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1.
Successful cryopreservation of Q. robur germplasm as plumules (i.e. shoot apical meristems of embryos) is described in this paper. After excision from the recalcitrant
seeds and preliminary storage in 0.5 M sucrose solution (18 h), the plumules were subjected to cryoprotection (in 0.75 M sucrose,
followed by 1.0 M sucrose and 1.5 M glycerol solutions), and next to desiccation (over silica gel or in nitrogen gas) and
cooling (in slush at –210°C or in vials filled with liquid nitrogen, LN, −196°C), and were then cryostored for 24 h. High
percentage of survival was obtained after cryostorage (21–67%, depending on pretreatment, assessed in vitro by greening plumules
that increased in size). Desiccation of plumules over silica gel resulted in significantly higher survival after cryopreservation
(58%) in comparison with desiccation in nitrogen gas (29%), with regrowth (shoots with leaves) 5–18%. The extent of plumule
desiccation was comparable in both methods, in which drying of plumules for 20 min decreased the water content to 0.5–0.6
g H2O g−1 dry weight before LN exposure. The type of LN exposure did not significantly influence plumule survival and regrowth after
cryostorage. Plumules isolated from acorns of four provenances survived cryostorage after cryoprotection followed by desiccation
over silica gel and direct cooling in vials with LN (survival 51–76%, regrowth 8–20%). Normal plants developed from the recovered
shoots after rooting. The presented protocol for Q. robur plumule cryopreservation may offer a potential approach for establishing germplasm conservation in gene banks for Quercus species. 相似文献
2.
Summary Plants of European chestnut (Castanea sativa) have been consistently recovered from cryopreserved in vitro-grown shoot apices by using the vitrification procedure. Factors found to influence the success of cryopreservation include
the source of the shoot tips (terminal buds or axillary buds), their size, the duration of exposure to the cryoprotectant
solution, and the composition of the post-cryostorage recovery medium. The most efficient protocol for shoot regrowth employed
0.5–1.0 mm shoot tips isolated from 1 cm-long terminal buds that had been excised from 3–5-wk shoot cultures and cold hardened
at 4°C for 2 wk. The isolated shoot tips were precultured for 2d at 4°C on solidified Gresshoff and Doy medium (GD) supplemented
with 0.2M sucrose, and were then treated for 20 min at room temperature with a loading solution (2M glycerol+0.4M sucrose) and for 120 min at 0°C with a modified PVS2 solution before rapid immersion in liquid nitrogen (LN). After 1 d in
LN, rapid rewarming and unloading in 1.2M sucrose solution for 20 min, the shoot tips were plated on recovery medium consisting of GD supplemented with 2.2 μM benzyladenine, 2.9 μM 3-indoleacetic acid, and 0.9 μM zeatin. This protocol achieved 38–54% shoot recovery rates among five chestnut clones (three of juvenile origin and two of
mature origin), and in all cases plant regeneration was also obtained. 相似文献
3.
Esther E. Uchendu Magfrat Muminova Sandhya Gupta Barbara M. Reed 《In vitro cellular & developmental biology. Plant》2010,46(4):386-393
Regrowth of plants after cryopreservation varies, and resulting regrowth ranges from poor to excellent. Oxidative stress is
a potential cause of damage in plant tissues. Antioxidants and anti-stress compounds may improve regrowth by preventing or
repairing the damage. Lipoic acid (LA), glutathione (GSH), glycine betaine (GB), and polyvinylpyrrolidone (PVP) were tested
during cryopreservation of shoot tips using the plant vitrification solution 2 (PVS2) protocol. Two in vitro-grown blackberry cultivars were cold acclimated and then cryopreserved in liquid nitrogen (LN). The antioxidant and anti-stress
compounds were added at four critical steps of the protocol: pretreatment, loading, rinsing, and regrowth. Three out of the
four compounds significantly improved regrowth of cryopreserved shoot tips. Regrowth ranged from 40% to 50% for controls to
>80% for treated shoot tips. LA (4-8 mM) produced high regrowth at pretreatment, loading, and rinsing for ‘Chehalem’ and at
all steps for ‘Hull Thornless’. Recovery improved at all steps with GSH (0.16 mM) and GB (10 mM). PVP had a neutral or negative
impact on regrowth. Overall addition of LA, GSH, and GB improved regrowth by ∼25% over the shoot tips cryopreserved using
the regular PVS2 protocol (control). This study shows that adding non-vitamin antioxidants and anti-stress compounds during
the PVS2-vitrification protocol improves regrowth of shoot cultures following cryopreservation. We recommend inclusion of
antioxidants as part of standard cryopreservation protocols. 相似文献
4.
