Dear Correspondents/Immortalists,

Though lofty ideals aren't to be discouraged , I do feel that
on occassion our feet aren't firmly upon the ground.
To help regain some focus , and show that politics need not
permeate every aspect of Scientific Research , I'm outlining
a possible experiment , that even an under-graduate might
attempt. Without an ethical review board being involved ,
if he/she only uses species other than human in their
investigations.

This is very much a tentative outline and may require
considerable revision.

I hope I posted this message , intended for all of the Yahoo,
immortalist/cloning group , in the right way.


Regards,
Edmont.


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Experiment 1
-----------------

Aim:
To determine the factors involved in the re-programming of
a differentiated adult cell to the developmental stage of an
Embryotic Stem Cell.

Apparatus:

High resolution microscope , possibly binocular.
Micro-pipette , with mechanical positioning.
Incubator , thermostatically controlled.
Cellular growth medium , suitable for culturing ESC's.
Various dyes , specific to particular cellular organelles.
and cell types.
Protein analysis equipment , electrophoresis or advanced.
Centrifuge , or other method of isolating cellular fractions
*Gene Chip[s] , possibly used to examine the genetic profile
at various stages of re-programming , or development.

Material:

Embryotic Stem Cells [ ESC's ] , in the first instance from a
species like Xenopus [ Giant African clawed tree frog ].
Differentiated Cells from an adult Xenopus.

Method:

The Xenopus ESC's are incubated at the appropriate
temperature and in a medium suitable for their development.
From a large number of these ESC's , the ooplasmic material
is removed ; excluding the chromosome.
Now particular fractions , from the ooplasmic material are
isolated through the application of the centrifuge and other
methods.
These fractions are carefully labelled and stored in the
necessary manner for their preservation.
Small amounts are used from these , in various combinations,
to [ hopefully ] determine the active components involved in
cellular re-programming.
Using the micropipette a specific combination of extracts
from the oocyte are inserted into a specific adult cell ,
from the Xenopus.
Computerized records help keep track of the combinations
and results.
The adult cells, that have undegone ooplasmic transfer, are
cultured in a suitable medium and at a suitable temperature
for the necessary amount of time.
Dyes are used to determine the type of cell that has resulted
from the ooplasmic transfer.
When a result indicates that an Embryotic type is present ,
this is cultured for an extended period of time.

After referring back to the particular combination that
produced these results , a small amount , from the stored
vial, is analyzed more extensively.

Repeat , using only the compounds found in the vials that
produced the favorable result[s].


Comments:

Ooplasmic transfer has been shown to be effective in
re-programming a differentiated adult cell.
This is just an extension of what I assume the methodology
might involve.

One of the crucial aspects of this experiment is the
identification of the components of the ooplasmic material
and the maintainance of this viability , during the
fragmentation.
The experimental tolerances involved aren't known at this
time.

This approach should be quite successful in isolating the
active component , this is essentially a binary search.
Identification and possible synthesis of the active component
would be performed elsewhere.

Once the feasibilty of this approach has been established ,
then a different species might be examined.
Of particular interest is the possibility of a substance that's
similar for most all species.
If this does appear to be the instance , then a cross-species
test might be performed .

-------------------------------------

Eventually the identification of the factor for humans might
be performed. This would however , require the use of human
adult cells and ESC's.

References:


Ooplasmic transfer has been used , quite sucessfully, with
In Vitro Fertilization , an internet search will provide details.
Advanced Cell Technology mention this as one approach to producing
clones , visit there website for further information.


--------- Ooplasmic Transfer ----------

New England Journal of Medicine. Vol 346 , issue 10, Mar 7 , 2002,
Pages 773-775.

Trends in biotechnology. Volumne 19, issue 10 , 1 October 2001 ,
Page 381.

Mitochondria in Human offspring derived from ooplasmic
transplantation.

Human reproduction. issue 3, March 2001 Pages 513 - 516 .

Mitochondrial DNA heteroplasmy after human ooplasmic transplantation.
Fertility and Society . Volumne 76 , issue 3 , September 2000.
Pages 573 - 578.

Spontaneous and artificial changes in human ooplasmic mitrochondria,
Human reproduction ( Oxford , England ) Volumne 15 , Supplement 2,
July 2000 , Pages 207 - 217 .

Ooplasmic transfer in mature human oocytes .
Molecular human reproduction. Volumne 4, issue 3 , March 1998,
Pages 269 - 280.

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