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1.
In an outbreeding species such as apple, haploid plants may be especially useful in breeding programmes for the production of homozygous material. However, methods must be available to induce chromosome doubling in the haploid plants. Two antimitotic agents, colchicine and oryzalin, were compared as regards their efficiency in inducing chromosome doubling of in vitro haploid apple shoots. Three colchicine levels (0.025, 0.25 and 1.25 mM) and three oryzalin levels (5, 15 and 30 μM) were evaluated. Three techniques were also used and compared. Survival rate and chromosome counts were determined. Differences were observed between the two antimitotic agents and between the three techniques. This study demonstrates that oryzalin could be a better choice than colchicine for chromosome doubling on haploid apple shoots in vitro.  相似文献   

2.
Prem P. Jauhar 《Euphytica》2003,133(1):81-94
Allopolyploidy, resulting from interspecific and intergeneric hybridization accompanied by sexual doubling of chromosomes, has played a major role in the evolution of crop plants that sustain humankind. The allopolyploid species, including durum wheat, bread wheat, and oat, have developed a genetic control of chromosome pairing that confers on them meiotic regularity (diploid-like chromosome pairing), and hence reproductive stability, and disomic inheritance. Being natural hybrids, they enjoy the benefits of hybridity as well as polyploidy that make them highly adaptable to diverse environments. Despite the complexities of sexual reproduction, it is widespread among plants and animals. Sexual polyploids are highly successful in nature. Sexual polyploidization is far more efficient than somatic chromosome doubling. Sexual polyploidization effected by functioning of unreduced (2n) gametes in the parental species or in their hybrids has been instrumental in producing our grain, fiber, and oilseed crops. Evidence is presented for the occurrence of sexual polyploidization in durum haploids. ThePh1-induced failure of homoeologous pairing is an important factor in the formation of first division restitution(FDR) nuclei and 2n gametes. The evolutionary and breeding significance of sexual polyploidization is discussed. It is emphasized that three factors, viz.,sexual reproduction, allopolyploidy, and genetic control of chromosome pairing,jointly constitute a perfect recipe for cataclysmic evolution in nature. This revised version was published online in July 2006 with corrections to the Cover Date.  相似文献   

3.
Doubling the chromosome number of bahiagrass via tissue culture   总被引：2，自引：0，他引：2  
Crop improvement in bahiagrass (Paspalum notatum Flüggé) is limited by apomixis in most natural tetraploids, however, diploid sexual types occur. Production of sexual tetraploids by chromosome doubling will allow hybridization with apomictic tetraploids. Diploid bahiagrass (Paspalum notatum Flüggé) embryogenic callus tissue was exposed to three concentrations of three antimitotic chemical agents, colchicine, oryzalin and trifluralin. Callus was generated to plants and ploidy was evaluated by stomata size, mitotic chromosome counts, and flow cytometry. A total of 310 plants were verified as tetraploid of 1,432 plants that reached transplanting size. All treatments yielded 4x plants. The mean percentage success over all treatments was 22%, with means of 31% for oryzalin, 24% for colchicine and 16% for trifluralin. The high rates of success indicate that all agents can be successfully used to double chromosome numbers in bahiagrass. The percentage of 4x plants ranged from 9% (20 μM trifluralin) to 43% (20 μM oryzalin). Several treatments adversely affected regeneration. Mitotic chromosome counts are difficult and labor intensive in bahiagrass. Therefore, leaf stomata measurements were used as a preliminary screen. Data gave a bimodal distribution with overlapping tails and based on chromosome counts would have given an error rate of 12%. Flow cytometry analysis of regenerated plants resulted in mean nucleus fluorescence distributions consistent with control diploid or tetraploid values. These values agreed with chromosome counts, and this method is recommended for determining bahiagrass ploidy level. Research goals and available resources should be taken into consideration when selecting a treatment for chromosome doubling in bahiagrass.  相似文献   