Moges Asmara D. Shibli Rida A. Karam Nabila S. 《In vitro cellular & developmental biology. Plant》2004,40(4):389-395
Summary Cryopreservation of African violet via encapsulation-dehydration, vitrification, and encapsulation-vitrification of shoot
tips was evaluated. Encapsulation-dehydration, pretreatment of shoot tips with 0.3 M sucrose for 2 d followed by air dehydration for 2 and 4 h resulted in complete survival and 75% regrowth, respectively. Dehydration
of encapsulated shoot tips with silica gel for 1 h resulted in 80% survival but only 30% regrowth. Higher viability of shoot
tips was obtained when using a step-wise dehydration of the material rather than direct exposure to 100% plant vitrification
solution (PVS2). Complete survival and 90% regrowth were achieved with a four-step dehydration with PVS2 at 25°C for 20 min
prior to freezing. The use of 2M glycerol plus 0.4M sucrose or 10% dimethyl sulfoxide (DMSO) plus 0.5M sucrose as a cryoprotectant resulted in 55% survival of shoots. The greatest survival (80–100%) and regrowth (80%) was obtained
when shoot tips were cryoprotected with 10% DMSO plus 0.5M sucrose or 5% DMSO plus 0.75M sucrose followed by dehydration with 100% PVS2. Shoot tips cryoprotected with 2M glycerol plus 0.4M sucrose for 20 min exhibited complete survival (100%) and the highest regrowth (55%). In encapsulation-vitrification, dehydration
of encapsulated and cryoprotected shoot tips with 100% PVS2 at 25°C for 5 min resulted in 85% survival and 80% regrowth. 相似文献
5.
R. L. KNIGHT ELIZABETH KEEP J. B. BRIGGS JILL H. PARKER 《The Annals of applied biology》1974,76(1):123-130
Field and insectary tests confirmed that the black-currant gall mite (Cecidophyopsis ribis) is unable to survive on gooseberry and red currant. A dominant gene Ce, controlling resistance to the gall mite, has been transferred from gooseberry to black currant. Resistant, large-fruited, self-fertile black currants of commercial potential have been obtained in the third backcross. One accession of Ribes bracteosum and three of R. americanum proved field susceptible to the gall mite, but twenty-four accessions of other Ribes species remained free from galled buds for at least 3 years in an infection plot. 相似文献
6.
Cryopreservation of apple (Malus×domestica Borkh.) shoot tips following encapsulation-dehydration or encapsulation-vitrification 总被引:1,自引:0,他引:1
Axillary shoot tips of apple cv. Golden Delicious isolated from shoot cultures were successfully cryopreserved using the
encapsulation-dehydration technique. After encapsulation in alginate gel, embedded shoot tips were dehydrated by exposure
to a sterile air flow before being frozen in liquid nitrogen and subsequent slow thawing. A preculture on modified MS medium
containing 0.75 M sucrose followed by 6 h of dehydration (21% residual water) led to the highest shoot regrowth of frozen, coated shoot tips
(83.7%). Among the sugars tested, sucrose and sorbitol presented the best cryoprotective effect. Four other scion apple varieties
and rootstocks were also successfully cryopreserved. Axillary shoot tips of five apple (Malus×domestica Borkh.) scion and rootstock cultivars were cryopreserved using the encapsulation-vitrification technique. Using a one-step
freezing method, we successfully cryopreserved axillary shoot tips without the requirement of a cold hardening pretreatment
of the shoot cultures. Cryopreserved shoot tips treated with aqueous cryoprotective mixture IV containing 180% (w/v) sucrose
and 120% (v/v) ethylene glycol showed the highest shoot regrowth rates, which varied from 64% to 77%, depending on the cultivar.
