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Kinetin-like activity in root apices of sunflower plants
Life Sciences Vol. 4, pp , 1323-1328, 1985 . Great Britain.
Pergamoa Preen Ltd. Printed in
~INSTIR-LI1~ ACTIVITY IN ROOT APICSB OF SUNFLOYER PLAlITB Chanan Yeiss and ?oash Vaadia Department of Plant Phyaiolog~, Regev Institute for Arid Zoae Research, Heersheva, Israel
(Received 19 April 1985)
There is accumulating circumstantial evidence for the presence of growth-regulating factors that originate in roots . These factors are said to affect growth (1), protein metabolise (2), senescence (3),and shoot response to root temperature (4), oseotic stress (5), water stress (6), and floo ding {~) . Kinina are able to substitute for such factors in several cases (8, 9, 10) .~Rich sources for kinins are rapidly-dividing tissue, e .g . fruitlets (11) and crown-gall tumors (12) . TY~ey were also detected in root exudate of tobacco (13), SrApevine (14),~and sunflower plants (15), but could not be found in maize-root tissues (16) . In the work presented here extracts of rapidly-dividing, meristematic root apices were assayed for kinin activity . The data support the hypothesis that root apices are the origin of root kinins .
Experimental Sunflower seeds, soakeù overnight in water, were transferred 1329
1324
KIIdETIN-LII~E ACTIVITY IId ROOT APICES
Yal . 4, No. 13
to flats lined with moist filter paper. After 48 hours, 1 ms sections of root apices (0-1 sun trom .the tip) and 2 mm sections of the radicles (1-3 mm from the tip) from 1200 roots, were cut
and freeze-killed separately in precooled (-37° C) absolute ethanol . The sections were extracted in a cold room by stirring for two hours in three portions of 75 ml of 80Sé ethanol . The original absolute ethanol traction and the three extraction aliquots were combined and the alcohol evaporated under vacuum at 34oC, the water phase then being lyophilized and the dry residue redissolved in 1 ml of water . This solution was applied as a streak to Whatman No .3 paper . Ascending-descending chromatograms were developed with n-butanol-acetic acid-water (4 :1 :1), over a distance of 30 cm, and air dried. Transverse strips corresponding to Rf values or U .V . absorption zones were cut and eluted with 80~ ethanol, the eluates being air dried and redissolved in 2 ml of mater . Each fraction was added to two 100 ml Erlenmeyer Sleeks containing 30 ml modified White's medium plus IAA (li) " Four pieces of callus originally derived from cotyledons of soybean (var . Lee) were placed in each flask.
Results and Discussion Kinetin-like activity ras found in root-apex extracts in the five experiments performed . Representative reéults are presented in Table I . Significant kinetin-like activity can be observed in root-apex extracts, but little, if any, in the physiologically older root tissue . The activity is present in two chromatogram zones .
Vol . 4, No. 13
1325
KINETIN-LII~E ACTIVITY IId ROOT APICES TABLE I
Kiaetia-like Activity as Estimated by Callus-growth Bioassay of Chromatographic Fractions of SSctracts .trom 1200 Roots . Chromatograms were cut to contain U.V, absorption bands at Rt 0 .15-0.1~ and 0 .~3-0 .80 . Other chromatogram z~egions showed no .kinetin-like activity . Total weight of Roots Mg
Rt 0.15-O.1Q Wt per Callus Mg
Fresh
Dry
474.2
163 .9
507 .9
Root sections 1-3 mm 867 .6 Srom Tips
115 .0
32 .1
Root apices O-1 mm from Tips
Kinetin Equivalents ~ 1200 roots
0 .13
O .QOl,
Rt 0.~3-0 .80 Yt per Callus
~Kinetia $quivalents Mgt 1200 roots
570.5
0 .17
14 .4
0.001
Kinins have not been shown to be a required addition to media used in continuous root-tissue culture (18) . This . a s well as the agreement of our results with those of Kende (15) for root exudate of sunflower plants, suggests that roots synthesize kinins . The successful demonstration of kinetin-like activity in roots is attributed to the use of the apices themselves rather than the whole roots . The data presented support Goldacre's assumption (19) that kinins may be formed in meristematic root apices . Acknowledgements The authors thank Dr . H. Kende for his advice . P . Cohen's
1lf6
B.IIdETIN-LII~ ACTIVITY IN ROOT APICES
Vol. ~, No . 19
coa~enta are appreciated. Rererences 1.