4.
The efficiency of in vitro polyploidization depends on several variables associated to the plant, the antimicrotubule agent and the interactions between them. In the present work, we have used response-surface methodology to determine the best operating conditions for plant recovery in polyploidization assays for shoot apices and somatic embryos of two seedless grape cultivars, employing colchicine and oryzalin. Explant type, tubulin-interfering compound and concentration were the critical factors determining plant recovery. Linear reduction in viable plant regeneration via organogenesis and somatic embryogenesis was obtained by increasing oryzalin concentrations and treatment time, whereas the effects of colchicine were better described by a quadratic design for both explants types. The conditions promoting higher rates of plant recovery were used in chromosome doubling experiments with oryzalin and colchicine for shoot apices and somatic embryos of ‘Crimson seedless’ and ‘BRS Clara’. The established protocols allowed the recovery of non-chimerical autotetraploid plants at rates higher than 30 % for both cultivars. Stomata size parameters statistically correlate to the ploidy level of the regenerants and were effective for preliminary polyploidy screening. Autotetraploid lines of seedless grapes were incorporated into the Vitis germplasm bank for further agronomical evaluations. To our knowledge, this is the first report of in vitro oryzalin induced polyploidization of grapevine and of the use of mathematical modeling to optimize chromosome doubling in plants.  相似文献   

5.
The regeneration of haploid plantlets is considered as a bottleneck in rice anther culture. In this study, an antimitotic chromosome doubling method, simple and efficient, of androgenic haploid plantlets resulted in an efficient doubled haploid obtainment. Through chromosome doubling capacity comparison of the three antimitotic compounds (colchicine, trifluralin and oryzalin), colchicine at 500 and 625 mg/L without supplementing with DMSO was found to be the best antimitotic treatment, with a chromosome doubling capacity of 40%. Furthermore, the in vitro growth of plantlets was followed to analyse the effects of antimitotic compounds. Colchicine treatments were more toxic than dinitroanilines, and colchicine DMSO-supplemented treatments had significant lower values on shoot growth. On the other hand, dinitroaniline compounds impeded root growth, provoked helical growth of shoot and caused the apparition of white nodules in the base of the plantlet due to sprouting abortion. In this study, a protocol for doubled haploid plant recovery was established taking advantage from androgenic haploid plantlets in order to increase the number of doubled haploid plantlets produced after an anther culture protocol.  相似文献   

6.
The aim was to develop an efficient chromosome doubling method for Miscanthus sinensis to enable the production of triploids and so avoid seed dispersal to the environment. Antimitotic treatments with colchicine or oryzalin were tested in M. sinensis cl. MS‐88‐110 on: (1) in vitro shoots and plants established in soil; (2) during propagation of embryogenic callus; and (c) during the initial stages of callus induction. All systems produced chromosome‐doubled plants. A higher percentage of tetraploids was found after antimitotic treatment at the explant or callus level compared with treatment of in vitro shoots or plants established in soil. In general, oryzalin was more toxic to plant material than colchicine. A higher frequency of chimeras was found among plants with altered ploidy level when the target was formed shoot buds compared with adventitious shoot formation from callus. Antimitotic treatment of embryogenic callus from shoot apices also resulted in a high degree of albinism.  相似文献   

7.
This study describes an efficient in vitro method using colchicine-treated triploid seedlings of Napiergrass and Pearl millet hybrids to produce hexaploid hybrids and their subsequent identification with flow cytometry. It also describes chromosome count and stomatal morphology and their use to ploidy analysis. Four-hundred and eighty triploid seeds, representing 12 different hybrids were sterilized and transferred to MS media to induce chromosome doubling. Surviving plants were analysed by flow cytometry. From six triploid (control) and hexaploid plants, the stomata sizes and frequency were analysed. Chromosome count was performed only in the plants identified as hexaploid. Seventeen plants were identified as hexaploid by flow cytometry analysis. Further confirmation of the hexaploid condition was performed with stomatal morphology (stomatal frequency reduction and stomatal length increase) and chromosome count (2n = 6x = 42). Chromosome doubling has numerous applications in Pennisetum breeding. It can be used to restore the fertility of interspecific hybrids and to improve seed size.  相似文献   