Received: 29 July 1999 / Revision received: 24 September 1999 / Accepted: 26 November 1999 相似文献
7.
Vitamins C and E improve regrowth and reduce lipid peroxidation of blackberry shoot tips following cryopreservation 总被引:2,自引:0,他引:2
Esther E. Uchendu Scott W. Leonard Maret G. Traber Barbara M. Reed 《Plant cell reports》2010,29(1):25-35
Oxidative processes involved in cryopreservation protocols may be responsible for the reduced viability of tissues after liquid
nitrogen exposure. Antioxidants that counteract these reactions should improve recovery. This study focused on oxidative lipid
injury and the effects of exogenous vitamin E (tocopherol, Vit E) and vitamin C (ascorbic acid, Vit C) treatments on regrowth
at four critical steps of the plant vitrification solution number 2 (PVS2) vitrification cryopreservation technique; pretreatment,
loading, rinsing, and regrowth. Initial experiments showed that Vit E at 11–15 mM significantly increased regrowth (P < 0.001) when added at any of the four steps. There was significantly more malondialdehyde (MDA), a lipid peroxidation product,
at each of the steps than in fresh untreated shoot tips. Vit E uptake was assayed at each step and showed significantly more
α- and γ-tocopherols in treated shoots than those without Vit E. Vit E added at each step significantly reduced MDA formation
and improved shoot regrowth. Vit C (0.14–0.58 mM) also significantly improved regrowth of shoot tips at each step compared
to the controls. Regrowth medium with high iron concentrations and Vit C decreased recovery. However, in iron-free medium,
Vit C significantly improved recovery. Treatments with Vit E (11 mM) and Vit C (0.14 mM) combined were not significantly better
than Vit C alone. We recommend adding Vit C (0.28 mM) to the pretreatment medium, the loading solution or the rinse solution
in the PVS2 vitrification protocol. This is the first report of the application of vitamins for improving cryopreservation
of plant tissues by minimizing oxidative damage. 相似文献
8.
A protocol was developed for short-term preservation and distribution of the medicinal and timber plantation tree, Khaya senegalensis, using alginate-encapsulated shoot tips. The study assessed the effects of culture medium, storage temperature, auxin concentration
and planting substrate on shoot regrowth or conversion into plantlets of four different clones. Optimal shoot growth was obtained,
with high frequencies (92–100%) of shoot emergence, on Murashige and Skoog (MS) culture media containing 4.4 μM benzyladenine
(BA). Encapsulated shoot tips survived longer at 25°C than at 4°C, with viability of 73–88% after 8 weeks. Conversion into
plantlets was achieved on half-strength MS medium by pre-culture treatment of shoot tips with 49–490 μM indole-3-butyric acid
(IBA) before encapsulation. Treatment with 245 μM IBA provided 52–98% conversion, and 90–95% of plantlets survived acclimatisation
under nursery conditions. To eliminate the in vitro culture step after encapsulation, synthetic seeds were allowed to pre-convert
before sowing directly onto a range of ex vitro non-sterile substrates. Highest frequencies of plantlet formation from pre-converted
synthetic seeds (42–86%) were obtained by transferring synthetic seeds to organic compost, and these plantlets exhibited almost
100% survival in the nursery without mist irrigation. Pre-conversion is a novel and convenient method for producing synthetic
seeds that are suitable for distribution to commercial nurseries. 相似文献
9.
A protocol was developed for short-term preservation and distribution of the plantation eucalypt, Corymbia torelliana × C. citriodora, using alginate-encapsulated shoot tips and nodes as synthetic seeds. Effects of sowing medium, auxin concentration, storage
temperature and planting substrate on shoot regrowth or conversion into plantlets were assessed for four different clones.
