Multiple shoot regeneration of kenaf (Hibiscus cannabinus L.) from a shoot apex culture system

 In this research, a medium was developed that would stimulate multiple shoot initiation from shoot apex explants of Hibiscus cannabinus L. (kenaf). Adventitious shoot formation on a shoot induction media supplemented with combinations of 2,4-dichlorophenoxyacetic acid (2,4-D) (0, 0.5, 2.3 μmol·l^–1) and thidiazuron (N-phenyl-N′-1,2,3-thiadiazol-5-ylurea; TDZ) (0, 1, 5, 20 μmol·l^–1) was evaluated. Multiple shoot induction medium with 1 μmol·TDZ l^–1 resulted in the highest number of regenerated shoots per explant for all three kenaf cultivars tested (Tainung 1, Tainung 2, and Everglades 71). The 2,4-D did not enhance multiple shoot formation. Additionally, kenaf cultivars 7N and SF459 also produced multiple shoots on the induction medium with 1 μmol·TDZ l^–1. Multiple shoot clumps formed after 2 weeks in culture without callus formation. Shoots elongated and rooted within 2 weeks after subculture on a plant growth regulator-free medium. A histological study demonstrated the de novo regeneration of shoots from the shoot apex. Received: 2 February 2000 / Revision received: 30 March 2000 / Accepted: 22 June 2000     

Genetic diversity and phylogenetic relationship of kenaf (Hibiscus cannabinus L.) accessions evaluated by SRAP and ISSR

Understanding genetic diversity and phylogenetic relationships is useful for plant breeding. In this study, we assessed the genetic diversity in a panel of 84 accessions of kenaf from 26 countries using SRAP and ISSR markers. The kenaf accessions could be divided into L1 (60 cultivated varieties) and L2 (24 wild accessions) at the level of 0.145 genetic dissimilarity coefficient by UPGMA. The L₂ group was further divided into two subgroups (16 relative-wide and 9 origin wide accessions) at the level of 0.207 genetic dissimilarity. Out of the 9 wild accessions in the L₂ group, 6 were from Tanzania and the remaining 3 lines were from Kenya. These results suggest that the center of origin for kenaf might be Tanzania and Kenya.     

Potential of kenaf (Hibiscus cannabinus L.) and corn (Zea mays L.) for phytoremediation of dredging sludge contaminated by trace metals

The potential of kenaf (Hibiscus cannabinus L.) and corn (Zea mays L.) for accumulation of cadmium and zinc was investigated. Plants have been grown in lysimetres containing dredging sludge, a substratum naturally rich in trace metals. Biomass production was determined. Sludge and water percolating from lysimeters were analyzed by atomic absorption spectrometry. No visible symptoms of toxicity were observed during the three- month culture. Kenaf and corn tolerate trace metals content in sludge. Results showed that Zn and Cd were found in corn and kenaf shoots at different levels, 2.49 mg/kg of Cd and 82.5 mg/kg of Zn in kenaf shoots and 2.1 mg/kg of Cd and 10.19 mg/kg in corn shoots. Quantities of extracted trace metals showed that decontamination of Zn and Cd polluted substrates is possible by corn and kenaf crops. Tolerance and bioaccumulation factors indicated that both species could be used in phytoremediation.     

Hibiscus chlorotic ringspot virus p27 and its isoforms affect symptom expression and potentiate virus movement in kenaf (Hibiscus cannabinus L.)

Hibiscus chlorotic ringspot virus (HCRSV), a member of the genus Carmovirus, encodes p27 (27-kDa protein) and two other in-frame isoforms (p25 and p22.5) that are coterminal at the carboxyl end. Only p27, which initiates at the 2570CUG codon, was detected in transfected kenaf (Hibiscus cannabinus L.) protoplasts through fusion to a Flag tag at either its N or C terminus. Subcellular localization of a p27-green fluorescent fusion protein in kenaf epidermal cells showed that it was localized to membrane structures close to cell walls. To study the functions of these proteins, a number of start codon mutants and premature translation termination mutants were constructed. Phenotypic differences were observed between the wild-type virus and these mutants during infection. Infectivity assays on plants indicated that p27 is a determinant of symptom severity. Without p25, appearance of symptoms on systemically infected kenaf leaves was delayed by 4 to 8 days. In a timecourse analysis, Western blot assays revealed that the delay corresponded to retardation in virus systemic movement, which suggested that p25 is probably involved in virus systemic movement. Mutations disrupting expression of p22.5 did not affect symptoms or virus movement.     