8.
In this work, different treatment combinations including colchicine concentrations and treatment methods as well as different time points was used to investigate the survival rate and chromosome doubling efficiency of potato tissue culture plantlets. The potato chromosome doubling by using colchicine had been successfully optimized. The potato plantlet stems treated with 0.1% colchicine for three days shaking at 120 r min-1 showed the highest doubling efficiency due to its better contact to the colchicine solution. Compared with other potato chromosome doubling techniques, this method has much higher chromosome doubling rate, shorter time treatment and easier to operate, so that it could provide a higher efficient method for potato ploidy operation. In the meantime, compared with the diploid, the tetraploid interspecific hybrids showed differences in the morphological characteristic, which had higher plants, thicker stem, bigger petals and pollen grain. In addition, no significant difference was found between diploid and tetraploid interspecific hybrids in terms of cold resistance, but both significantly enhanced cold resistance compared with the common potato cultivar. Taken together, the doubled interspecific hybrids could sever for improving cold resistance of potato cultivars.  相似文献   

9.
In cereals, chromosome doubling of microspore-derived haploid plants is a critical step in producing doubled haploid plants. This investigation was undertaken to study the effect of incorporation of colchicine in the induction medium for anther culture, and the effect of colchicine on anther culture-derived plants of triticale grown under controlled greenhouse conditions. In the latter case, chromosome doubling of adult sterile plants derived from anther culture of fourteen triticale populations was attempted, where androgenetic plants with non-dehiscent anthers were cloned and subjected to the colchicine treatment, and then grown with the aid of hydroponics. The hydroponic system provided optimal conditions for recovery of the affected haploids from the toxic effects of colchicine treatment and all colchicine-treated plants survived. A topcross-F1 (TC1F1) population with timopheevii cytoplasm produced the highest percentage of plants with seed-set either due to chromosome doubling by colchicine (98%) or spontaneous doubling of chromosome number (15%). Colchicine-treated anthers performed inferior than control in both induction and regeneration phases. One of the key observation of this study was the reversal from reproductive stage back to the vegetative stage which in turn enabled further cloning of haploid plants under hydroponic conditions once they were identified as sterile. The one hundred percent survival rate of in vitro-derived plants, 100% survival rate of colchicine treated haploid plants and the high chromosome doubling success rate (X = 82.3) observed in this study imply that a temperature-controlled greenhouse with an hydroponic system provides an efficient environment for inducing chromosome doubling of haploid plants in cereals. This revised version was published online in August 2006 with corrections to the Cover Date.  相似文献   

10.
In vitro chromosome doubling during ovule culture of sugar and fodder beets (Beta vulgaris L.) was studied with four anti-microtubule herbicides: amiprophos-methyl (APM), oryzalin, pronamide, and trifluralin at concentrations of 0–300 μM. Best chromosome doubling results were obtained by treatment of the ovules with 100 μM APM which produced 4.7 diploid plants per 100 ovules. Highest chromosome doubling was found with oryzalin using 1 μM, with trifluralin at 10 μM, and with pronamide at 10 μM producing 2.8, 2.0, and 2.0 diploid plants per 100 ovules, respectively. The APM treatments showed relatively low toxicity on embryo formation which in combination with a high chromosome doubling effect, resulted in up to 89 diploids per 100 plants regenerated. Oryzalin and trifluralin had more severe toxic effects, which reduced embryo formation, thereby lower percentages of chromosome doubled plants were obtained from these treatments. Pronamide had no significant toxic effect but it induced chromosome doubling at lower frequencies. Compared to colchicine, APM seems to be as efficient for chromosome doubling during beet ovule culture, but at molar concentrations 100 times lower than those used for chromosome doubling with colchicine. This revised version was published online in July 2006 with corrections to the Cover Date.  相似文献   