High frequencies of shoot regrowth (76–100%) from encapsulated explants were consistently obtained in hormone-free half- and
full-strength Murashige and Skoog (MS) sowing media. Conversion into plantlets from synthetic seeds was achieved on half-strength
MS medium by treating shoot tips or nodes with 4.9–78.4 μM IBA prior to encapsulation. Pre-treatment with 19.6 μM IBA provided
62–100% conversion, and 95–100% of plantlets survived after acclimatisation under nursery conditions. Synthetic seeds containing
explants pre-treated with IBA were stored for 8 weeks much more effectively at 25°C than at 4°C, with regrowth frequencies
of 50–84% at 25°C compared with 0–4% at 4°C. To eliminate the in vitro culture step after encapsulation, synthetic seeds were
allowed to pre-convert before sowing directly onto a range of ex vitro non-sterile planting substrates. Highest frequencies
(46–90%) of plantlet formation from pre-converted synthetic seeds were obtained by transferring shoot tip-derived synthetic
seeds onto an organic compost substrate. These plantlets exhibited almost 100% survival in the nursery without mist irrigation.
Pre-conversion of non-embryonic synthetic seeds is a novel technique that provides a convenient alternative to somatic embryo-derived
artificial seeds. 相似文献
10.
T. Ganino A. Silvanini D. Beghé C. Benelli M. Lambardi A. Fabbri 《Biologia Plantarum》2012,56(1):78-82
This study was carried out on Kober 5BB (Vitis Berlandieri × V. riparia) grape rootstock shoot tips during the preparatory steps preceding the direct immersion in liquid nitrogen, in order to overcome
until now unsuccessful cryopreservation with this species. The exposure of shoot tips to 0.3–0.4 M sucrose leads to a high
cell solute concentration. The treatment with plant vitrification solution (PVS2) alone, i.e., not followed by storage in liquid nitrogen, markedly affected shoot tip survival. After a 30 min exposure, regrowth percentage
of shoot tips decreased from 94 % (control) to 57 %, and dropped to 15 % when the treatment was prolonged up to 60 min. After
a 90 min exposure, no regrowth occurred. In addition, plantlets regenerated from shoot tips which underwent 60 min or more
exposure to PVS2 showed signs of malformation. Microscope observations of shoot tips treated with 0.3 or 0.4 M sucrose and
30 min PVS2 showed the presence of cells starting to plasmolyze, localized in the area surrounding the apical meristem. A
limited presence of starch grains in meristem and bract cells was also noted. However, the most conspicuous consequence of
prolonged PVS2 treatment was convex plasmolysis. The phenomenon was dependent on the time of PVS2 exposure. Indeed, after
a 30 min treatment, plasmolysis was minimal or absent, but it increased with longer exposure to PVS2 at 4 °C. 相似文献
11.
V. W. L. JORDAN 《The Annals of applied biology》1968,61(3):399-406
It is shown that conidia of Sphaerotheca mors-uvae (Schw.) Berk, from gooseberry readily infect black currant; therefore it is unlikely that a new race of S. mors-uvae specifically pathogenic to black currants has recently appeared. However, the life-cycle on black currants differs from that on gooseberry. Incubation, infection and sporulation of the fungus have been examined under the optimum conditions of 18 °C and 100% relative humidity. The establishment of infections and sporulation was encouraged by a relatively low soil moisture content, temperatures above 15 °C, 60% relative humidity and good illumination-factors which promote vigorous plant growth. High phosphorus and high potassium nutrition also increased the susceptibility of black currants to infection. 相似文献
12.