Surface topography of kenaf (Hibiscus cannabinus) sized papers

Bleached kenaf handsheets sized with different polymers such as chitosan, polyvinyl alcohol and cationic starch, were used for the determination of surface topography. A non-contact profilometer, the AltiSur 500, was used to characterize the topography of structural details in the paper surface. Numerical and visual characterization of surface roughness indicated that the surface of chitosan-sized paper was less rough than other surfaces, and all the sized papers were commonly smoother than the unsized paper. One property of bio-polymer of chitosan is its ability to form films that improve the surface properties of paper when it is applied to the surface of the sheet.     

Phenolic constituents from the core of kenaf (Hibiscus cannabinus)

Four lignans, boehmenan H [2-(4-hydroxy-3-methoxyphenyl)-5-[3-(4-hydroxy-3-methoxycinnamoyloxy)propyl]-3-hydroxymethyl-7-methoxybenzodihydrofuran], boehmenan K [2-(4-hydroxy-3-methoxyphenyl)-5-[3-(4-hydroxycinnamoyloxy)-1-propenyl]-3-(4-hydroxy-3-methoxycinnamoyloxymethyl)-7-methoxybenzodihydrofuran], threo-carolignan H [threo-1-(4-hydroxy-3-methoxyphenyl)-2-[4-[3-(4-hydroxy-3-methoxycinnamoyloxy)propyl]-2-methoxyphenoxy]-1,3-propanodiol], and threo-carolignan K [threo-1-(4-hydroxy-3-methoxyphenyl)-3-(4-hydroxy-3-methoxycinnamoyloxy)-2-[4-[3-(4-hydroxycinnamoyloxy)-1-propenyl]-2-methoxyphenoxy]-1-propanol] as well as several other lignans, aldehydes and a tyramine derivative were isolated from the acetone extract of core of kenaf (Hibiscus cannabinus). All the structures were established by spectroscopic methods. The hitherto unreported 13C NMR spectra of some compounds are also presented and discussed. 2D NMR techniques have allowed the revision of certain previously reported 13C NMR assignments of some scarce naturally occurring compounds.     

Comparative studies of genetic diversity in kenaf (Hibiscus cannabinus L.) varieties based on analysis of agronomic and RAPD data

Kenaf (Hibiscus cannabinus L.) is a fiber crop classified in the genus Hibiscus (Malvaceae), and has a great potential for its multipurpose utilization, in addition to its traditional usage. Varietal identification of kenaf is always problematic and knowledge on genetic diversity of kenaf varieties is also limited, which significantly hindered our effective utilization and conservation of the valuable kenaf germplasm. In order to find a proper method for identifying kenaf varieties and studying their variation, morpho-agronomic characters and random amplified polymorphic DNA (RAPD) markers were analyzed among 14 kenaf varieties commonly used in Japan. Data from morphological analysis showed that the included kenaf varieties could be divided into three major groups. The characters, such as middle stem diameter, whole stalk weight, and days to 50% flowering, are highly responsible for the variation of the kenaf varieties, but it is difficult to identify individual varieties merely by the morpho-agronomic characters. On the other hand, clearly separation of the kenaf varieties was achieved based on the RAPD variation patterns. Genetic relationship of the kenaf varieties can also be traced through the analysis of RAPD and morph-agronomic variation. It is concluded from the present study that RAPD analysis is an effective tool in identifying of kenaf varieties and determining their genetic relationships, particularly when combined with the analysis of morpho-agronomic characters.     

Assessment of molecular diversity and evolutionary relationship of kenaf (Hibiscus cannabinus L.), roselle (H. sabdariffa L.) and their wild relatives