11.
Cucumber is one of the most important vegetable crops worldwide, which makes it a good candidate to produce doubled haploid (DH) lines to accelerate plant breeding. Traditionally, these approaches involved induction of gynogenesis or parthenogenesis with irradiated pollen, which carries some disadvantages compared to androgenesis. Despite this, studies on anther/microspore cultures in cucumber are surprisingly scarce. Furthermore, most of them failed to unambiguously demonstrate the haploid origin of the individuals obtained. In this work we focused on anther cultures using two cucumber genotypes, different previously published protocols for anther culture, different in vitro culture variants to make it more efficient, and most importantly, a combination of flow cytometry and microsatellite molecular markers to evaluate the real androgenic potential and the impact of anther wall tissue proliferation. We developed a method to produce DH plants involving a bud pretreatment at 4 °C, a 35 °C treatment to anthers, culture with BAP and 2,4-D, and induction of callus morphogenesis by an additional 35 °C treatment and sequential culture first in liquid medium in darkness and second in solid medium with light. We also found that factors such as genotype, proliferation of anther wall tissues, orientation of anthers in the culture medium and growth regulator composition of the initial anther culture medium have a remarkable impact. Our rate of chromosome doubling (81%) was high enough to exclude additional chromosome doubling steps. Together, our results present androgenesis as an improvable but yet more convenient alternative to traditional gynogenesis and parthenogenesis-based approaches.  相似文献   

12.
Of 3272 plants regenerated from protoplasts of 10 Saintpaulia ionantha genotypes, 98.4% survived transfer to the greenhouse. The frequency of regenerants with chlorophyll deficiencies, i.e. variegated leaves or albinos, was low (1.5%). There was a higher number of polyploid, in most cases tetraploid plants, regenerated from protoplasts (16%) which were identified by their altered morphology. Measurements of stomatal length and counting the number of chloroplasts per guard cell also allowed a clear differentiation between diploid and polyploid plants. The classification was confirmed by DNA content determination using flow cytometry. Mechanisms leading to polyploidization included spontaneous protoplast fusion as well as chromosome doubling during callus growth and shoot regeneration. Two genotypes with instabilities in flower colour showed completely altered flower colours in plants regenerated from protoplasts as well as in plants regenerated on leaf explants in vitro.  相似文献   

13.
The production of doubled haploid (DH) lines has become a key technology in maize (Zea mays L.) research and breeding. However, most of the haploid plants are sterile and in many cases artificial chromosome doubling involves the use of costly and toxic chemicals. Here, we report a special kind of doubled haploid named the early doubled haploid (EH) that was generated directly by in vivo haploid induction. We found 83 EH plants induced from the hybrid Zhengdan958, 55 families of its F2:3 population and the parental lines, all of which were confirmed to be homozygous diploids via flow cytometry and 104 SSR markers. The progeny of EH0 (EH1) behaved in the same manner and showed the same potentialities as the parents of Zheng58 and Chang7-2. EH plants were also detected in other genetic backgrounds at a frequency of 1–3.5 % based on the total number of haploid plants. Because the EH lines exhibited completely fertility and were obtained from induction directly in one step, they could be used in DH breeding as a new breeding strategy. According to our observations, it is likely that spontaneous doubling in EH occurred during embryo development when haploid induction. The possible mechanism of EH is also discussed.  相似文献   