Shizheng Deng Zhongdong Dong Kehui Zhan Yanmin Hu Dongmei Yin Dangqun Cui 《In vitro cellular & developmental biology. Plant》2009,45(1):99-103
Tissue and cell culture systems are vital to many areas of maize research and improvement. Efficient shoot regeneration remains
a limiting factor for most elite lines. To enhance shoot regeneration, calluses derived from immature embryos of four genotypes
were subjected to 6, 12, 24, 48, and 72 h of desiccation on sterile filter paper before shoot induction. We achieved up to
32% desiccation (measured as mass) and 48% imbibition (measured as mass gain in 48 h incubation) which was 3.4-fold higher
than nondesiccated controls. This rapid desiccation/imbibition procedure enhanced shoot regeneration and development, but
its efficiency was crucially dependent on the degree and duration of desiccation. All four genotypes calluses regenerated
shoots best after 48 h of desiccation corresponding to a 23.7% average desiccation percentage, and regeneration frequencies
reached 42–74%, which increased 1.5–2.1-fold compared with that of nondesiccated calluses. The effect of desiccation treatment
on the shoot regeneration increase took place mainly during the early phase of induction. The weekly average shoot regeneration
frequency of calluses with 48 h desiccation in the first wk reached 8.8% (the control was no regeneration); then it increased
3.0 and 2.3-fold in the second and third wk induction, respectively. We demonstrate that the moderate desiccation of calluses
increases the yield of shoot regeneration and speeds up regeneration course in maize. 相似文献
13.
Sarab A. Sharaf Rida A. Shibli Mahmoud A. Kasrawi Savinaz H. Baghdadi 《Plant Cell, Tissue and Organ Culture》2012,108(3):437-444
Artemisia herba-alba, called Shih is a medicinal herbal plant found in the wilds. The biodiversity of this plant is heavily subjected to loss because
of heavy grazing, land cultivation and collection by people to be used in folk medicine. In the current study, two cryopreservation
dependent techniques to conserve the shoot-tips of in vitro grown Shih were evaluated: encapsulation- dehydration and encapsulation-
vitrification. Shoot-tips of Shih were encapsulated into sodium-alginate beads. In encapsulation- dehydration, the effect
of sucrose concentration (0.5, 0.75 or 1.0 M) and dehydration period (0, 2, 4 or 6 h) under sterile air-flow on survival and
regrowth of encapsulated shoot tips were studied. Maximum survival (100%) and regrowth (27%) rates were obtained when encapsulated
unfrozen Artemisia herba-alba shoot tips were pretreated with 0.5 M sucrose for 3 days without further air dehydration. After cryopreservation the highest
survival (40%) and regrowth (6%) rates were achieved when Artemisia herba-alba shoot tips were pretreated with 1.0 M sucrose for 3 days without further air dehydration. Viability of Artemisia herba-alba shoot tips decreased with increased dehydration period. In encapsulation-vitrification, the effect of dehydration of encapsulated
Artemisia herba-alba shoot tips with 100% PVS2 for various dehydration durations (10, 20, 30, 60 or 90 min) prior to freezing was studied. After
cryopreservation the dehydration of encapsulated and vitrified shoot tips with 100% PVS2 for 30 min resulted in 68% survival
and 12% regrowth rates. Further conservation techniques must be evaluated to increase both survival and regrowth percentages. 相似文献
14.
Plant regeneration from Rosa shoot tips cryopreserved by a combined droplet vitrification method 总被引:1,自引:0,他引:1
Shoot tips obtained from in vitro Rosa plants (three cultivars) were successfully cryopreserved by a combined droplet vitrification method and subsequently shoots
regenerated. The excised shoot tips (1–4 mm long) were incubated in a liquid MS medium supplemented with 2.5 mg l−1 thiamine, 0.2 mg l−1 biotin, 0.2 mg l−1 pyridoxine, 0.25 mg l−1 6-benzylaminopurine (BAP), 0.5 mg l−1 gibberellic acid (GA3) and 0.08 M sucrose, for 24 h. Following that incubation shoot tips were pre-cultured in this MS medium containing 0.1 till
1.0 M sucrose for 24 and 48 h, respectively. Pre-cultured shoot tips were dehydrated with concentrated PVS2 cryoprotective
solution for 10–30 min at room temperature, prior to a direct plunge in liquid nitrogen. After rapid rewarming in the above
mentioned liquid medium shoot tips were plated on a modified MS medium (5 g l−1 agar) supplemented with vitamins and plant growth regulators as mentioned above for regrowth. Cryopreserved shoot tips resumed
growth within 10 days and regenerated shoots within 3 weeks. The highest numbers of regrowing shoot tips were 64.44% for cv.
