<i>VvAGAMOUS</i> Affect Development of Four Different Grape Species Ovary

Grape pistil has an important influence on fruit size and quality. However, there were few studies on grape ovary, and the development process of the ovary is still unclear. Therefore, in this paper, four different grape varieties with different lengths of small inflorescences, namely ‘Musct Hambourg’ grape (Vitis vinifera), ‘Concord’ grape (Vitis labrusca), ‘ShanPuTao’ grape (Vitis amurensis) and ‘GongNiang2Hao’ grape (Vitis amurensis × Vitis vinifera) were used as test materials. Four varieties ovary were significant differences by means of stereomicroscope, paraffin section. The expression of ovary determining gene VvAGAMOUS (VvAG) and its development related genes VvCRABS CLAW (VvCRC) andVvAGAMOUS-LIKE 11 (VvAGL11) with similar functions during the development of different grape varieties were preliminarily explored using fluorescence quantitative test. The relationship between VvAG and VvCRC, VvAG and VvAGL11 were analyzed using Y1H assay. Our results showed that there were obvious abdominal sutures on the surface of expect for ‘Musct Hambourg’ grape, and existing poly carpels. The ovary development of ‘ShanPuTao’ and ‘GongNiang2Hao’ grape was completed when the inflorescence length was less than 1 cm, while the ‘Concord’ and ‘Musct Hambourg’ grape were fully developed when the length of inflorescence was 3–4 and 4–5 cm, respectively. VvAG and VvCRC began to express in large quantities after the formation of stamen primordia, while VvAGL11 during the forming of ovule primordia. Therefore, VvAG and VvCRC mainly regulated the development of stamens and carpels and also promote the development of ovules, while VvAGL11 major regulated the development of ovules. The promoters of VvCRC and VvAGL11 were bound by VvAG. This study provides an important theoretical basis for further research on the molecular mechanism of grape ovary development.     

Differential Responses of <i>NHX1</i> and <i>SOS1</i> Gene Expressions to Salinity in two <i>Miscanthus sinensis</i> Anderss. Accessions with Different Salt Tolerance

The lignocellulosic crop Miscanthus spp. has been identified as a good candidate for biomass production. The responses of Miscanthus sinensis Anderss. to salinity were studied to satisfy the needs for high yields in marginal areas and to avoid competition with food production. The results indicated that the relative advantages of the tolerant accession over the sensitive one under saline conditions were associated with restricted Na⁺ accumulation in shoots. Seedlings of two accessions (salt-tolerant ‘JM0119’ and salt-sensitive ‘JM0099’) were subjected to 0 (control), 100, 200, and 300 mM NaCl stress to better understand the salt-induced biochemical responses of genes involved in Na⁺ accumulation in M. sinensis. The adaptation responses of genes encoding for Na⁺ /H⁺ antiporters, NHX1 and SOS1 to NaCl stress were examined in JM0119 and JM0099.The cDNA sequences of genes examined were highly conserved among the relatives of M. sinensis based on the sequencing on approximate 600 bp-long cDNA fragments obtained from degenerate PCR. These salt-induced variations of gene expression investigated by quantitative real-time PCR provided evidences for insights of the molecular mechanisms of salt tolerance in M. sinensis. The expression of NHX1 was up-regulated by salt stress in JM0119 shoot and root tissues. However, it was hardly affected in JM0099 shoot tissue except for a significant increase at the 100 mM salt treatment, and it was salt-suppressed in the JM0099 root tissue. In the root tissue, the expression of SOS1 was induced by the high salt treatment in JM0119 but repressed by all salt treatments in JM0099. Thus, the remarkably higher expression of NHX1 and SOS1 were associated with the resistance to Na⁺ toxicity by regulation of the Na⁺ influx, efflux, and sequestration under different salt conditions.     

