Research Advances of Black Spot on Marigold

Marigold （ Tagetes erecta） originated in South America is the main raw material of lutein, which plays a vital role in treating maeula lutea retinae, im- proving immunity and delaying aging. Black spot is the main disease reducing marigold production. This paper concluded occurrence and damage of black spot, the species and symptoms of pathogens, pathogenic factors, selection of disease-resistant varieties and effective means against black spot, and made a briefly analysis of the existing problems and the corresponding solutions, in order to provide a theoretical basis for disease-resistant breeding of marigold.

Molecular cloning,identification,and chromosomal localization of two MADS box genes in peach(Prunus persica)

MADS box proteins play an important role in floral development. To find genes involved in the floral transition of Prunus species, cDNAs for two MADS box genes, PpMADS1 and PpMADSIO, were cloned using degenerate primers and 5＇- and T-RACE based on the sequence database of P. persiea and P. duleis. The full length of PpMADS1 cDNA is 1,071 bp containing an open reading frame （ORF） of 717 bp and coding for a polypeptide of 238 amino acid residues. The full length of PpMADSIO cDNA is 937 bp containing an ORF of 633 bp and coding for a polypeptide of 210 amino acid residues. Sequence comparison revealed that PpMADS1 and PpMADSIO were highly homologous to genes API and PI in Arabidopsis, respectively. Phylogenetic analysis indicated that PpMADS1 belongs to the euAP1 clade of class A, and PpMADSIO is a member of GLO/PI clade of class B. RT-PCR analysis showed that PpMADS1 was expressed in sepal, petal, carpel, and fruit, which was slightly different from the expression pattern ofAPl; PpMADS10 was expressed in petal and stamen, which shared the same expression pattern as PI. Using selective mapping strategy, PpMADSI was assigned onto the Binl：50 on the G1 linkage group between the markers MCO44 and TSA2, and PpMADSIO onto the Bin1：73 on the same linkage group between the markers Lap- 1 and FGA8. Our results provided the basis for further dissection of the two MADS box gene function.

Jasmonic acid and ethylene signaling pathways participate in the defense response of Chinese cabbage to Pectobacterium carotovorum infection

Chinese cabbage(Brassica rapa subsp.pekinensis)suffers from soft rot disease caused by Pectobacterium carotovorum(Pc).To uncover the mechanisms underlying the defense response of Chinese cabbage to Pc,we constructed a suppression subtractive hybridization(SSH)library from Pc-infected cabbage and obtained 1919 non-redundant expressed sequence tags(ESTs),which were used for cDNA microarray.We detected 800 differentially expressed genes(DEGs)in cabbage at different time points post-Pc inoculation,which were further confirmed by quantitative real-time PCR.One quarter of these DEGs were involved in the biotic stress pathways visualized by MapMan.Among them,8,8,1,3,and 2 DEGs were related to jasmonic acid(JA),ethylene(ET),JA+ET,auxin,and abscisic acid(ABA)signaling pathways,respectively,while no DEG was detected for salicylic acid(SA)signaling.Assessment of phytohormone production in the Pc-infected leaves showed that JA and ET production was increased,while SA production was decreased.Treatment with JA,methyl jasmonate(MeJA),the ET precursor 1-aminocyclopropane-1-carboxylate(ACC),or combinations thereof,reduced the disease severity,and the JA and JA+ACC treatments were superior and performed equally well.Our findings suggest that JA and ET may act synergistically against Pc infection in Chinese cabbage,and JA-mediated signaling might be the most significant.

A maize bundle sheath defective mutation mapped on chromosome 1 between SSR markers umc1395 and umc1603

The bsd-pg（bundle sheath defective pale green） mutant is a novel maize mutation, controlled by a single recessive gene, which was isolated from offspring of maize plantlets regenerated from tissue callus of the maize inbred line 501. The characterization was that the biogenesis and development of the chloroplasts was mainly interfered in bundle sheath cells rather than in mesophyll cells. For mapping the bsd-pg, an F2 population was derived from a cross between the mutant bsd-pg and an inbred line Xianzao 17. Using specific locus amplified fragment sequencing（SLAF-Seq） technology, a total of 5 783 polymorphic SLAFs were analysed with 1 771 homozygous alleles between maternal and paternal parents. There were 49 SLAFs, which had a ratio of paternal to maternal alleles of 2：1 in bulked normal lines, and three trait-related candidate regions were obtained on chromosome 1 with a size of 3.945 Mb. For the fine mapping, new simple sequence repeats（SSRs） markers were designed by utilizing information of the B73 genome and the candidate regions were localized a size of 850 934 bp on chromosome 1 between umc1603 and umc1395, including 35 candidate genes. These results provide a foundation for the cloning of bsd-pg by map-based strategy, which is essential for revealing the functional differentiation and coordination of the two cell types, and helps to elucidate a comprehensive understanding of the C4 photosynthesis pathway and related processes in maize leaves.