Kardinal, 67.73% for cv. Fairy and 57.57% for cv. Maidy. 相似文献
15.
Alginate encapsulation is a simple and cost-effective technique to preserve plant germplasm but there are only a few reports
available on preservation of encapsulated explants of two highly valuable groups of tropical trees, the eucalypts (Myrtaceae)
and mahoganies (Meliaceae). This study investigated alginate encapsulation for preservation of the eucalypt hybrid, Corymbia torelliana × C. citriodora, and the African mahogany, Khaya senegalensis. We assessed shoot regrowth of encapsulated shoot tips and nodes after storage for 0, 3, 6 and 12 months on media varying
in sucrose and nutrient content, under storage conditions of 14°C and zero-irradiance. Encapsulated explants of both trees
were preserved most effectively on high-nutrient (half-strength Murashige and Skoog) medium containing 1% sucrose, which provided
very high frequencies of shoot regrowth (92–100% for Corymbia and 71–98% for Khaya) and excellent shoot development after 12 months’ storage. This technique provides an extremely efficient means for storage
and exchange of eucalypts and mahoganies, ideally suited for incorporation into plant breeding and germplasm conservation
programs. 相似文献
16.
In vitro-grown shoot tips of Emmenopterys henryi Oliv. were successfully cryopreserved by vitrification. Shoot tips excised from 3-month old plantlets were precultured in
a liquid hormone-free Murashige and Skoog (MS) medium supplemented with 0.5 M sucrose for 3 days at 25°C and then treated
with a mixture of 2 M glycerol plus 0.4 M sucrose (LS solution) for 40 min at 25°C. Osmo-protected shoot tips were first dehydrated
with 60% vitrification solution (PVS2) for 30 min at 0°C and followed by 100% PVS2 for 40 min at 0°C. After changing the solution with fresh 100% PVS2, the shoot tips were directly plunged into liquid nitrogen. After rapid warming in a water-bath at 40°C for 2 min, the shoot
tips were washed for 20 min at 25°C with liquid MS medium containing 1.2 M sucrose and then transferred onto solid MS medium
supplemented with kinetin 2 mg l−1, α-naphthaleneacetic acid 0.1 mg l−1, 3% (w/v) sucrose and 0.75% (w/v) agar. The shoot tips were kept in the dark for 7 days prior to exposure to the light (12 h
photoperiod cycle). Direct shoot elongation was observed in approximately 12 days. The regeneration rate was approximately
75–85%. This method appears to be a promising technique for cryopreserving shoot tips of Emmenopterys henryi Oliv. germplasm. 相似文献
17.
In vitro-grown shoot tips of the LN33 hybrid (Vitis L.) and cv. Superior (Vitis vinifera L.) were successfully cryopreserved by encapsulation-dehydration. Encapsulated shoot tips were precultured stepwise on half-strength
MS medium supplemented with increasing sucrose concentrations of 0.25, 0.5, 0.75 and 1.0 M for 4 days, with one day for each
step. Following preculture, encapsulated shoot tips were dehydrated prior to direct immersion in liquid nitrogen for 1 h.
After thawing, cryopreserved shoot tips were post-cultured on a post-culture medium for survival. An optimal survival of cryopreserved
shoot tips was achieved when encapsulated shoot tips were dehydrated to 15.6 and 17.6% water content for the LN33 hybrid and
cv. Superior, respectively. Comparison between the effects of dehydration with silica gel and by air drying on cryopreserved
shoot tips, showed that survival was dependent on water content, not on dehydration method. The thawing method markedly affected
survival of cryopreserved shoot tips, and thawing at 40 °C for 3 min was found best. No callus formation and fastest shoot
elongation were obtained when cryopreserved shoot tips were post-cultured on the post-culture medium composed of half-strength
MS supplemented with 1 mg l−1 BA and 0.1 mg l−1 NAA. With these optimized parameters, 60 and 40% survival of cryopreserved shoot tips were obtained for the LN33 hybrid and
cv. Superior, respectively.
