Protocols for Micropropagation of Woody Trees and Fruits

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CHAPTER 18 MICROPROPAGATION OF BLACK LOCUST (ROBINIA PSEUDOACACIA L.) J. ZHANG, Y. LIU AND H. WANG* College of Biological Sciences and Biotechnology, Beijing Forestry University, Beijing 100083, China, ∗ Corresponding author, E-mail: hfwang @ bjfu.edu.cn 1. INTRODUCTION Robinia genus has about 10 species and 4 hybrids that are native to eastern North America and Mexico. They are Robinia boyntonii*, R. elliottii*, R. hartwegii* (R. viscosa var. hartwegii), R. hispida, R. kelseyi*, R. luxurians*, R. nana*, R. neomexicana, R. pseudoacacia, R. viscosa (*: not accepted as distinct by all authorities), R. × ambigua (R. pseudoacacia × R. viscose), R. × holdtii (R. neomexicana × R. pseudoacacia), R. × longiloba (R. hispida × R. viscose), and R. × margarettiae (R. hispida × R. pseudoacacia) (http://en.wikipedia.org/wiki/Robinia). Robinia pseudoacacia (black locust) is a commercial species. It is natively distributed in south eastern United States; on the lower slopes of the Appalachian Mountains, with separate outliers north along the slopes and forest edges of southern Illinois, Indiana, and Missouri (http://www.nps.gov/plants/alien/fact/rops1.htm). Black locust is the first forest tree species introduced in Europe from North America. The Parisian Botanical Garden introduced this species already in 1601 (as reviewed by Davis & Keathley, 1987). The species was introduced to Qingdao City of China in 1897 (Pan & You, 1994). Black locust is able to fix atmospheric nitrogen and enrich the soil fertility. The wood is valued for its durability and high fuel value. The tree provides good forage for bees and generally planted on reclaimed drought and barren lands to control erosion. It is also used for ornamental purposes. Moreover, its fast growth rate and great potential for environmental restoration have drawn attention to woody biomass plantation, establishment of energy plantation that would transfer existing traditional forests into energy forests (Rédei, 1999). The research is being carried out to improve wood properties of the species for construction, furniture, and other industrial uses (Molnár & Bariska, 2006). The wider 193 S.M. Jain and H. Häggman (eds.), Protocols for Micropropagation of Woody Trees and Fruits, 193–199. © 2007 Springer.
194 J. ZHANG ET AL. uses of wood have led to the nursery propagation and plantation forestry. However, the conventional plant propagation by grafting, cutting or root sucker is too slow to meet the needs of large-scale plantations. Micropropagation is an effective route to produce plants rapidly (Shu & Liu, 2003) and a strategy to minimize somaclonal variation among regenerated plants (Scowcroft, 1985). The explants collected from young black locust trees or twigs have high multiplication rate in vitro and the micropropagted shoots have high rooting efficiency (Barghchi & Chi, 1998). In this paper, we will describe the experimental protocol to micropropagate black locust efficiently. The protocol can also be utilized in germplasm conservation and genetic studies. 2.1. Preparation of Explants 2. EXPERIMENTAL PROTOCOL 2.1.1. Acquiring of Explants Well suited mother plants (Figure 1A) showing vigorous growth rates in the field are most appropriate for micropropagation of black locust. The young shoots sprouted from the lower trunk or from underground roots were collected in spring before budding (Kamlesh et al. 1995). About 30 to 40 cm long segments excised from the shoots were cultured in water in a phytotron. The water was changed every day. The temperature in the phytotron was between 24 and 26ºC, the relative humidity (RH) 70–80% during shoot cultivation, 16-h photoperiod and with light illumination at 80 μmol m –2 s –1 . In these conditions, the axillary buds sprouted after 14 days. Within 28 days, the new green shoots reached 3–4 cm in length. The new shoots were excised in 1.0–2.0 cm long segments with at least one axillary bud on each segment. Thereafter, they were used as explants (Figure 1B–C). 2.1.2. Surface Sterilization Shoot segments were cleaned by brushing with suds and rinsed by running tap water for 20–30 min. They were sterilized by dipping into 70% ethanol for 30 s, and then in 0.4% active chlorine of sodium hypochlorite solution (bleach) to which Tween-20 was added at the rate of one drop per 50 ml and shaken gently for about 20 min. The explants were rinsed by distilled sterile water three times (each for 3 min) and kept in the sterilized distilled water for 5 min in order to wash out residual bleach sticking to the plant material. All of these steps were carried out in a laminar flow hood. The shoot segments were individually dried on a sterilized filter paper in a Petri dish (10cm diameter). Some necrotic ends, which were probably caused by sterilization, were cut off. The explants were cultured on initiation medium (one explant per vessel) (for medium formula, see Table 2). 2.2. Culture Media and Conditions 2.2.1. Culture Media The black locust culture media were modified from the basic Murashige & Skoog medium (1962). The changes were mainly made in the concentrations of minerals, plant growth regulators and sucrose. The substances supplemented are listed in Table 2. Different compositions were used in initiation, proliferation and induction of roots.
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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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PROPAGATION OF SELECTED PINUS GENOT
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Page 152 and 153: ROOT INDUCTION OF PINUS SYLVESTRIS
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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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2.6. Flow Cytometry Analysis MICROP
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CHAPTER 43 MICROPROPAGATION OF BAMB
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BAMBOO MICROPROGATION BY AXILLARY S
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CHAPTER 44 IN VITRO CULTURE OF TREE
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IN VITRO CULTURE OF TREE PEONY 479
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500 M. MHATRE from various natural
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502 2.2. Disinfection of Plant Mate
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504 M. MHATRE soil in polythene bag
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506 M. MHATRE Figure 2. Pineapple,
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508 M. MHATRE Escalona, M., Lorenzo
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510 J.M. AL-KHAYRI Tunisia, Morocco
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512 J.M. AL-KHAYRI 2.1.2. Separatio
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514 2.2. Culture Medium J.M. AL-KHA
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516 J.M. AL-KHAYRI 5. Isolate callu
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518 J.M. AL-KHAYRI 3. Water the tra
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520 J.M. AL-KHAYRI expressed from c
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522 J.M. AL-KHAYRI alcohol and twic
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524 J.M. AL-KHAYRI potential. Simil
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526 J.M. AL-KHAYRI Barreveld, W.H.
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528 D.T. NHUT ET AL. arsenide chip
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530 D.T. NHUT ET AL. Figure 2. Plan
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532 D.T. NHUT ET AL. Figure 4. Plan
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534 D.T. NHUT ET AL. plantlets. The
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536 D.T. NHUT ET AL. Figure 11. Fre
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538 D.T. NHUT ET AL. Figure 13. Sub
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540 D.T. NHUT ET AL. The response o
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CHAPTER 48 IN VITRO MUTAGENESIS IN
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IN VITRO MUTAGENESIS IN BANANA 545
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3.3. Observations to be Recorded IN
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