Plant Regeneration from Immature Inflorescence Culture and Genetic Transformation of Wheatgrass(Agropyron cristatum×A. desertorum cv. Hycrest-Mengnong)

The plants of hybrid wheatgrass （A. cristatum×A. desertorum cv. Hycrest-Mengnong） were directly induced from embryogenic callus regenerated from immature inflorescence. Immature inflorescence was cultured on improved MS medium containing 2.0-3.0 mg L^-1 2,4-D to regenerate callus. The calli were then transferred to hormone-free MS medium for differentiation and 1/2 MS medium for rooting. Results showed that callus initiation frequency was 83.4% and plant regeneration frequency was 59.6%. Phosphinothricin acetyltransferase （bar） gene was transformed into the hybrid wheatgrass by particle bombardment. Resistant callus was obtained using selecting agent, herbicide glufosinate of 0.5 mg L^-1, and some transgenic plants were recovered in vitro. The transgenic plants were identified by PCR and Southern blot analysis and these plants developed normally in the glufosinate medium, whereas the nontransgenic plants did not. The results demonstrated that bar cDNA integrated into the genomic DNA of the transgenic plants. The transgenic frequencies of bar gene were 1.1%.

Condensation of STM is critical for shoot meristem maintenance and salt tolerance in Arabidopsis

The shoot meristem generates the entire shoot system and is precisely maintained throughout the life cycle under various environmental challenges.In this study,we identified a prion-like domain(PrD)in the key shoot meristem regulator SHOOT MERISTEMLESS(STM),which distinguishes STM from other related KNOX1 proteins.We demonstrated that PrD stimulates STM to form nuclear condensates,which are required for maintaining the shoot meristem.STM nuclear condensate formation is stabilized by selected PrD-containing STM-interacting BELL proteins in vitro and in vivo.Moreover,condensation of STM promotes its interaction with the Mediator complex subunit MED8 and thereby enhances its transcriptional activity.Thus,condensate formation emerges as a novel regulatory mechanism of shoot meristem functions.Furthermore,we found that the formation of STM condensates is enhanced upon salt stress,which allows enhanced salt tolerance and increased shoot branching.Our findings highlight that the transcription factor partitioning plays an important role in cell fate determination and might also act as a tunable environmental acclimation mechanism.

LettuceGDB:The community database for lettuce genetics and omics

As a globally popular leafy vegetable and a representative plant of the Asteraceae family,lettuce has great economic and academic significance.In the last decade,high-throughput sequencing,phenotyping,and other multi-omics data in lettuce have accumulated on a large scale,thus increasing the demand for an integrative lettuce database.Here,we report the establishment of a comprehensive lettuce database,LettuceGDB(https://www.lettucegdb.com/).As an omics data hub,the current LettuceGDB includes two reference genomes with detailed annotations;re-sequencing data from over 1000 lettuce varieties;a collection of more than 1300 worldwide germplasms and millions of accompanying phenotypic records obtained with manual and cutting-edge phenomics technologies;re-analyses of 256 RNA sequencing datasets;a complete miRNAome;extensive metabolite information for representative varieties and wild relatives;epigenetic data on the genome-wide chromatin accessibility landscape;and various lettuce research papers published in the last decade.Five hierarchically accessible functions(Genome,Genotype,Germplasm,Phenotype,and O-Omics)have been developed with a user-friendly interface to enable convenient data access.Eight built-in tools(Assembly Converter,Search Gene,BLAST,JBrowse,Primer Design,Gene Annotation,Tissue Expression,Literature,and Data)are available for data downloading and browsing,functional gene exploration,and experimental practice.A community forum is also available for information sharing,and a summary of current research progress on different aspects of lettuce is included.We believe that LettuceGDB can be a comprehensive functional database amenable to data mining and database-driven exploration,useful for both scientific research and lettuce breeding.

