SSRG International Journal of Medical Science (SSRG-IJMS) Volume 4 Issue 7 July 2017

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1 SSRG International Journal of Medical Science (SSRG-IJMS) Volume Issue 7 July 7 Te Essential Role of Different Factors in Germination and Tube Growt of Heliotropium Hirsutissimum Grauer (Boraginaceae) Maa A. Hamdan*, Dina A. Haddad*, Hananamame* * Postgraduate student,department of Botany, Faculty of Science,TisreenUniversity,Latakia, Syria * Associate professor, Department of Botany, Faculty of Science,Tisreen University, Latakia, Syria * Lecturer, Department of Botany, Faculty of Science,Tisreen University, Latakia, Syria Abstract Te elongation of te pollen tube in flowering plants is exceedingly rapid and its requirements, in general, seem quite unimpressive, i.e., water, oxygen and a suitable osmotic milieu. Despite extensive attempts to asten tis growt process wit te conventional ost of growt factors, few ave met wit convincing suc We study te essential role of different factors (temperature, PH, Sucrose, Boric acid, Calcium) in pollen germination and pollen tube growt of eliotropiumirsutissimum (Boraginaceae).We used basal medium (Brewbaker&Kwack, 9) wit different temperatures (-- C), PH (--7-), Sucrose (---) g/l, Boric acid (-- -) mg/l, Calcium (---) mg/l. Te best result for eac factor in tis study was:- Temperature: C, PH:, Sucrose: g/l, Boric acid: mg/l, Calcium: mg/l. Keywords: pollen germination, eliotropium, Boric acid, Sucrose, PH I. INTRODUCTION In plants, post-pollination sexual selection can occur by competition between pollen grain and by female coice (stepens&bertin, 9) grains are believed to compete for ovules wen tere are more pollen grains on te stigma tan tere are ovules in te ovary. It is assumed tat under conditions of pollen competition, only te fast growing pollen tubes acieve fertilization (MuLCy, 979) Selection due to differences between pollen donors can occur eiter befor fertilization, as competition between pollen grains on te stigma and in te style, or after fertilization between te embryos(herrero&hormaza, 99; pasonenet al., 999) grains of many plant species germinate, wen placed in an artificial medium, forming a pollen tube tat extends by tip growt. Apart from non-permeant solutes (sucrose, polyetylene glycol, etc ), te medium need only contain te mineral ions os potassium, calcium and borate wit appropriate anions suc as pospate and nitrate or clorine (Weisenseelet al., 97) Te pollen tube is a cellular protuberance formed by te pollen grain, and is caracterized by a rapid and unidirectional growt. tubes grow at very rapid rates tat can reac up to.7 cm/ (scleiden, 9 Liliumlongiflorum, as a pollen tube tat grows at mm/ in vivo, wile tobacco pollen grows at.7 mm/ (sancezet al., ) It sould be mentioned tat pollen tube growt in vitro is muc slower tan tat of pollen growing in te style of te flower (Daer, ) II. HeliotropiumirsutissimumGrauer. Annual, covered trougout wit ± patent airs, especially on calyx. Leaves ovate to ovate- Lanceolate, obtuse, rarely subacute. Cymes manyflowered, very dense at first, becoming laxer and elongated later. Stigma squat.nutlets.glabrous, densely tuberculate. Distribution area: Bulgaria, Greece, Aegean, N. Africa, Syria, Palestine. (Davis, 97) Heliotropiumamultipotent medicinal erb belonging to te family Boraginaceae as a wide range of medicinal values. It as been used traditionally, like it is useful for local applications for ulcers, sores, wound, skin infections, stings of insects, and reumatism (Bagadekar&Jayaraj, ; Kumar &Rao, 7) Nutlet ISSN: Page

