Protocols for Micropropagation of Woody Trees and Fruits

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11. Determine the mean channel number of the sample G0/G1 nuclear DNA peak and that of internal reference standard. 12. Estimate the nuclear DNA content of the sample using the formula: sample 2C DNA content (pg) = PLANT GENETIC FIDELITY 73 sample G / G peak mean standard 0 1 G / G 0 1 peak mean × standard 2C DNAcontent If needed, the DNA content values can be converted in number of base pairs (bp), taking in consideration that 1 pg DNA = 978 Mbp (Doležel et al., 2003). The genome size in base pairs is usually shown in terms of haploid size (1C) of the genome, whether genome size in mass values are typically shown per 2C value. 13. Using the regions that were defined, record the full peak coefficient of variation (FPCV) of both FS and SS and the half peak coefficient of variation (HPCV) of sample and standard G0/G1 peaks. 14. Carefully and critically interpret the results. Small differences in the peak position of both samples and internal standard should be interpreted with caution as they may be due to instrumental drift or to the possible effect of cytosolic compounds. Whether in the first case, these differences may be reduced by planning the experience in at least three different days, to eliminate the second hypothesis and ascertain the real occurrence of DNA differrences, a new sample containing the tissues where the differences were obtained may be prepared. If only one peak is observed it is a strong indication that the observed differences were probably artefactual. 2.1.3. Troubleshooting 1. Empirical experience has strongly advised the chopping procedure instead of slicing for obtaining a high output of nuclei and little debris. Before the chopping step it is important that the plant tissue does not dry. Also, the chopping procedure should be fast (60 s of chopping should be enough). However, in some plant tissues containing compounds that affect histograms quality, chopping should be less intense. If only few nuclei are isolated, the amount of plant material can be increased. As an example, in Q. suber leaves chopping must consist of only several cuts and nuclear suspension should be quickly filtered. Callus tissue and somatic embryos of Q. suber are easier to be analysed than leaves. Friable callus (e.g. from Pinus pinaster) may be difficult or in some cases inappropriate for FCM analysis, as the method is only able to isolate a reduced number of nuclei. 2. After using the suggested buffer, if the obtained histograms present low resolution, other lysis buffers should be tested. A review on nuclear isolation buffers can be found in Greilhuber et al. (2007). (1)
74 C. SANTOS ET AL. Figure 1. A) Histogram of PI fluorescence and B) cytograms of forward-angle light scatter (FS LOG) vs. side-angle light scatter (SS LOG) both in logarithmic scale, C) PI fluorescence vs. time, D) SS LOG vs. PI and E) PI fluorescence pulse integral vs. PI fluorescence pulse height of nuclei of Quercus suber (peaks: A – G 0/G 1 nuclei, coloured in green, and C – G2 nuclei coloured in blue) and Glycine max (as internal reference standard with peaks B – G 0/G 1 nuclei, coloured in red, and D – G2 nuclei, coloured in purple) simultaneously isolated in WPB and stained with PI. Mean FL channel (FL), DNA index (DI=mean channel of Q. suber/mean channel of G. max) and HPCV % of each peak are given in the histogram A and mean channel of FS and SS and (Mean channel) FPCV % of each parameter are given in cytogram B.
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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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Page 44 and 45: MICROPROPAGATION OF PINUS PINEA L.
Page 50 and 51: 42 2.1. Organ Culture K. ISHII ET A
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Page 54 and 55: 46 Chemicals (mg/l) K. ISHII ET AL.
Page 56 and 57: 48 Table 3. Effects of plant growth
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Page 64 and 65: 56 2.1.3. Organogenesis from Field-
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Page 74 and 75: CHAPTER 7 GENETIC FIDELITY ANALYSES
Page 76 and 77: PLANT GENETIC FIDELITY 69 Plant mat
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Page 88 and 89: PLANT GENETIC FIDELITY 81 microprop
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Page 92 and 93: 86 2.1. Explant Preparation M.G. OS
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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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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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