Kenaf (Hibiscus cannabinus L.) and roselle (H. sabdariffa L.) are valuable fibre crop species with diverse end use. Phylogenetic relationship of 73 accessions of kenaf, roselle and their wild relatives from 15 countries was assessed using 44 inter-simple sequence repeat (ISSR) and jute (Corchorus olitorius L.) specific simple sequence repeats (SSR) markers. A total of 113 alleles were identified of which 61.95 % were polymorphic. Jute specific SSR markers exhibited high polymorphism and resolving power in kenaf, although ISSR markers exhibited higher resolving power than SSR markers. Number of polymorphic alleles varied from 1 to 5 for ISSR and 1 to 6 for SSR markers. Cultivated species exhibited higher allele polymorphism (57 %) than the wild species (35 %), but the improved cultivars exhibited lower genetic diversity compared to germplasm accessions. Accessions with common genetic lineage and geographical distribution clustered together. Indian kenaf varieties were distinct from cultivars bred in other countries and shared more genetic homology with African accessions. High genetic diversity was observed in the Indian (J = 0.35–0.74) and exotic kenaf germplasm collections (J = 0.38–0.79), suggesting kenaf might have been introduced in India from Africa through Central Asia during early domestication. Genetic similarity-based cluster analysis was in close accordance with taxonomic classification of Hibiscus.     

利用红麻复垦多金属污染酸化土壤

对大宝山矿区周边酸性多金属污染农田土壤进行了5年的红麻原位种植试验,研究不同改良剂对土壤改良的持效性及其对红麻生长的影响,以探索具重金属耐性的经济作物红麻用于重金属污染土壤大田复垦的可行性和有效性.结果表明:施加改良剂后,红麻能在重金属Pb、Zn、Cu、Cd和As含量分别为1600、440、640、7.6和850 mg·kg-1的土壤上定植.其中,白云石和粉煤灰的改良效果优于石灰石和有机肥.施加白云石或粉煤灰后,红麻地上部产量(干质量)为14～15 t·hm-2,达到一般农田产量水平,且红麻杆和红麻皮中重金属含量显著降低.红麻韧皮纤维(麻皮)含量达到32％ ～38％,韧皮纤维中可萃取重金属含量低于《生态纺织品技术要求》的标准,具有潜在的经济效益.白云石/粉煤灰改良红麻复垦的化学-植物联合修复模式是复垦酸性多金属污染农田土壤的有效措施.     

Comparative analysis of diversification and population structure of kenaf (Hibiscus cannabinus L.) and roselle (H. sabdariffa L.) using SSR and RGA (resistance gene analogue) markers

Multiple DNA marker systems and complementary analytical approaches are often useful in population genetic analysis and speciation of plants. We investigated population structure of kenaf (Hibiscus cannabinus) and roselle (H. sabdariffa) for gaining insight in evolution and geographic separation of these crop species using SSR and RGA (resistance gene analogues) markers through Bayesian clustering and principal coordinate analysis (PCoA) methods. Genotyping by 12 SSR and 16 RGA markers amplified a total of 172 loci in the study population. The RGA markers generated higher number of alleles per marker (8.2) as compared to SSR (3.4), but exhibited lower heterozygosity in the population. Genetic variance and heterozygosity in roselle population for both marker systems were lower than in kenaf. RGA markers revealed higher variation among populations. Bayesian structure as well as PCoA analysis using RGA marker revealed distinct cluster for roselle, while SSR-based classification revealed high admixture. Results indicate geographic isolation and natural selection for adaptive RGA alleles in kenaf. The Indian kenaf landraces were distinct from the exotic kenaf accessions, suggesting separate lineage formation by geographic separation. Possible origin and domestication of roselle in South India is proposed.     

Lignanamides and other phenolic constituents from the bark of kenaf (Hibiscus cannabinus).

Two new acyclic phenylpropane lignanamides, grossamide K and erythro-canabisine H, and the naphthol glucoside 2,5-dimethyl-3-O-beta-D-glucopyranosylnaphthol, along with six known compounds were isolated from the acetone extract of bark of Hibiscus cannabinus. All structures were established by spectroscopic methods including 2D NMR techniques, which allowed the correction of certain previously reported 13C NMR assignments of grossamide.     

First report on molecular detection of a stolbur phytoplasma (Group16SrXII-A) associated with kenaf (Hibiscus cannabinus L.) in India

Infection of stolbur phytoplasma was detected in kenaf (Hibiscus cannabinus) plants at CRIJAF research farm, Barrackpore, India. The infected plants formed profuse short branches at the top with bushy and bunchy top appearance. PCR with universal 16S rDNA phytoplasma primers P1/P7 yielded amplicons of 1.5 kb from all symptomatic leaf samples. Nested PCR with 16S-rDNA-specific nested primer pair R16F2n/R2 generated an amplicon of 1241 bp confirming the presence of a phytoplasma. The nested PCR products were sequenced and BALSTn analysis revealed 100% identity with 16S rRNA gene of phytoplasma. Phylogenetic analysis showed kenaf phytoplasma having 99% identity with both “Bois noir” stolbur phytoplasma 16SrXII group (Accession no: JQ181540). The RFLP data also supported the phylogenetic analysis. Multi-locus sequence characterisation assay was conducted by using different locus-specific primers viz. tuf, rpsC-rplV, rplF-rplR, map-SecY and uvrB-degV. The infected phytoplasma samples amplified only SecY gene and generated 1224 bp product which was deposited at NCBI (accession no: KC508636).     