Vegetative Compatibility and Virulence Diversity of <i>Verticillium dahliae</i> from Okra (<i>Abelmoschus esculentus</i>) Plantations in Turkey and Evaluation of Okra Landraces for Resistance to <i>V. dahliae</i>

Forty-four V. dahliae isolates were collected from symptomatic vascular tissues of okra plants each from a different field in eight provinces located in the eastern Mediterranean and western Anatolia regions of Turkey during 2006- 2009. Nitrate-nonutilizing (nit) mutants of V. dahliae from okra were used to determine heterokaryosis and genetic relatedness among isolates. All isolates from okra plants were grouped into two vegetative compatibility groups (VCGs) (1 and 2) and three subgroups as 1A (13.6%, 6/44), 2A (20.5%, 9/44) and 2B (65.9%, 29/44) according to international criteria. Pathogenicity tests were performed on a susceptible local okra (A. esculentus) landrace in greenhouse conditions. All isolates from VCG1A and VCG2B induced defoliation (D) and partial defoliation (PD) symptoms, respectively. Other isolates from VCG2A gave rise to typical leaf chlorosis symptoms without defoliation. The obtained data showed that the virulence level of V. dahliae isolates from okra was related to their VCG belongings. Eighteen okra landraces from diverse geographical origins were screened for resistance to VCG2B and VCG1A of V. dahliae. The results indicated that all landraces were more susceptible to highly virulent VCG1A-D pathotype displaying D or PD symptoms depending on their susceptibility levels with a mean disease severity index of 3.52 than to less virulent VCG2B-PD pathotype of V. dahliae displaying PD and ND symptoms with a mean disease severity index of 2.52. Significant differences were observed among the landraces; however, none of them exhibited a level of resistance. Okra landraces; Çorum, Hatay Has and Şanlıurfa displayed the lowest level of susceptibility or little tolerance to both D and PD pathotypes. VCG2B of PD was prevailing in the surveyed areas and VCG1A of D was the most virulent of the VCGs identified. Introduction of resistant genotypes to Turkish okra germplasm from different sources and breeding new resistant okra cultivars are critical for the sustainability of okra production.     

Potential structural and biochemical mechanisms of compositae wild species resistance to <Emphasis Type="Italic">Alternaria tenuissima</Emphasis>

Disease assessment to measure severity of alternaria leaf spot (Alternaria tenuissima (Fr.) Wiltsh.) was performed in 32 wild species of Compositae family by seedling inoculation. It was found that two species were resistant, four were moderately resistant, and others were susceptible to various degrees. Some leaf morphological traits of two resistant and two highly susceptible wild species were studied. Trichome density and height and wax content were found to be associated with plant passive resistance, while stomata density was not associated. After A. tenuissima inoculation, enzymes phenylalanine ammonia-lyase, β-1,3-glucanase, and chitinase were also assayed. Their higher activities in the selected resistant wild species and lower activities in the highly susceptible ones were found. This suggests the possibility that these enzymes are not only constituents of resistance but also could be used as biochemical markers for screening Compositae wild species for alternaria leaf spot resistance.     

Conventional Breeding and Molecular Markers for Blast Disease Resistance in Rice (<i>Oryza sativa</i> L.)

