Protocols for Micropropagation of Woody Trees and Fruits

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514 2.2. Culture Medium J.M. AL-KHAYRI 2.2.1. Basal Medium Components For medium used to induce adventitious organogenesis, the reader is referred to other protocols reviewed above since this protocol focuses on indirect somatic embryogenesis. In both cases, however, the basal medium commonly used for date palm tissue culture is based on MS salts formulation (Murashige & Skoog, 1962) with modifications by different authors to achieve genotype-specific and stage-dependent optimizations, by particularly manipulating the hormonal and vitamin content. Either commercially available prepackaged MS salt formulation or in-house prepared stock solutions can be used for medium preparation. Table 1 lists the components of MS medium and specifies other inorganic and organic additives. 2.2.2. Medium Preparation Prepare the basal medium including all additives except plant growth regulators and activated charcoal which are added according to culture stage as shown in Table 1. Adjust medium to ph 5.7 with 1 M KOH and HCl, and dispense in 150 × 25-mm culture tubes (15 ml per tube), 125-ml culture flasks (30 ml per flask), or GA-7 magenta vessels (50 ml per vessel). These vessels are autoclaved for 15 min at 121°C and 1 × 10 5 Pa (1.1 Kg cm –2 ) and kept in a cool place ready for culturing. Culture tubes are more suitable for culture initiation and rooting stages but the other vessels are more appropriate for callus multiplication and regeneration stages. 2.3. Callus Induction and Plant Regeneration 2.3.1. Callus Induction and Multiplication 1. Inoculate explants on culture initiation medium prepared according to Table 1. Place explants vertically with the basal end slightly inserted, 1–2 mm deep, into the medium (Figure 2 I). Because of potential latent internal contamination, a persistent problem in date palm tissue culture, it is advisable to culture explants in individual vessels throughout the procedure to limit cross contamination. 2. Incubate cultures at 23 ± 2°C in complete darkness for 9 weeks, during which transfer at a 3-week interval. Note explants swelling and slight expansion, a useful criterion to determine viability of newly cultured explants (Figure 3A, B). 3. While avoiding cutting the tissue to minimize browning, transfer entire explants to callus induction medium (Table 1) and maintain in darkness for 3 weeks. Callus formation becomes clearly visible. Note that auxin was switched from 2,4-D to NAA to reduce potential mutations. However, it is possible to use the initiation medium (Table 1) throughout callus stages from initiation to maintenance. 4. Again, transfer entire explants to callus proliferation medium (Table 1) and maintain in darkness for 9 weeks. More callus growth occurs that can be easily separated from the original explants at the end of this stage.
DATE PALM MICROPROPAGATION Table 1. Formulation of culture medium used for date palm micropropagation based on modified MS salt augmented with culture stage-specific plant growth regulators. Components Chemical formula Stock (g/L) Medium (mg/L) Major nutrients, 10 × stock, use 100 ml per L medium Ammonium nitrate NH4NO3 16.5 1650 Potassium nitrate Calcium chloride-2H2O KNO3 CaCl2 19.0 1900 . Magnesium sulfate-7H2O 2H2O MgSO4 4.4 440 . 7H2O 3.7 370 Potassium orthophosphate Sodium phosphate KH2PO4 NaH2PO4 1.7 170 . 2H2O 1.7 170 Minor nutrients, 100 × stock, use 10 ml per L medium Potassium iodide KI 0.083 0.83 Boric acid Manganese sulfate-4H2O H3BO3 MnSO4 0.62 6.2 . Zinc sulfate-7H2O 4H2O ZnSO4 2.23 22.3 . Sodium molybdate-2H2O 7H2O Na2MoO4 0.86 8.6 . Cupric sulfate-5H2O 2H2O CuSO4 0.025 0.25 . Cobalt chloride-6H2O 5H2O CoCl2 0.0025 0.025 . 6H2O 0.0025 0.025 Iron –EDTA, 100 × stock, use 10 ml per L medium Iron sulfate-7H2O FeSO4 . Ethylenediamine tetraaceticacid disodium 7H2O Na2EDTA 2.78 27.8 . 2H2O 3.73 37.3 Vitamins, 100 × stock, use 10 ml per L medium Myo-Inositol 12.5 125 Nicotinic acid 0.1 1 Pyridoxine hydrochloride 0.1 1 Thiamine hydrochloride 0.1 1 Glycine 0.2 2 Calcium pantothenate 0.1 1 Biotin 0.1 1 Glutamine Other additives 200 Ascorbic acid 100 Citric acid 100 Sucrose 40000 Agar 7000 Plant growth regulators and activated charcoal, add according to culture stage Final concentrations (mg/L) Culture stage 2,4-D 2iP NAA Charcoal Culture initiation 100 3 – 1500 Callus induction — 30 10 1500 Embryogenic callus — 6 10 1500 Callus multiplication — 1.5 10 — Embryogenesis — — — — Rooting — — 0.2 — 515
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PROTOCOLS FOR MICROPROPAGATION OF W
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A C.I.P. Catalogue record for this
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vi TABLE OF CONTENTS 14. Root induc
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viii TABLE OF CONTENTS 43. Micropro
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x PREFACE on precise stepwise proto
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CHAPTER 1 TOTIPOTENCY AND THE CELL
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TOTIPOTENCY AND THE CELL CYCLE 5 a
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2.2. Plant Bioregulators and the Ce
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TOTIPOTENCY AND THE CELL CYCLE 9 in
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TOTIPOTENCY AND THE CELL CYCLE 11 F
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Bartel, D. (2004) MicroRNAs: genomi
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CHAPTER 2 MICROPROPAGATION VIA ORGA
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MICROPROPAGATION OF SLASH PINE Tabl
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MICROPROPAGATION OF SLASH PINE Amon
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2.6. Field Testing MICROPROPAGATION
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CHAPTER 3 MICROPROPAGATION OF COAST
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MICROPROPAGATION OF SEQUOIA SEMPERV
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CHAPTER 4 MICROPROPAGATION OF PINUS
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MICROPROPAGATION OF PINUS PINEA L.