2 SSRG International Journal of Medical Science (SSRG-IJMS) Volume Issue 7 July 7 Stigma Total no.of germinated and non-germinated pollen grains flower stigma Style ovary For eac experiment, at least five images per sample were taken at random, positions and te percentage of germination was quantified. To determine pollen tube lengt, at least tirty tube were measured for eac experiment. Figure : HeliotropiumirsutissimumGrauer. (addad, ) III. MATERIALS AND METHODS. Plant material Te pollen used in all te essays related came from flowers of H.irsutissimum (Boraginaceae) from Syria (Latakia).. Germination medium We used Brewbaker and Kwac, s medium (9) containing: g/l sucrose, mg/l boric acid (HBo), mg/l Ca(No ).Ho, mg/l Mgso, mg/l KNo, g/l Agar. Wit a PH adjusted to. Agar was added to te mix and eated to dissolve. Hot agar containing medium was poured on to a petri dises ( cm diameter) to form a layer wit a tickness of a bout ( cm) and left to cool.. Study of different factors on germination, growt rate and pollen tube lengt We study te effect: - Temperature (-- C) - PH (--7-) - Sucrose (---)g/l -Boric acid (---) mg/l -Calcium (---) mg/l. Determination of te germination, growt rate and pollen tube lengt grains collected from te fresly deisced anters for germination on solid medium (petri dises) was ten sprinkled on te surface using (ml) from liquid growt medium witout agar, wit moving of dises. Te dises were placed in a umid camber wit different temperatures. grains, wic ad germinated and produced pollen tubes in different conditions was recorded (-----) minutes after te commencement of germination and percentage of pollen germination was calculated and te average lengt of pollen tube was recorded. Percentage of pollen germination was calculated by te following metod(amad et al., ) IV. Results and discussion. Te effect of temperature on pollen germination and pollen tube growt Wen pollen was distributed onto te germination medium, differents temperatures affected pollen viability under te tree incubation temperatures, and C, being C significantly iger. Table : Means of pollen germination and pollen tube growt in agar medium under te effects tree different temperatures Incubation temperature C germination (%) tube (µ m ) Te optimal temperature for in vitro germination assays can be species dependent. Te pollen from many species germinates well at c; owever, differences exist. For example, cotton pollen as an optimum germination temperature of c to c (Burke et al., ), to c in Agave genus (Lopes &Rodrigue, ), c in Erlenmeyer flasks (Boavidaet al., 7) Temperature is among te most important environmental factors affecting plant reproductive processes suc as pollen germination, pollen tube growt and fruit-set. In te present study, in vitro pollen germination and pollen tube growt of all cultivars were severely reduced under bot ig and low temperature conditions. Weaver and Timm (9) suggested tat pollen is more sensitive to ig temperatures tan femal reproductive organs, wic could account for a lack of fertilization under ig- temperatures stress. Te low temperature reduces te molecular mobility in te cytoplasm, wic may be a controlling factor in pollen longevity (Buitinket al., ). % pollen germination= No. of germinated pollen grains X ISSN: Page 9

3 SSRG International Journal of Medical Science (SSRG-IJMS) Volume Issue 7 July 7. Te effect of PH on pollen germination and pollen tube growt We used te basel medium after fixation te best temperature c, ten, we study te effect four PH on pollen germination and pollen tube growt. We observed, tere is no germination in a medium wit PH 7 and. In a medium wit PH, pollen grains were able to germinate, and te percentage of germination reaced a ig value tat at tis point of time was iger tan in a medium wit PH after our. Table : Means of pollen germination and pollen tube growt under te effects of four PH after PH 7 growt (µ m ) 9 Medium PH is critical condition for in vitro pollen tube growt. For lilium, solanum and camellia pollen tubes, te optimum PH is situated between and. Lower or iger PH values drastically reduce te percentage of germination and are unable to sustain pollen tube growt ( Cebli&Geitmann, 7) Since te pollen tube does not perform potosyntesis, a carbon source is required for energy supply and carboydrate skeleton formation and metabolized by te pollen grains or tubes (Montaner, ) During its development in te anter, pollen accumulates large quantities of carboydrates, wic constitute a large part of its dry weigt (Paciniet al., ) Sugar in te pistil ( or in pollen germination media) are necessary to maintain pollen tube growt (Labarca&Loewus, 97) Terefore, sucrose is generally added to pollen germination media, but te optimal concentration varies greatly between species. For instance, optimal papaver pollen growt in vitro occurs at % sucrose (Bowmik&Datta, ), Camellia at %, Lilium,Solanum and Tobacco (Nicotianatabacum) at % (Loguercio, ).. Te effect of Sucrose on pollen germination and pollen tube growt We used te basel medium after fixation te best temperature c and best PH, ten, we study te effect of sucrose (--- g/l). We observed tat in a medium wit g/l sucrose, pollen grains were able to germinate faster tan in a medium wit (- -) g/l sucrose. )Table, Figure ) Table : Means of pollen germination and pollen tube growt under te effects of sucrose Figure : Effect of Sucrose on pollen germination and tubes growt. Te effect of Boric acid on pollen germination and pollen tube growt We used te basel medium after fixation te best temperature c, best PH and sucrose g/l ten, we study te effect of Boric acid wit (,,, mg/l) We observed, In a medium wit mg/l, te percentage of germination reaced a value % wit µ m long pollen tubes. Sucros e g/l germination % growt (µ m ) Table : Means of pollen germination and pollen tube growt under te effects of Boric acid Boric acid mg/l growt (µ m ) 7 ISSN: Page

