Optimisation of a highly efficient shoot regeneration system using leaf explants of Chinese jujube (<i>Ziziphus jujuba</i> Mill.) by response surface methodology

<p>Chinese jujube (<i>Ziziphus jujuba</i> Mill.) is a major fruit crop in Asia. In this study, response surface methodology (RSM) was successfully employed to establish a highly efficient <i>in vitro</i> propagation and regeneration system for the ‘Teapot’ jujube via shoot organogenesis. Among the tested factors, gibberellic acid (GA₃) concentration showed the most significant positive effect. The pre-culture darkness timing and medium were also important factors for highly efficient shoot regeneration of the ‘Teapot’ jujube. The highest regeneration (> 75%) was achieved by 1 week in darkness and culture on wood plant medium (WPM) containing 0.25 mg·L⁻¹ GA₃, 0.5 mg·L⁻¹ 6-benzylaminopurine (BAP) and 0.1 mg·L⁻¹ 3-indoleacetic acid (IAA). <i>In vitro</i>-derived shoots rooted very well in the modified ¹/₂ Murashige and Skoog (MS) medium containing 0.4 mg·L⁻¹ 3-indolebutyric acid (IBA), resulting in a 100% rooting rate. These findings suggest that the RSM can be employed to optimise the protocols needed for successful <i>in vitro</i> plant regeneration of jujube cultivars, with potential applications in plant genetic transformation practices, polyploidy induction and germplasm preservation.</p

Comparison of the differential proteins expression level in two groups.

<p>CK: the control, samples treated with cotton balls soaked distilled water only. T: the test, samples treated with cotton balls soaked 0.8% (8 mg/mL) colchicine solution.</p

Obs. pI and Mr, observed pI, Mr calculated from the 2D-gels with PDQuest 8.0.1 software according to standard marker proteins.

<p>ER: endoplasmic reticulum, PM: Plasma Membrane, M: Mitochondrion.</p

Histogram of GO classifications of assembled Chinese jujube (<i>Ziziphus jujuba M.</i>) unigenes.

<p>Results are summarized for three main GO categories: biological process, cellular component and molecular function.</p

Representative 2-DE gels of untreated (A) and colchicine treated (B) microsporangia.

<p>Twenty-seven of the spots showing at least a 2-fold change with <i>p</i><0.05 were analyzed by MALDI-TOF/TOF-MS.</p

Functional categorization of different proteins. Digitals stand for the protein number of each functional category.

<p>Functional categorization of different proteins. Digitals stand for the protein number of each functional category.</p

Summary of transcriptome sequencing of Chinese jujube fruit using 454 GS FLX Titanium.

<p>Summary of transcriptome sequencing of Chinese jujube fruit using 454 GS FLX Titanium.</p

<i>De Novo</i> Assembly and Characterization of the Fruit Transcriptome of Chinese Jujube (<i>Ziziphus jujuba</i> Mill.) Using 454 Pyrosequencing and the Development of Novel Tri-Nucleotide SSR Markers

<div><p>Chinese jujube (<i>Ziziphus jujuba</i> Mill.) is an economically important deciduous tree that has high therapeutic value and health benefits. However, a lack of sequence data and molecular markers have constrained genetic and breeding studies for better fruit quality and other traits in Chinese jujube. In this study, two combined cDNA libraries of ‘Dongzao’ fruit representing the early and late stages of fruit development were constructed and sequenced on the 454 GS FLX Titanium platform. In total, 1,124,197 reads were generated and then <i>de novo</i> assembled into 97,479 unigenes. A total of 52,938 unigenes were homologous to genes in the NCBI non-redundant sequence database. A total of 33,123 unigenes were assigned to one or more Gene Ontology terms, and 16,693 unigenes were classified into 319 Kyoto Encyclopedia of Genes and Genomes pathways. The results showed that the Smirnoff-Wheeler pathway was the main pathway for the biosynthesis of ascorbic acid in Chinese jujube. The number of differentially expressed genes between the two stages of fruit development was 1,764, among which 974 and 790 genes were up-regulated and down-regulated, respectively. Furthermore, 9,893 sequences were identified containing SSRs. 93 primer pairs designed from the sequences with a tri-nucleotide repeat showed successful PCR amplification and could be validated in Chinese jujube accessions and <i>Z. mauritiana Lam</i> and <i>Z. acidojujuba</i> as well, of which 71 primer pairs were polymorphic. The obtained transcriptome provides a most comprehensive resource currently available for gene discovery and the development of functional markers in <i>Z. jujuba</i>. The newly developed microsatellite markers could be used in applications such as genetic linkage analysis and association studies, diversity analysis, and marker-assisted selection in Chinese jujube and related species.</p></div

Frequency of classified novel developed SSR repeat types.

<p>Frequency of classified novel developed SSR repeat types.</p

Silver stained polyacrylamide gels of four example SSR loci showed microsatellite polymorphisms in Chinese jujube and related species.

<p>The SSR loci BFU2038, BFU2113, BFU2134 and BFU2143 are corresponding to A, B, C and D, respectively. Lane1, ‘Hupingzao’; Lane2, ‘Yuanling’; Lane3, ‘Dongzao’; Lane4, ‘Junzao’; Lane5, ‘Jidanzao’; Lane6, ‘Mayizao’; Lane7, ‘Jinsixiaozao’; Lane8, ‘Huizao’; Lane9, ‘Anonymous’; Lane10, ‘Wanglang’.</p