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42 2.1. Organ Culture K. ISHII ET A
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44 K. ISHII ET AL. scent light, 70
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46 Chemicals (mg/l) K. ISHII ET AL.
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48 Table 3. Effects of plant growth
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50 K. ISHII ET AL. Gupta, P.K. & Du
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52 C. HARGREAVES AND M. MENZIES (Me
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54 C. HARGREAVES AND M. MENZIES Con
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56 2.1.3. Organogenesis from Field-
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58 C. HARGREAVES AND M. MENZIES Fig
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60 C. HARGREAVES AND M. MENZIES Fig
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62 C. HARGREAVES AND M. MENZIES For
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64 C. HARGREAVES AND M. MENZIES and
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CHAPTER 7 GENETIC FIDELITY ANALYSES
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PLANT GENETIC FIDELITY 69 Plant mat
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e added to samples, as this fluoroc
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11. Determine the mean channel numb
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3. In addition to testing various b
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GeneScan internal size standards: 4
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3500 3000 2500 2000 1500 1000 500 0
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PLANT GENETIC FIDELITY 81 microprop
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PLANT GENETIC FIDELITY 83 Steinkell
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86 2.1. Explant Preparation M.G. OS
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88 WPM (Lloyd & McCown, 1980) Macro
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90 M.G. OSTROLUCKÁ ET AL. on the m
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CHAPTER 9 MICROPROPAGATION OF MEDIT
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2.1. Explant Preparation MICROPROPA
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MICROPROPAGATION OF MEDITERRANEAN C
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108 D.T. NHUT ET AL. Larue (1953) a
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110 D.T. NHUT ET AL. HgCl2 added wi
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112 2.4. Establishment of Shoot Cul
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114 D.T. NHUT ET AL. Table 5. Effec
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116 D.T. NHUT ET AL. culture to be
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118 D. EWALD ability of the plant m
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120 D. EWALD Figure 1. Yew micropro
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122 D. EWALD Root development. Afte
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CHAPTER 12 MICROPROPAGATION OF LARI
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MICROPROPAGATION OF LARIX SPECIES V
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CHAPTER 13 PROPAGATION OF SELECTED
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PROPAGATION OF SELECTED PINUS GENOT
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CHAPTER 14 ROOT INDUCTION OF PINUS
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ROOT INDUCTION OF PINUS SYLVESTRIS
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ROOT INDUCTION OF PINUS SYLVESTRIS
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CHAPTER 15 MICROPROPAGATION OF BETU
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MICROPROPAGATION OF BETULA PENDULA
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CHAPTER 16 PROTOCOL FOR DOUBLED-HAP
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DOUBLED-HAPLOID MICROPROPAGATION IN
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CHAPTER 17 IN VITRO PROPAGATION OF
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IN VITRO PROPAGATION OF FRAXINUS SP
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Axillary shoots (number) IN VITRO P
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CHAPTER 18 MICROPROPAGATION OF BLAC
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MICROPROPAGATION OF BLACK LOCUST 19
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2.5. Rooting and Acclimatization MI
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MICROPROPAGATION OF BLACK LOCUST 19
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202 V. RAJESWARI AND K. PALIWAL tha
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204 V. RAJESWARI AND K. PALIWAL Mak
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206 V. RAJESWARI AND K. PALIWAL 2.3
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208 V. RAJESWARI AND K. PALIWAL Fig
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210 2.8. Hardening V. RAJESWARI AND
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CHAPTER 20 MICROPROPAGATION OF SALI
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MICROPROPAGATION OF SALIX CAPREA L.