Monogenic lines, which carried 23 genes for blast resistance were tested and used donors to transfer resistance genes by crossing method. The results under blast nursery revealed that 9 genes from 23 genes were susceptible to highly susceptible under the three locations (Sakha, Gemmeza, and Zarzoura in Egypt); Pia, Pik, Pik-p, Piz-t, Pita, Pi b, Pi, Pi 19 and Pi 20. While, the genes Pii, Pik-s, Pik-h, Pi z, Piz-5, Pi sh, Pi 3, Pi 1, Pi 5, Pi 7, Pi 9, Pi 12, Pikm and Pita-2 were highly resistant at the same locations. Clustering analysis confirmed the results, which divided into two groups; the first one included all the susceptible genes, while the second one included the resistance genes. In the greenhouse test, the reaction pattern of five races produced 100% resistance under artificial inoculation with eight genes showing complete resistance to all isolates. The completely resistant genes: Pii, Pik-s, Piz, Piz-5 (=bi2) (t), Pita (=Pi4) (t), Pita, Pi b and Pi1 as well as clustering analysis confirmed the results. In the F₁ crosses, the results showed all the 25 crosses were resistant for leaf blast disease under field conditions. While, the results in F₂ population showed seven crosses with segregation ratio of 15 (R):1 (S), two cross gave segregated ratio of 3 R:1 S and one gave 13:3. For the identi- fication of blast resistance genes in the parental lines, the marker K3959, linked to Pik-s gene and the variety IRBLKS-F5 carry this gene, which was from the monogenic line. The results showed that four genotypes; Sakha 105, Sakha 103, Sakha 106 and IRBLKS-F5 were carrying Pik-s gene, while was absent in the Sakha 101, Sakha 104, IRBL5-M, IRBL9-W, IRBLTACP1 and IRBL9-W(R) genotypes. As for Pi 5 gene, the results showed that it was present in Sakha 103 and Sakha 104 varieties and absent in the rest of the genotypes. In addition, Pita-Pita- 2 gene was found in the three Egyptian genotypes (Sakha 105, Sakha 101 and Sakha 104) plus IRBLTACP1 monogenetic. In F2 generation, six populations were used to study the inheritance of blast resistance and specific primers to confirm the ratio and identify the resistance genes. However, the ratios in molecular markers were the same of the ratio under field evaluation in the most population studies. These findings would facilitate in breeding programs for gene pyramiding and gene accumulation to produce durable resistance for blast using those genotypes.     

Isozymes variability among <Emphasis Type="Italic">Fusarium udum</Emphasis> resistant cultivars of pigeonpea (<Emphasis Type="Italic">Cajanus cajan</Emphasis> (L.) (Millsp)

The present study was carried out to select the different pigeonpea cultivars for resistance against wilt caused by Fusarium udum and to assess the genetic variability among the resistant and susceptible cultivars. These cultivars were screened by root dip inoculation and classified into resistant (ICP 8863 and 9145), moderately resistant (ICP 11681 and Selection-1), susceptible (ICP 7118, TRG-1 and LRG-30) and highly susceptible cultivars (ICP-2376 and LRG-41). The peroxidase activity (PEO) in both leaf and root tissues of four pigeonpea cultivars (ICP 8863, Selection-1, ICP 2376 and LRG-30) were determined at 1^st, 4^th and 7^th day after inoculation (DAI) in healthy and F. udum infected tissues. Higher PEO activity in both leaf and root was observed and at 4^th DAI in susceptible cultivars. In native-PAGE analysis of isozymes, the induction of specific leaf peroxidase band (Em=0.17) and two root peroxidase bands (Em=0.24 and 0.55) were observed in ICP 8863 after inoculation. Significant differences were observed in the leaf phosphatase and esterase banding profiles of all the cultivars. The presence of leaf phosphatase band at Em of 0.04 was observed only in ICP 8863 and 11681. The leaf esterase band (Em=0.3) was well expressed in ICP 8863 when compared to other cultivars. The significance of peroxidase in plant defense mechanism and utility of biochemical markers in breeding programmes are discussed. Part of M.Sc. (Ag) thesis of the first author and approved by the Acharya N.G. Ranga Agricultural University during March 2002.     

Genome-Wide Analysis of the <i>KANADI</i> Gene Family and Its Expression Patterns under Different Nitrogen Concentrations Treatments in <i>Populus trichocarpa</i>

