! gt; There is a website called kitchen culture, or kitchen
>>>> tissue culture, or
>>>> such.  Google will bring it up.
>>>> In addition to the information on the site,, there is a
>>>> link to an egroup
>>>> of home tissue culture practicers, mostly amatures but
>>>> including
>>>> professionals and professors, etc.
>>>> They are full of good information, and will answer
>>>> questions.
>>>> The chemicals are available from several chemical supply
>>>> companies, the
>>>> medium pre-measured for one liter of sterile distilled
>>>> water. Our standard
>>>> culture vessels are babyfood jars, which are not
>>>> terribly expensive at your
>>>> local food store full of babyfood, cheaper than buying
>>>> babyfood jars
>! ;>>> online.  The jars might be cheaper at a daycare if you
>>>> can find one.
>>>> I used to be quite active on the group, but I  have been
>>>> too busy this
>>>> winter.
>>>> Walter
>>>>
>>>>
>>>>
>>>>    On Tuesday, April 14, 2015 8:15 PM, "Sean Zera
>>>> z*@umich.edu
>>>> [iris-species]"<i*@yahoogroups.com>  wrote:
>>>>
>>>>
>>>>
>>>>   Anita,
>>>>
>>>> It would be nice to be able to use that technique on
>>>> hybrid seeds; I don't
>>>> think it's too involved, but requires some basic plant
>>>> mi! cropropagation
>>>> skills I don't have. :)
>>&! gt;>
>>>> As for the *koreana*, if it's really self-incompatible,
>>>> I can just have
>>>> someone mail me some pollen, or try wide crosses on it
>>>> like Dennis.
>>>>
>>>> Sean Z
>>>>
>>>>
>>>>
>>>>
>>>>
>>>>
>>>>
>>>>
>>>>
>>>>
>>>>
>>
>>
>> ------------------------------------
>> Posted by:<k*@astound.net>
>> ------------------------------------
>>
>>
>> ------------------------------------
>>
>> Yahoo Groups Links
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>>
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>>
>



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Posted by: <k*@astound.net>
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