14.
Dinitroanilines represent a class of compounds that are widely used in herbicide formulations as they depolymerise plant microtubles, causing chromosome doubling. The potential of microtubule depolymerising herbicides trifluralin, oryzalin, and amiprophosmethyl (APM) for in vitro chromosome doubling of Rosa was studied. Five concentrations (0, 3, 6, 12 and 24 μM) and three exposure periods (12, 24 and 48 h) for each of the compounds were compared. Oryzalin, trifluralin and APM were not significantly different in their ability to induce chromosome doubling of R. hybrida cv Iceberg. At concentration of 6 μM and exposure period of 24 h, chromosome doubling of R. hybrida cv Iceberg was not significantly different with each of the polyplodising agents. At higher concentration (24 μM) and longer exposure period (48 h), 66.7% and 62.5% chromosome doubling was achieved with APM and trifluralin, respectively. However, the application of 6 μM oryzalin to R. persica (2n = 2x), R. hybrida cv Iceberg (2n = 3x) and R. hybrida cv Akito (2n = 4x), resulted in 60.0%, 6.3% and 0% chromosome doubling, respectively, which suggest that chromosome doubling is genotype dependent and plants with lower ploidy level have a higher propensity for chromosome doubling. Flow cytometry results at 18 and 24 weeks after herbicide treatment, indicated that the best time to test the treated plants was after 24 weeks.  相似文献   

15.
Sexual polyploidization via the formation of 2n gametes has been acknowledged as the most significant evolutionary mode of polyploidization among plant species. The present study was conducted in order to determine whether 2n gametes are present in the C-genome diploid Avena ventricosa Bal. ex Coss., a species that contributed to the evolution of the cultivated hexaploid species (Avena sativa L). Individual plants belonging to four different Cypriot populations, were screened for pollen grain size variation with the aim to distinguish 2n gametes. Avena ventricosa ARI00-845 was identified to produce large pollen grains at a low percentage (1.21%). Subsequent analysis using flow cytometry confirmed the presence of 2n gametes in the pollen. Cytogenetic analyses of pollen mother cells revealed cells with twice the typical chromosome number at metaphase I (i.e., 28 chromosomes). We postulate that irregularities in cell wall formation preceding meiosis could have contributed to the mode of chromosome doubling.  相似文献   

16.
芦笋单倍体染色体加倍技术研究   总被引：2，自引：1，他引：1  
研究了芦笋单倍体染色体加倍的方法。在离体培养条件下,以秋水仙素为诱变剂,分别用浸泡法和培养基添加法处理芦笋单倍体幼苗的茎尖,比较了秋水仙素不同浓度、不同处理时间的诱导效果。结果表明,培养基添加法的诱导效果好于浸泡法,当在培养基中添加0.3%秋水仙素并处理7天时诱导效果最佳,染色体加倍频率与成活率分别可达82.50%和80%。加倍后的二倍体植株与单倍体植株相比,茎干变粗,气孔与保卫细胞增大,保卫细胞内的叶绿体数增多。  相似文献   

17.
Chemically induced polyploids were obtained by the colchicine treatment of shoot tips of Humulus lupulus L. ‘Sybilla’. Flow cytometry revealed that most of the treatments resulted in the production of tetraploids. The highest number of tetraploids was obtained when explants were immersed in 0.05% colchicine for 48 h. A field experiment was conducted to compare diploid and tetraploid plants and assess the effect of genome polyploidization on the morphological and chemical characteristics. Tetraploids showed significant differences in relation to diploids. They had thinner and shorter shoots. The influence of chromosome doubling was also reflected in the length, width and area of leaves. The length of female flowers in the tetraploids was significantly shorter than that observed in diploids. Tetraploids produced a diverse number of lupuline glands that were almost twice as large as those observed in diploids. The most distinct effect of genome polyploidization was a significant increase in the weight of cones and spindles. Contents of major chemical constituents of hop cones was little affected by ploidy level. Total essential oils were significantly lower than those in diploids. However there was a significant increase in the proportion of humulene, caryophyllene and farnesene, oils desired by the brewing industry.  相似文献   