F.Y . Yali?, Plant Ph7siol. ].8i 51 (1943) .
2.
".C . CHIBNILL, }~ev Phytol . ~, 31 (1954) .
3.
L. BNGELBREC~, Flora ~, $7 (1964) "
4.
R.M . D"VIS and J .L . LINGLZ, Plant Pi~siol . ~6, 153 (1961) .
5.
V. MEYER and V. GINGRICH, Science 144, 1463 (1964) .
6.
Q. YEISS and T. VIADIA, in preparation .
7"
P.J . äRAI+ISR, Plant Ph7siol . 26, 722 (1951) .
8.
R. irOLLGIEHli and B . PARTHISR, Phytoche~. ~, 241 (1964) .
9.
C.B : BBAH aad R .S . LOOIÜB . Plvsiol . Plant . 18, 240 (1965) .
10 . " .C . LÉOPOLD and M . iUYASE, . Am . J. Bot. ~, 294 (1964) " 11 . D.S . LETHAM, Lite Sci . ~, 152 (1963) . 12 . H.N-. YOOD, ReRUlateura Naturels ds la Croissance Ve~xetale , Editions du Centre National de la Recherche Scientiliqus, Paria, 1964, P " 97 " 13 . O .lf. RULAI~V" , Soviet Plant Ph~aiol. (Sngl . traps.) (1962) .
s,
182
14. J.E . LOEFFL&R and J. VAN OVERBEEK, Regulateura Naturels de la Croissance Yepcetale, Editions du Centre National de la Recherche 8cisntitiqus, Paria, 1964, p . 77 . 15 . H. KEDTDE, Proc . Nat. "cad . Sci ., in press. 16 . F. H. YITHAM and C.O . MILLER, Plant Physiol. aupp . ~8, 7CYYYIII (1963) .
-
17 . C .O . MILL$R, Modern Methods of Plant Analysis , H.F . Linskens and M.V . Traced Sds . Vol . 6, 194, Springer, Berlin (1963) . 18 . D.N . BUTCHER and H .E . STREET, Bot . Rev . _~, 513 (1964) . 19 . P.L . GOLDACRE, Auat . J. Biol . 8ci . 12, 388 (1959) "
Pergamoa Preen Ltd. Printed in
~INSTIR-LI1~ ACTIVITY IN ROOT APICSB OF SUNFLOYER PLAlITB Chanan Yeiss and ?oash Vaadia Department of Plant Phyaiolog~, Regev Institute for Arid Zoae Research, Heersheva, Israel
(Received 19 April 1985)
There is accumulating circumstantial evidence for the presence of growth-regulating factors that originate in roots . These factors are said to affect growth (1), protein metabolise (2), senescence (3),and shoot response to root temperature (4), oseotic stress (5), water stress (6), and floo ding {~) . Kinina are able to substitute for such factors in several cases (8, 9, 10) .~Rich sources for kinins are rapidly-dividing tissue, e .g . fruitlets (11) and crown-gall tumors (12) . TY~ey were also detected in root exudate of tobacco (13), SrApevine (14),~and sunflower plants (15), but could not be found in maize-root tissues (16) . In the work presented here extracts of rapidly-dividing, meristematic root apices were assayed for kinin activity . The data support the hypothesis that root apices are the origin of root kinins .
Experimental Sunflower seeds, soakeù overnight in water, were transferred 1329
1324
KIIdETIN-LII~E ACTIVITY IId ROOT APICES
Yal . 4, No. 13
to flats lined with moist filter paper. After 48 hours, 1 ms sections of root apices (0-1 sun trom .the tip) and 2 mm sections of the radicles (1-3 mm from the tip) from 1200 roots, were cut
and freeze-killed separately in precooled (-37° C) absolute ethanol . The sections were extracted in a cold room by stirring for two hours in three portions of 75 ml of 80Sé ethanol . The original absolute ethanol traction and the three extraction aliquots were combined and the alcohol evaporated under vacuum at 34oC, the water phase then being lyophilized and the dry residue redissolved in 1 ml of water . This solution was applied as a streak to Whatman No .3 paper . Ascending-descending chromatograms were developed with n-butanol-acetic acid-water (4 :1 :1), over a distance of 30 cm, and air dried. Transverse strips corresponding to Rf values or U .V . absorption zones were cut and eluted with 80~ ethanol, the eluates being air dried and redissolved in 2 ml of mater . Each fraction was added to two 100 ml Erlenmeyer Sleeks containing 30 ml modified White's medium plus IAA (li) " Four pieces of callus originally derived from cotyledons of soybean (var . Lee) were placed in each flask.