KANADI (KAN) is a plant-specific gene that controlled the polarity development of lateral organs. It mainly acted on the abaxial characteristics of plants to make the lateral organs asymmetrical. However, it had been less identified in woody plants. In this study, the members of the KAN gene family in Populus trichocarpa were identified and analyzed using the bioinformatics method. The results showed that a total of 8 KAN family members were screened out, and each member contained the unique GARP domain and conserved region of the family proteins. Phylogenetic analysis and their gene structures revealed that all KAN genes from P. trichocarpa, Arabidopsis thaliana, and Nicotiana benthamiana could be divided into four subgroups, while the eight genes in P. trichocarpa were classified into three subgroups, respectively. The analysis of tissue-specific expression indicated that PtKAN1 was highly expressed in young leaves, PtKAN6 was highly expressed in young leaves and mature leaves, PtKAN2, PtKAN5, and PtKAN7 were highly expressed in nodes and internodes, PtKAN8 was highly expressed in roots, and PtKAN3 and PtKAN4 showed low expression levels in all tissues. Among them, PtKAN2 and PtKAN6, and PtKAN4 and PtKAN5 might have functional redundancy. Under high nitrogen concentrations, PtKAN2 and PtKAN8 were highly expressed in mature stems and leaves, respectively, while PtKAN4, PtKAN5, and PtKAN7 were highly expressed in roots. This study laid a theoretical foundation for further study of the KAN gene-mediated nitrogen effect on root development.     

Molecular Basis of Unique Branching Phenotypes in <i>Salvia splendens</i> and the Role of PSY

The branching system of higher plants plays a very important role in plant morphogenesis, and the number of branches can directly affect crop yield and the ornamental value of plants. It is a complicated development process involving complex molecular mechanisms. The ‘Cailinghong’ variety of Salvia splendens is characterized by its great branching ability with the ability to grow into a spherical form naturally, without pinching. To gain insight into the molecular events during the branching development of S. splendens, suppressive subtractive hybridization (SSH) technology was used to screen differentially expressed genes between the erect plant type (strain 35) and the spherical plant type (‘Cailinghong’). In total, 96 and 116 unigenes were annotated. Four and eight unigenes up-regulated in ‘Cailinghong’ and strain 35, respectively, were associated with plant hormone anabolism and signal transduction, suggesting that they participate in the branching process. One of these genes, phytoene synthase (PSY), is a precursor of the new plant hormone group strigolactones. Using the PSY fragment (192 bp) as a template, the cDNA sequence of PSY in S. splendens was cloned and named SsPSY. A relative expression analysis and transgenic test results indicated that SsPSY plays an important role in lateral branch development in ‘Cailinghong’. These results provide new insight into the molecular mechanisms underlying branching in S. splendens.     

Identification and Evaluation of Insect and Disease Resistance in Transgenic <i>Cry1Ab13-1</i> and <i>NPR1</i> Maize

PCR detection, quantitative real-time PCR (q-RTPCR), outdoor insect resistance, and disease resistance identification were carried out for the detection of genetic stability and disease resistance through generations (T₂, T₃, and T₄) in transgenic maize germplasms (S3002 and 349) containing the bivalent genes (insect resistance gene Cry1Ab13-1 and disease resistance gene NPR1) and their corresponding wild type. Results indicated that the target genes Cry1Ab13-1 and NPR1 were successfully transferred into both germplasms through tested generations; q-PCR confirmed the expression of Cry1Ab13-1 and NPR1 genes in roots, stems, and leaves of tested maize plants. In addition, S3002 and 349 bivalent gene-transformed lines exhibited resistance to large leaf spots and corn borer in the field evaluation compared to the wild type. Our study confirmed that Cry1Ab13-1 and NPR1 bivalent genes enhanced the resistance against maize borer and large leaf spot disease and can stably inherit. These findings could be exploited for improving other cultivated maize varieties.     

Overexpression of <Emphasis Type="Italic">AP1-</Emphasis>like genes from <Emphasis Type="Italic">Asteraceae</Emphasis> induces early-flowering in transgenic <Emphasis Type="Italic">Chrysanthemum</Emphasis> plants

Chrysanthemum is one of the most important commercial cut flowers in the world. Early-flowering cultivars are required to produce quality chrysanthemum flowers with a lower cost of production. To shorten the vegetative growth phase of chrysanthemum, three AP1-like genes from Asteraceae were constitutively overexpressed in 80 independent transgenic chrysanthemum lines. All lines were characterized by PCR and RT-PCR and demonstrated that overexpression of compositae AP1-homologs in transgenic chrysanthemum under long-day conditions had no effect on plant development compared to non-transgenic controls. Conversely, under short-day conditions, transgenic plants commenced bud initiation 2 wk earlier than non-transgenic chrysanthemum plants. Subsequently, transgenic chrysanthemum flowers showed color earlier and resulted in full opening of inflorescences 3 wk prior to non-transgenic control plants. These results open new possibilities for genetic improvement and breeding of chrysanthemum cultivars.     