Characterization and Cytopathology of Hibiscus Latent Ringspot Virus Isolated from Kenaf (Hibiscus cannabinus L.) in Italy

Hibiscus latent ringspot virus (HLRV) was isolated from kenaf plants (Hibiscus cannabinus L.) in Italy. and identified as III. RV by its host range, purification, electron microscopy and serology. Ultrastructural studies showed that infected plants contained membranous cytoplasmic inclusion bodies. characteristic of nepoviruses, and sometimes also crystalline aggregates.     

Protoplast isolation, culture, and fusion protocols for kenaf (Hibiscus cannabinus)

Protoplast isolation and culture protocols were developed for ten cultivars of Hibiscus cannabinus L. (kenaf). Leaves from seedling lines maintained in vitro were used as donor tissues. Optimal cell wall digestions were achieved with a combination of cellulysin (1.0%) and macerase (0.5%). Average yields ranged from 0.9×10⁵ to 5.9×10⁶ protoplasts g fw^-1 leaf tissue with viability estimates ranging from 53% to 87%. This protocol was ineffective for leaf tissue taken from plants grown in vivo. Protoplasts harvested from plantlets maintained in vitro produced rapidly growing calluses when plated in semi-solid medium after an initial culture in liquid medium. First cell divisions were observed within four to six days after initial culture in medium containing plant growth regulators 2,4-dichlorophenoxyacetic acid (1.4 M) and kinetin (13.8 M). An electrofusion protocol which did not significantly reduce protoplast viabilities was developed for kenaf protoplasts. The maximum fusion frequency (4.6%) was obtained with an electrofusion voltage of 2.0 kV cm^-1.Abbreviations 2,4-d 2,4-dichlorophenoxyacetic acid - BA 6-benzylaminopurine - FDA fluorescein diacetate - MS Murashige and Skoog - NAA 1-naphthaleneacetic acid - PGRs plant growth regulators - SCL seed clonal line     

X-ray irradiation of excised embryos of mesta (Hibiscus cannabinus L. and H. sabdariffa L.)

Summary Excised embryos of Hibiscus spp. were treated with 1 kR to 6 kR of X-ray. Results indicate that germination was unaffected at this level of employed doses in both species, which in turn implies that the factors responsible for inhibition of germination are not present in the embryo. LD₅₀ values differed between varieties and species. Early varieties of both species were more sensitive to radiation than late varieties. Strikingly similar effects were observed for the varieties with smaller embryos over those with larger ones. Allopolyploid H. sabdariffa (2n=72) was more susceptible than diploid H. cannabinus (2n=36).Differences in mutation frequency exist between species with different levels of ploidy and between varieties within the same species. Most of the HC mesta varieties yielded higher mutation frequencies than those of HS mesta. Optimal dose for triggering mutations in all varieties (except the chlorophyll mutation variety of HC mesta) of the two species lies within a narrow range of 1 kR to 2 kR. Cent per cent seedling abnormalities is concomitant to LD₅₀; nevertheless, optimum dose for mutation frequency is independent of LD₅₀. Hence, the response should be viewed in terms of respective genotype. The advantages of the embryo irradiation technique are mentioned.     

红麻品种(系)表型性状的因子和聚类分析

依据红麻高产育种目标,利用因子分析和聚类分析对40个红麻品种(系)的9个表型性状进行分析。结果表明:供试材料9个表型性状的变异系数为5.13%~25.33%,具有较大差异和丰富的遗传多样性;对供试红麻材料进行聚类分析,可分为3大类,其中,第Ⅲ类群属理想的高产品系。二维排序分析,评选出7个综合性状表现较好的亲本,T17-2、福红951、FH航992、H1301、ZHKX-01、福航优1号、福航优3号共7个品系,可在生产和育种中加以利用。