4 SSRG International Journal of Medical Science (SSRG-IJMS) Volume Issue 7 July 7 In te pollen tube, Boric acid is involved in cell wall formation and protein assembly into membranes and cell wall (Daer, ), toug its effect on H + - ATPase activity, Boric acid affects pollen germination, tube growt (Wang et al., ), it promotes absorption of sugar, and increases oxygen uptake, it is involved in te syntesis of pectic material for te wall of actively growing pollen tube (Amad et al., ) ppm boric acid was found to be essential for pollen germination (Brewbaker&kwack, 97), decrease of it concentration in species Tribulusterrestris, peac, Almond, piceameyeri, gave a negative results in pollen germination and formation abnormal tubes (Imaniet al., ) range (picton& steer, 9), tat varies between species (steer & steer, 99). Witin tis range, pollen tube tip extension rates are relatively insensitive to small canges in te calcium concentration (picton& steer, 9), wereas outside of tis range, growt is severely ampered.. Te effect of Calcium on pollen germination and pollen tube growt After fixation te best temperature c, PH, sucrose g/l Boric acid mg/l, we study te effect of Calcium wit (,,, mg/l) on pollen germination and pollen tube growt.we found, te percentage of germination in a medium wit mg/l of Calcium reaced a value 7.% wit µ m long pollen tubes. (Table, Figure ) Table : Means of pollen germination and pollen tube growt under te effects of Calcium Calciu m mg/l growt (µ m ) Calcium plays an essential role in pollen tube growt (Zou et al., ), an ionic current of calcium ions enters at te pollen tube tip as a result of te localized positioning of calcium ions cannels in te plasma membrane by secretory vesicle fusion at te tube apex. Tese processes create optimal conditions for growt mediated by te cytoskeleton at te calcium ion-entry site (steer, 99). Te presence of Ca + in te growt medium is known to be required for in vitro pollen germination and tip growt of most plant species. It plays a role in cell wall formation and rigidity, directs vesicle trafficking, controls actin dynamics (cebli&geitmann, 7). Normal pollen tube growt can only take place in te presence of a calcium concentration tat is situated witin a certain Figure : Effect of Calcium on pollen germination and tubes growt V. CONCLUSION different factors ave an essential roles in pollen germination and pollen tube growt.carbon source is required for energy supply and metabolized by te pollen grains, Boric acid and calciumare involved in cell wall formation and protein assembly into membranes and cell wall, PH and Temperature ave positive or negative effect on germination. REFERENCES [] S. Amad, A. Rana,R. Sarma, R. Agniorti, " Effect of different media and Boric acid on pollen germination and tube growt of Tribulusterrestris-A traditional medicinal plant ", (),. [] A.N. Bagadekar, M. Jayaraj, " Invitrorizogenesis from leaf and stem callus of Heliotropiumindicum, L.- Medicinal Herb ", International Journal of plant, Animal and Environmental Science, (), -,. [] S. Bomick, B. K. Datta, " In vitro pollen germination of EicbborniaCrassipes (Mart.) solms: An In sigt into its preferred mode of clonal reproduction ", (), -7,. [] L. C. Boavida, S. McCormick, " Temperature as a determinat factor for increased and reproducible in vitro pollen germination in Arabidopsis taliana", Te plant Journal, :7-, 7. [] J. L Brewbaker," Te distribution and pylogenetic significance of binucleate and trinucleate pollen grains in te angiospermis", American journal of Botany, 9-, 97. [] J. Buitink, O. Leprince, M. A. Hemminga, F. A. Hoekstra, " Te effect of moisture moisture and temperature on te ageing kinetics of pollen: interpretation based on cytoplasmic mobilityplant ", Cell and Environment, : 97-97,. ISSN: Page

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