الفهرس 
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Abstract
Azotobacter Chroococcum, Azotobacter vinelandii and
Escherichia coli. were subjected to genetic transformation in order to
aquiresaponin for producing ability. The transforming materials in case
of Azotobacter sp. were Saponaria high molecular weight DNA,
Saponaria officinalis fresh roots DNA, Poinciana regia chromatin
DNA and Asparagus officinalis fresh roots DNA, each of
concentration 22 ug/ml, In case of E.coli, the transforming material was
Saponaria fresh roots DNA of concentrations 66, 50 and 30 J.1g/ml.
Some biolgocial activites of saponin in each ofliquid culture of
transfonnant and untransfonnant Azotobcter, culture supernatant, cell
suspensions and extracted (TLC) saponin bands of culture supernatants
of isolate (1) of transfonnant, untransfonnant and isolate ”2” of
transfonnat’ltE. coli were tested with comparison to aqueous extracts of
3 plants (Saponaria, Poinciana: and Asparagus) of different concentrations
as follow:
. I. Haemolytic activity:
About 0.5 ml from each saponin solution mentioned above was
added to 5 ml of blood corpuscles suspension, Kept at room temp. for 5
minutes, centrifuged at 3000 rpm for 15 minutes. The content ofH.b in
supernatant was measured in coloremeter at 540 G.D. The saponin
content of unknown samples were caleculated from the linear portion--_.of the standrd curve which was constructed for white saponin of
Merck.
n Moluscicidal activity:
Ten snails (Biomphalaria alexandrina) were transferred to
about 10 ml from each tested saponin solutionand dechlorinated H2O
as control for 24 hrs in duplicate then transferred to dechlorinated
water for 24 hr. After recovery, mortality was calculated.
m. Nematicidal activity:
About 1000 2-nd stage juveniles were transferred to 10 ml of
each tested saponin soultion and distilled water as control in triplicates
for 24 hrs then were transferred to aerated water for 24 hrs. Mortality
percentages of nematodes were calculated after recovery.
- Type of saponin in plants was determined by the colour produced by
application of the liebennann- Burchard reaction.
- Various transformation frequencies in case of Azotobacter were
observed. The highest frequency was obtained from the two types of
Saponaria DNA.
- Transfonnant E.Coli were obtained with the concentration 66 ~glml
and the transformation frequency was 0.00096.
- Untransfonnant bacterial strain don’t produce saponin and don’t
exhibit any of its examined biological activities.
- Transfonnant strains exhibited various degrees of biological activites
of saponin as follows:-
- The Azotobacter chroococcum transformed by Saponaria high
molecular wieght DNA gave the highest (60%) molluscicidal activity.
This mortality value was equal to that obtained from standard saponin
(Merck) cone, of 0.015mglml). While the highestmolluscicidal
activity of cell suspensions of E.coli was that of the isolate (2) of
transformed strain (30%, equivalent to 0.014 mglml of Merck
saponin) which cause decrease in DNA content of snails to the lowest
value (5.2 mglg of snail tissue) with comparison to control which have
(23.1 mglg of snail tissue).
- The purified saponin extracted from culture supernatant of
transfonnant E.coli mortalized snails at the maximum percentage
(100%), which was equivalent to the mortality caused by 0.0175
mglml of Merck saponin, the DNA content of dead snail tissue was
reduced to about 4.1 mglg of snail tissue. Cell suspension and the
spots dervied from the wild E.Coli showed no molluscicidal activity.
- The best potent saponin examined in 3 plants was that of Saponaria
and Asparagus (LCso were 35, 40 mglml) respectively for snails,
which equivalent ot the mortality caused by 0.015 mglml of Merck
saponm.
Nematicidal activity:
- A. chroococcum transformed by Saponaria high molecular weight
DNA gave the highest nematodes mortality (50%) which corresponds
to 80 mglml concentation of Merck saponin.
-Cells suspension of isolate (1) oftransfonnant E.Coli gave the highest
nematodes mortality percentage (60%) in comparison to the cell
suspension of wild type and isolate (2) oftransfonnant E.Coli
-Purified saponin extracted from culture supernatant of isolate (1) of
transformant and isolate (2) of transformant E.coli. caused 100%
nematodes mortality, equivalent to 160 mglml of Merck saponin.
- The best potent saponin examined in 3 plants was that of Saponaria
and Asparagus LCso for nematodes were 100 mg plant! ml from each
plant.
Haemolytic activity:
Culture of A. chroococcum transformed by fresh Saponaria
.DNA could hemolyze red blood cells to an extend equal to the obtained
by 40.8 mgl ml of standard saponin.
- Haemolytic activity of isolate(l) of transformant and isolate (2) of
transformant E.coli cell suspensions was greater than that of the
supernatants. Isolate (1) of transformed strain gave stronger
hemolytic action than isolate (2) of transformed strain in both cases.
- The hemolytic activity of purified saponin extracted from culture
supernatant of isolate (1) oftransfonnant E.coli was lower than that
of isolate (2) oftransfonnant one (0.5 and 0.736) respectively.
- Aqueous extracts of Saponaria roots and Poinciana seeds at cone. of
100 mg plant! ml gave stronger hemolytic action than aqueous
extract of the same cone, from Asparagus, this may be due to the
differene in.raponin type (triterpenoid in Saponaria and Poinciana,
but it its steroid in Asparagus).
- Aqueous extract of Saponaria could hemolyze red blood cells to an
extend equal to that obtained by 0.DI5 mglml of standard saponin, so
Saponaria is considered to be the most producer for saponin than
Asparagus and Poinciana.