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In vitro pollen germination in citrus

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Pollen viability is very important for breeders, geneticists and growers. It increases research efficiency and also help in section suitable Pollenizer for established orchards.

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In vitro pollen germination in citrus

  1. 1. In vitro Pollen Germination in Citrus PRESENTED BY: NOORANI GUL NABI KHAN, M2 DATE: 28-5-2015 ADVISOR: DR. CHITOSE HONSHO
  2. 2. Outline  Section 1: Introduction  Section 2: Receptiveness of the stigma and In vitro germination of orange pollen, submitted to different temperature • Knowledge gape • Material and Methods • Results and discussion • Conclusion  Section 3 : Assessment of pollen viability and germination in seven varieties of lemon • Knowledge gape • Material and Methods • Results and discussion • Conclusion  Section 4: Questions and Discussion  References
  3. 3. Section 1: Introduction Good quality, delicious taste and efficiency in consumption. How to produce Seedless Citrus fruits ? By Breeding approaches Interploidy Sexual Hybridization Required Control (Manual Pollination) followed by fertilization to set fruits. One factor that affects fruit setup is Pollen Viability
  4. 4. Pollen viability section 1 Pollen viability, tube growth and morphological homogeneity are important properties of quality pollen, useful for breeder, geneticists and breeders (Bolat and Pirlak, 1999). Advantages:  Increase the breeding efficiency  Help in selection of suitable pollinizer for established orchards. How to test the pollen viability in fruit crops ?
  5. 5. Pollen viability by staining tests SECTION 1 Many researchers investigated different staining tests to determine pollen viability of fruit crops. They are given below Acetocarmine or Alexander’s stain test Propone carmine Aniline blue IKI (Iodine potassium Iodide FDA (Fluorescent Diacetate) NBT (p-nitro blue tetrazolium) MTT (2, 5-diphenyletetrazolium bromide) TTC (2, 3, 5-triphenyletetrazolium chloride) Advantages: • Faster and easier compare to in vitro pollen germination test but in some cases different results may be obtained (Bolat, and Pirlak, 1999). •For accurate pollen viability, germination test is necessary.
  6. 6. In vitro Pollen Germination Section 1 Pollen of many flowering plants can be germinated on artificial media consisting of aqueous solutions of mineral salts (Brewbaker and Kwack, 1963). But The rate of pollen germination depending on the chemical concentration of medium and cultivar (Bolat, and Pirlak, 1999). Thus, suitable germination medium should be investigated for each cultivar of same species in fruit crops.
  7. 7. Basic Component of In Vitro Pollen germination Medium Section 1 Generally, pollen germination require 1. Sucrose (Tunistra and Wedel, 2000; Bair and Loomis, 1941 and DeBruyn, 1966a) 2. Boric acid (Bair and Loomis, 1941 and DeBruyn, 1966a) 3. Calcium ions (DeBruyn, 1966b) Additional component for pollen germination 1. Agar ( Provide stability ) 2. K+ ( Stimulatory effect on pollen germination and growth of tube) and so on Amount of the chemicals components can be vary, depending on the plant species (Cook and Walden, 1967; Sahar and Spiegel, 1984 and Taylor and Hepler, 1997). <PH of media, temperature and incubation time can also affect the pollen germination rate>
  8. 8. RECEPTIVENESS OF THE STIGMA AND IN VITRO GERMINATION OF ORANGE POLLEN, SUBMITTED TO DIFFERENT TEMPERATURES (Pio et al., 2004) Section 2 Knowledge gape: This study was conducted to find out the suitable temperature for in vitro pollen germination and stigma receptiveness in three orange cultivars (Valencia, Pera and Natal)
  9. 9. Material and Methods For stigma receptiveness experiment: w 3% Oxygenated (H2O2) water, 3 min The release of oxygen bubbles indicate the stigma is receptive Flower collection in three different growth stages
  10. 10. Experiment for germination temperature Dehiscence (26 C for 24 h ) 100 g/L Sucrose, 10g/l agar, 200mg/l boric acid, 800mg/l calcium nitrate, PH, 6.5 BOD chamber, 23, 24, 25, 26 C for 12h light period10 ml media
  11. 11. Results and discussion The enzymatic activity of peroxidase enzyme was greater in open flowers
  12. 12. Size, color and water droplet on stigma
  13. 13. Temperature
  14. 14. Conclusion 1. Valencia and Pera cultivars, manual pollen can be performed at the balloon stage 2. Natal cultivar, receptivity was greater in open flowers. Therefore, it can be pollinated during open stage of flowers when there is no risk of cross pollination. 3. The optimum temperature was 25C 4. Valencia has greater percentage of pollen germination.
  15. 15. Section 3 ASSESSMENT OF POLLEN VIABILITY AND GERMINATION IN SEVEN VARIETIES OF LEMON (Demir et al., 2015) Knowledge Gape: To determine the pollen viability by TTC and In vitro germination percentage of seven different lemon cultivars.
  16. 16. Materials and Methods Section 3 Dehiscence (23°C for 24 h ) One or two drop of TTC were dropped on a clear slides, and pollen were spread on it. Red stained pollen= Viable Light red= semi-viable Colorless= non-viable BATEM Sarisi, BATEM Pinari, Interdonato, Italyan Memeli, Kutdiken, Lamas, and Meyer. Control conditions for 3-4h (Norton, 1966). TTC Stain test for pollen viability
  17. 17. In vitro pollen germination on agar-plate Section 3 Five different conc. Of sucrose 5,10,15,20, 25% Adding 1% agar, 0.1% boric acid and incubated 21°C for 24h in dark
  18. 18. Results and Discussion Section 3 Pollen Viability
  19. 19. Pollen germination on agar-plat Pollen germination among cultivars were significant at all sucrose concentration The optimum sucrose concentration was 20% and 25%
  20. 20. Conclusion Section 3 1. Sucrose concentration affected the pollen germination of lemon varieties 2. The optimum sucrose concentrations were 20% and 25% in line with Ateyyeh (2005). 3. Staining tests provide only a rough estimate of pollen viability. 4. The exact amount of viable pollen maybe determine by in vitro pollen germination.
  21. 21. References •Demir, G., Turgutoglu, E. & Kurt, S., 2015. Assessment of pollen viability and germination in seven varieties of lemon, TURKEY: Journal of Crop Breeding and Genetics . •Pio, L. A. S. et al., 2004. RECEPTIVENESS OF THE STIGMA AND IN VITRO GERMINATION OF ORANGE POLLEN, SUBMITTED TO DIFFERENT TEMPERATURES, s.l.: Univeristy of Lavras.
  22. 22. Section 4: Questions and Discussion The end Thank you so much for your kind attention

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