Resistance to Tomato leaf curl New Delhi virus in Cucurbita spp.

Tomato leaf curl New Delhi virus (ToLCNDV) is a bipartite begomovirus (family Geminiviridae) first reported in India and its neighbouring countries. ToLCNDV severely affects zucchini crop (Cucurbita pepo) in the main production areas of Southern Spain since 2012. This emerging begomovirus is a serious threat to this and other cucurbit crops. Breeding resistant cultivars is the most promising method for disease control, but requires the identification of sources of resistance in the Cucurbita genus. In this work, we screened for ToLCNDV resistance a large collection of Cucurbita spp. accessions, including landraces and commercial cultivars of the main cultivated species, C. pepo, Cucurbita moschata and Cucurbita maxima and wild species. The screening was performed using mechanical and whitefly inoculation. The level of resistance was assessed by scoring symptom severity and by measuring the virus content with quantitative polymerase chain reaction in selected genotypes. Diversity in the response was observed within and among species. Severe symptoms and high viral amounts were found at 30 days after mechanical and whitefly inoculation in C. pepo, in all accessions belonging to the Zucchini morphotype and to other morphotypes of both subspecies, pepo and ovifera, and even in the wild relative Cucurbita fraterna. C. maxima was also highly susceptible. This species showed characteristic symptoms of leaf decay and intense yellowing, different from those of mosaic, curling and internode shortening found in C. pepo. The only species showing resistance was C. moschata. Four accessions were symptomless or had some plants with only mild symptoms after three independent rounds of mechanical inoculation with different inoculum sources. Two of them also remained symptomless after virus inoculation with viruliferous whiteflies. ToLCNDV was detected in these asymptomatic accessions at 15 and 30 days post inoculation, but viral amounts were much lower than those found in susceptible genotypes, suggesting a high level of resistance. The symptoms in the susceptible accessions of this species were also different, with a characteristic leaf mottling, evolving to a severe mosaic. The newly identified C. moschata resistant accessions are good candidates for breeding programmes to avoid the damage caused by ToLCNDV.     

Comparative Analyses and Phylogenetic Relationships between <i>Cryptomeria fortunei</i> and Related Species Based on Complete Chloroplast Genomes

Cryptomeria fortunei (Chinese cedar) is a highly adaptable woody species and one of the main forest plantation trees in subtropical high-altitude areas in China. However, there are few studies on its chloroplast (cp) genome. In this study, the complete cp genome of C. fortunei was sequenced and evaluated via comparative analyses with those of related species (formerly the Taxodiaceae) in Cupressaceae. The C. fortunei cp genome was 131,580 bp in length, and the GC content of the whole genome was 35.38%. It lost one relevant large inverted repeat and contained 114 unique genes, including 82 protein-coding genes, 28 tRNAs and 4 rRNAs. The relative synonymous codon usage (RSCU) of codons ending with A/U was more than twice that of codons ending with G/C. Thirty long repeat structures (LRSs) and 213 simple sequence repeat (SSR) loci were detected in the C. fortunei cp genome. Comparative analyses of 10 cp genomes revealed that substantial rearrangements occurred in the gene organization. Additionally, 6 cp hotspot regions (trnS-GGA, ycf1, trnP-GGG, trnC-GCA, psbZ and accD) were identified, and 4 genes (petL, psbM, rpl22 and psaM) had likely underwent positive selection. Phylogenetic analysis showed that Cupressaceae, Taxaceae and Cephalotaxaceae clustered to form a clade and that C. fortunei was most closely related to C. japonica (Japanese cedar), C. japonica cv. Wogon Hort and Taxodium distichum (baldcypress). These results provide references for future studies of population genetics, phylogenetic status and molecular markers among Cupressaceae species and for the cultivation of improved varieties.