-- [ From: Yang, Doo Suck * EMC.Ver #2.5.02 ] --
Strepcomyces is a genus included in Actinomycetes.
It grows in mycelia, like fungi. So it's difficult, even impossible to
measure
OD. Usually Streptomyces growth is measured by Dried Cell Weight (DCW)
or Packed mycelial Volume (PMV).
There are three ways to make sample equally suspended. One thing is using
mild homoginization, which makes fragments of mycelium.
The other method is using enzyme which breaks cell wall, like lysozyme. That
makes protoplast.
The third is using medium on which Streptomyces makes spores.
Then use spores to prepare equally suspended samples.
See below for medium composition.
Sporulation medium (R2YE, Hopwood et al., 1985)
Sucrose 103 g
K2SO4 0.25 g
MgCl2¡¤6H2O 10.12 g
Glucose 10 g
Casamono acids 0.1 g
Yeast extract 5 g
Agar 22 g
Distilled water 800 ml
After autoclaving add following constituent;
KH2PO4 (0.5%) 10 ml
CaCl2¡¤2H2O (3.68%) 80 ml
L-proline (20%) 15 ml
TES buffer (5.73%, pH 7.2) 100 ml
NaOH (1N) 5 ml
Trace element solution* 2 ml
*Trace element solution (per liter)
ZnCl2 40 mg
FeCl3¡¤6H2O 200 mg
CuCl2¡¤2H2O 10 mg
MnCl2¡¤4H2O 10 mg
Na2B4O7¡¤10H2O 10 mg
(NH4)6Mo7O24¡¤4H2O
-------- REPLY, Original message follows --------
> Date: Thursday, 05-Dec-96 03:19 PM
>> From: aj mccardell \ Internet: (mccardaj at esvax.dnet.dupont.com)
> To: microbio at net.bio.net \ Internet: (microbio at net.bio.net)
>> Subject: Streptomyces/O.D. reading
>> I am trying to develop a sample preparation procedure with Streptomyces
and am
> having some difficulty. Our standard sample prep procedure is to use a
> plastic stick to pick colonies from an agar plate. Since the Streptomyces
I am
> growing are quite 'crusty' and grow into the agar somewhat, I can't
recover
> them from the agar surface in this way. I thought I would switch to
growing
> them up in broth and then adjust the O.D. to my target range. The problem
is
> that I can't vortex to achieve a uniform suspension of cells. These
things are
> like big strings that don't disperse. Also, when I have dug colonies from
the
> plates, I haven't been able to vortex the sample into any kind of
suspension.
>> I am currently using BHI media to grow these samples. They are growing
> alright. I was thinking about switching to Bennett's medium in the hopes
of
> getting a little bit more luxurious growth. Does anyone have any
suggestions
> for media? How do people take O.D. readings of Streptomyces? (I've tried
> incubating plates at higher humidity and have been thinking about
ammending the
> media with a little Tween but that's the limit of my less-than-brilliant
> ideas.)
>> Hope someone can help.
>> Thanks,
> A.J.
>>
-------- REPLY, End of original message --------
--
+++++++++++++++++++++++++++++++++++++++++++++++++++
Hanhyo Institutes of Tehchnology
Fermentation Lab.
461-6 Chunmin-dong, Yoosung-ku,
Taejun, 305-390, KOREA
Tel. 82-42-866-9136
Fax. 82-42-866-9129
Research Scientist
Yang, Doo Suck
+++++++++++++++++++++++++++++++++++++++++++++++++++