18.
S. S&#;kof    B. Bohanec    D. Kastelec    Z. Luthar 《Plant Breeding》2007,126(4):416-421
In the process of breeding triploid hop cultivars, diploid varieties need to be raised to the tetraploid level. Existing methods are predominantly based on treatment of the apices with antimitotic substances, in vivo or in vitro , resulting in a reported maximum of 26% tetraploids and a large proportion (over 60%) of mixoploids. This study was set-up to test whether an adventitious shoot regeneration protocol can be used as an alternative. In the first part of the experiments, regeneration media and a choice of explants (internodes, petioles and leaf discs) were optimized using three hop cultivars with putatively low, medium or high adventitious shoot regeneration capacity. On an optimized induction medium supplemented with 6 mg/l N 6-(2-isopentenyl) adenine (2iP), a regeneration rate up to 17.7% was obtained from cultured internodes, even of the most recalcitrant 'Aurora', while the regeneration rate of the most responsive 'Tettnanger' was 56.9%. Flow cytometric analysis of ploidy level revealed a high frequency of tetraploid induction, the highest (58.6%) being in 'Savinjski golding'. In 616 regenerants of all three cultivars, 30.4% were found to be tetraploid, the others were diploid. Cytokinin content was found to be of minor importance in tetraploid induction. The high frequency of spontaneous tetraploid induction and, in particular, the complete absence of mixoploids makes this protocol an alternative to the currently established methods of chromosome doubling based on the use of antimitotic substances. The occurrence of a high proportion of tetraploid adventitious regenerants in hop is discussed in view of certain other biotechnological applications.  相似文献   

19.
Isolated microspore cultures from two doubled haploid (DH) lines of wheat, Triticum aestivum L., were used to develop an in vitro chromosome-doubling protocol. During the initial 24 h or 48 h of culture the microspores were treated with either of the two antimicrotubule herbicides trifluralin or amiprophos-methyl (APM) in concentrations ranging from 0.1 μM to 10μM. Untreated control cultures yielded 209 embryos per 100000 microspores, which is the equivalent of one spike. Among the regenerated plantlets 67% were green, and 15% of the flowering plants were spontaneously chromosome doubled. Treatments with both the herbicides had a significant effect on chromosome doubling, measured as the percentage of fertile regenerants. With the best combination of treatment duration (48 h) and herbicide concentration (10/μM) the percentage of fertile plants among regenerants could be increased up to 74% with APM and up to 65% with trifluralin. The largest numbers of DH plants per spike could be obtained with herbicide concentrations at 1–3 μM. Treatments with either herbicide at these concentrations resulted in an estimated average between the two genotypes of 27 DH plants per 100 000 microspores. These results demonstrate the high potential of APM and trifluralin as chromosome-doubling agents in isolated microspore cultures. The in vitro treatment integrated into tissue culture procedures will constitute an efficient method for chromosome doubling in future wheat breeding  相似文献   

20.
为进一步了解秋水仙素对农作物根尖细胞染色体畸变的遗传毒害效应,以地方品种敏感型绿皮小粒蚕豆和饲用玉米为材料,研究不同浓度秋水仙素（0、0.01%、0.05%、0.10%、0.15%、0.20%）及不同培养时间（24、48、72h）对2种作物染色体的畸变影响。结果表明：秋水仙素能诱发根尖膨大,当秋水仙素浓度为0.20%处理24h,蚕豆根尖膨大率100%;秋水仙素浓度为0.20%处理72h,玉米根尖膨大率65.8%;适当浓度秋水仙素（0.01%、0.05%）可促进蚕豆和玉米根尖细胞的有丝分裂,当浓度≥0.10%,反而抑制蚕豆和玉米细胞分裂。在染色体畸变类型中,微核最多,其次是染色体断片,最少是染色体桥。秋水仙素浓度0.20%,处理时间72h,蚕豆畸变率和玉米畸变率均最高,分别为11.92%和7.25%;秋水仙素浓度0.10%,处理时间72h,蚕豆染色体加倍指数最高为7.97%;秋水仙素浓度0.20%,处理时间24h,玉米细胞加倍指数最高为4.64%。  相似文献   

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