Results and Discussion Kinetin-like activity ras found in root-apex extracts in the five experiments performed . Representative reéults are presented in Table I . Significant kinetin-like activity can be observed in root-apex extracts, but little, if any, in the physiologically older root tissue . The activity is present in two chromatogram zones .
Vol . 4, No. 13
1325
KINETIN-LII~E ACTIVITY IId ROOT APICES TABLE I
Kiaetia-like Activity as Estimated by Callus-growth Bioassay of Chromatographic Fractions of SSctracts .trom 1200 Roots . Chromatograms were cut to contain U.V, absorption bands at Rt 0 .15-0.1~ and 0 .~3-0 .80 . Other chromatogram z~egions showed no .kinetin-like activity . Total weight of Roots Mg
Rt 0.15-O.1Q Wt per Callus Mg
Fresh
Dry
474.2
163 .9
507 .9
Root sections 1-3 mm 867 .6 Srom Tips
115 .0
32 .1
Root apices O-1 mm from Tips
Kinetin Equivalents ~ 1200 roots
0 .13
O .QOl,
Rt 0.~3-0 .80 Yt per Callus
~Kinetia $quivalents Mgt 1200 roots
570.5
0 .17
14 .4
0.001
Kinins have not been shown to be a required addition to media used in continuous root-tissue culture (18) . This . a s well as the agreement of our results with those of Kende (15) for root exudate of sunflower plants, suggests that roots synthesize kinins . The successful demonstration of kinetin-like activity in roots is attributed to the use of the apices themselves rather than the whole roots . The data presented support Goldacre's assumption (19) that kinins may be formed in meristematic root apices . Acknowledgements The authors thank Dr . H. Kende for his advice . P . Cohen's
1lf6
B.IIdETIN-LII~ ACTIVITY IN ROOT APICES
Vol. ~, No . 19
coa~enta are appreciated. Rererences 1.
F.Y . Yali?, Plant Ph7siol. ].8i 51 (1943) .
2.
".C . CHIBNILL, }~ev Phytol . ~, 31 (1954) .
3.
L. BNGELBREC~, Flora ~, $7 (1964) "
4.
R.M . D"VIS and J .L . LINGLZ, Plant Pi~siol . ~6, 153 (1961) .
5.
V. MEYER and V. GINGRICH, Science 144, 1463 (1964) .
6.
Q. YEISS and T. VIADIA, in preparation .
7"
P.J . äRAI+ISR, Plant Ph7siol . 26, 722 (1951) .
8.
R. irOLLGIEHli and B . PARTHISR, Phytoche~. ~, 241 (1964) .
9.
C.B : BBAH aad R .S . LOOIÜB . Plvsiol . Plant . 18, 240 (1965) .
10 . " .C . LÉOPOLD and M . iUYASE, . Am . J. Bot. ~, 294 (1964) " 11 . D.S . LETHAM, Lite Sci . ~, 152 (1963) . 12 . H.N-. YOOD, ReRUlateura Naturels ds la Croissance Ve~xetale , Editions du Centre National de la Recherche Scientiliqus, Paria, 1964, P " 97 " 13 . O .lf. RULAI~V" , Soviet Plant Ph~aiol. (Sngl . traps.) (1962) .
s,
182
14. J.E . LOEFFL&R and J. VAN OVERBEEK, Regulateura Naturels de la Croissance Yepcetale, Editions du Centre National de la Recherche 8cisntitiqus, Paria, 1964, p . 77 . 15 . H. KEDTDE, Proc . Nat. "cad . Sci ., in press. 16 . F. H. YITHAM and C.O . MILLER, Plant Physiol. aupp . ~8, 7CYYYIII (1963) .
-
17 . C .O . MILL$R, Modern Methods of Plant Analysis , H.F . Linskens and M.V . Traced Sds . Vol . 6, 194, Springer, Berlin (1963) . 18 . D.N . BUTCHER and H .E . STREET, Bot . Rev . _~, 513 (1964) . 19 . P.L . GOLDACRE, Auat . J. Biol . 8ci . 12, 388 (1959) "