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CHAPTER 21 MICROPROPAGATION OF CEDR
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2.1. Establishment of Shoot Culture
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MICROPROPAGATION OF CEDRELA FISSILI
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238 programmes is somatic embryogen
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240 Figure 2. A) Induction of organ
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242 2.4.1. Rooting of Apical and Ba
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244 of donor individuals and/or by
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246 J. MALÀ ET AL. Karnosky, D.F.,
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CHAPTER 23 MICROGRAFTING IN GRAPEVI
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MICROGRAFTING IN GRAPEVINE 251 and
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MICROGRAFTING IN GRAPEVINE 253 Tabl
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2.4. Culture Conditions MICROGRAFTI
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MICROGRAFTING IN GRAPEVINE 257 Feuc
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CHAPTER 24 MICROGRAFTING GRAPEVINE
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MICROGRAFTING GRAPEVINE FOR VIRUS I
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CHAPTER 25 APRICOT MICROPROPAGATION
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APRICOT MICROPROPAGATION Figure 1.
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APRICOT MICROPROPAGATION Figure 2.
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APRICOT MICROPROPAGATION 2.2.2. Hyp
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2.4. Acclimatization APRICOT MICROP
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APRICOT MICROPROPAGATION occurs fre
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CHAPTER 26 IN VITRO CONSERVATION AN
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IN VITRO CONSERVATION AND MICROPROP
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CHAPTER 27 MICROGRAFTING OF PISTACH
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MICROGRAFTING OF PISTACHIO Constitu
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MICROGRAFTING OF PISTACHIO 2.2.2. C
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MICROGRAFTING OF PISTACHIO 6. Incub
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MICROGRAFTING OF PISTACHIO 9. The r
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CHAPTER 28 PROTOCOL FOR MICROPROPAG
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MICROPROPAGATION OF CASTANEA SATIVA
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CHAPTER 29 MICROPROPAGATION OF CASH
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MICROPROPAGATION OF CASHEW 2.2.1. E
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MICROPROPAGATION OF CASHEW axillary
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MICROPROPAGATION OF CASHEW Table 1.
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MICROPROPAGATION OF CASHEW shade an
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CHAPTER 30 IN VITRO MUTAGENESIS AND
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IN VITRO MUTAGENESIS AND MUTANT MUL
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336 R.I. IYER compounds (Iyer et al
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A 338 2.2. Culture Medium R.I. IYER
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340 R.I. IYER Figure 2. Formation o
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342 R.I. IYER Figure 3. Formation o
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344 R.I. IYER Rao, Y.S., Mathew, K.
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346 B.K. BISWAS AND S.C. GUPTA marg
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348 B.K. BISWAS AND S.C. GUPTA (iv)
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350 B.K. BISWAS AND S.C. GUPTA 2.4.
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352 B.K. BISWAS AND S.C. GUPTA Figu
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354 B.K. BISWAS AND S.C. GUPTA tree
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356 B.K. BISWAS AND S.C. GUPTA Figu
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358 B.K. BISWAS AND S.C. GUPTA Drew
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CHAPTER 33 MICROPROPAGATION PROTOCO
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MICROPROPAGATION FOR MICROSPORE EMB
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374 L. TIAN AND S.I. SIBBALD younge
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376 L. TIAN AND S.I. SIBBALD Table
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378 L. TIAN AND S.I. SIBBALD 2.4. R
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CHAPTER 35 MICROPROPAGATION OF JUGL
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MICROPROPAGATION OF JUGLANS REGIA L
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CHAPTER 36 TISSUE CULTURE PROPAGATI
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PROPAGATION OF MONGOLIAN CHERRY AND
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410 M. PASQUAL AND E.A. FERREIRA 2.
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414 M. PASQUAL AND E.A. FERREIRA ch
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418 D.T. NHUT ET AL. its economical
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420 D.T. NHUT ET AL. Table 1. Shoot
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422 D.T. NHUT ET AL. Figure 3. Orga
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424 D.T. NHUT ET AL. mg l -1 NAA +
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426 D.T. NHUT ET AL. Nakasone, H.Y.
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428 C. LIU ET AL. C. cujete L. is c
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430 C. LIU ET AL. Table 1. Composit
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432 C. LIU ET AL. 6. Excise the sho
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434 Fresh weight per plantlet (mg)
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436 C. LIU ET AL. 4. REFERENCES Aga
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438 M. MISHRA ET AL. micropropagati
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440 M. MISHRA ET AL. each plate ino
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Section C
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446 M.G. OSTROLUCKÁ ET AL. from me
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448 M.G. OSTROLUCKÁ ET AL. regener
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450 M.G. OSTROLUCKÁ ET AL. Figure
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452 M.G. OSTROLUCKÁ ET AL. 2.6.3.
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454 counts counts M.G. OSTROLUCKÁ
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CHAPTER 42 PROTOCOL FOR MICROPROPAG
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AN medium (Anderson, 1980) MICROPRO
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MICROPROPAGATION OF VACCINIUM VITIS
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Protocols for Micropropagation of Woody Trees and Fruits

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