This revised version was published online in June 2006 with corrections to the Cover Date. 相似文献
18.
Sugar beet shoot tips from cold-acclimated plants were successfully cryopreserved using a vitrification technique. Dissected
shoot tips were precultured for 1 day at 5 °C on solidified DGJ0 medium with 0.3 M sucrose. After loading for 20 min with
a mixture of 2 M glycerol and 0.4 M sucrose (20 °C), shoot tips were dehydrated with PVS2 (0 °C) for 20 min prior to immersion
in liquid nitrogen. Both cold acclimation and loading enhanced the dehydration tolerance of shoot tips to PVS2. After thawing,
shoot tips were deloaded for 15 min in liquid DGJ0 medium with 1.2 M sucrose (20 °C). The optimal exposure time to both loading
solution and PVS2 depended on the in vitro morphology of the clone. With tetraploid clones a higher sucrose concentration
during cold acclimation and preculture further enhanced survival after cryopreservation. Survival rates ranged between 60%
and 100% depending on the clone. Since only 10–50% of the surviving shoot tips developed into non-hyperhydric shoots, regrowth
was optimized.
Received: 13 September 1999 / Revision received: 2 March 2000 / Accepted: 16 March 2000 相似文献
19.
Samia S. Al-Ababneh Nabila S. Karam Rida A. Shibli 《In vitro cellular & developmental biology. Plant》2002,38(6):602-607
Summary The objective of this study was to establish a cryopreservation protocol for sour orange (Citrus aurantium L.). Cryopreservation was carried out via encapsulation-dehydration, vitrification, and encapsulation-vitrification on shoot
tips excised from in vitro cultures. Results indicated that a maximum of 83% survival and 47% regrowth of encapsulated-dehydrated and cryopreserved
shoot tips was obtained with 0.5M sucrose in the preculture medium and further dehydration for 6 h to attain 18% moisture content. Dehydration of encapsulated
shoot tips with silica gel for 2h resulted in 93% survival but only 37% regrowth of cryopreserved shoot tips. After preculturing
with 0.5M sucrose, 80% of the vitrified cryopreserved shoots survived when 2M sucrose plus 10% dimethyl sulfoxide (DMSO) was used as a cryoprotectant for 20 min at 25°C. Survival and regrowth of vitrified
cryopreserved shoot tips were 67% and 43%, respectively, when 0.4M sucrose plus 2M glycerol was used as a loading solution followed by application of 100% plant vitrification solution (PVS2) for 20 min. Increased
duration of exposure to the loading solution up to 60 min increased survival (83%) and regrowth (47%) of cryopreserved shoot
tips. With encapsulation-vitrification, dehydration with 100% PVS2 for 2 or 3 h at 0°C resulted in 50 or 57% survival and
30 or 40% regrowth, respectively, of cryopreserved shoot tips. 相似文献
20.
The ability of shoot tips from carnation (Dianthus caryophyllus L., var. Eolo) cultured in vitro to develop resistance to freezing in liquid nitrogen depends on the physiological state of the cell material and the pretreatment conditions. Regrowth rates close to 100% have been obtained with apical shoot tips isolated from 2 month-old stems, precultured on medium supplemented with sucrose (0.75M) and treated with dimethylsulfoxide (5% or more). Resistance of axillary shoot tips decreased progressively as a funtion of their distance from the apical shoot tip. During the development of the stem from axillary buds (obtained by cutting), progressive increases in the regrowth rate of frozen apices were noted, from 30% before cutting (axillary buds) to 98% after 3 weeks of culture.Abbreviations DMSO
dimethylsulfoxide
- LN
liquid nitrogen 相似文献