Genetic diversity of Chinese summer soybean germplasm revealed by SSR markers

There are abundant soybean germplasm in China. In order to assess genetic diversity of Chinese sum- mer soybean germplasm, 158 Chinese summer soybean ac- cessions from the primary core collection of G. max were used to analyze genetic variation at 67 SSR loci. A total of 460 alleles were detected, in which 414 and 419 alleles oc- curred in the 80 Huanghuai and the 78 Southern summer accessions, respectively. The average number of alleles per locus was 6.9 for all the summer accessions, and 6.2 for both Huanghuai and Southern summer accessions. Marker diver- sity (D) per locus ranged from 0.414 to 0.905 with an average of 0.735 for all the summer accessions, from 0.387 to 0.886 with an average of 0.708 for the Huanghuai summer acces- sions, and from 0.189 to 0.884 with an average of 0.687 for the Southern summer accessions. The Huanghuai and Southern summer germplasm were different in the specific alleles, allelic-frequencies and pairwise genetic similarities. UPGMA cluster analysis based on the similarity data clearly separated the Huanghuai from Southern summer soybean accessions, suggesting that they were different gene pools. The results indicate that Chinese Huanghuai and Southern summer soybean germplasm can be used to enlarge genetic basis for developing elite summer soybean cultivars by ex- changing their germplasm.

Optimization of butanol production from corn straw hydrolysate by Clostridium acetobutylicum using response surface method

Butanol is a new kind of very potential biofuels.Enzymatic hydrolysis of corn stalk was utilized in this study to produce butanol by Clostridium acetobutylicum CICC 8008.Plackett-Burman (P-B) design and Central Composite Design (CCD) were adopted to screen crucial factors during fermentation as well as the optimization of experimental conditions.The result demonstrated that among the seven factors,namely,Yeast extract,(NH 4) 2 SO 4,KH 2 PO 4,MgSO 4,FeSO 4,CuSO 4 and CaCO 3,only CaCO 3 was selected as the most critical factor.The optimization experiment results for CaCO 3 usage,temperature and reaction time by CCD were determined to be 5.04 g/L,35°C and 70 h,respectively.A corresponding mathematical model was established to predict the fermentation experiment and maximum butanol yield of 6.57 g/L was acquired.The result of verification experiment under the optimum conditions showed that 6.20 g/L was the maximum butanol yield.This demonstrated that statistical method was a powerful tool for the optimization of butanol production from enzymatic hydrolysis of corn stalk.

Hydrogen peroxide modulates abscisic acid signaling in root growth and development in Arabidopsis

Exogenous abscisic acid(ABA)can inhibit root growth and promote formation of more root hairs in the root tip of Arabidopsis.However,the molecular mechanisms that underlie root ABA signaling are largely unknown.We report here that hydrogen peroxide(H2O2)reduces the root growth of wild type, and the phenotype of H2O2 on the root growth is similar to ABA response.Meanwhile ABA-induced changes in the morphology of root system can be partly reversed by ascorbic acid in wild type and abolished in NADPH oxidase defective mutant atrbohF and atrbohC.Further,ABA can induce H2O2 accumulation in the root cells and enhance transcription level of OXI1,which is necessary for many more AOS-dependent processes such as root hair growth in Arabidopsis.Our results suggest that H2O2 as an important signal molecule is required for the ABA-regulated root growth and development in Arabidopsis.

Nucleotide Variation in the NCED3 Region of Arabidopsis thaliana and its Association Study with Abscisic Acid Content under Drought Stress

Drought tolerance is a comprehensive quantitative trait that is being understood further at the molecular genetic level. Abscisic acid （ABA） is the main drought-induced hormone that regulates the expression of many genes related to drought responses. 9-cis-epoxycarotenoid dioxygenase （NCED3） is thought to be a key enzyme in ABA biosynthesis. In this paper, we measured the ABA content increase under drought stress, and sequenced and compared the sequence of AtNCED3 among 22 Arabidopsis thaliana accessions. The results showed that the fold of ABA content increase under drought stress was highly variable among these accessions. High density single nucleotide polymorphism （SNP） and insertion/deletion （indel） were found in the AtNCED3 region, on average one SNP per 87.4 bp and one indel per 502 bp. Nucleotide diversity was significantly lower in the coding region than that in non-coding regions. The results of an association study with ANOVA analysis suggested that the 274th site （P←→S） and the 327th site （P←→R） amino acid variations might be the cause of ABA content increase of 163av accession under drought stress.

SSR Analysis of Genetic Relationship and Classification in Chrysanthemum Germplasm Collection

To further test the accuracy and rationality of the Chinese classification system for chrysanthemum(Chrysanthemum morifolium), 10 simple sequence repeat(SSR) markers were used to identify a collection of 88 chrysanthemum and its related genera accessions. In total, 42 effective alleles across 88 accessions were detected; 3 429 bands were obtained by PCR amplification, including 2 630 polymorphic bands. The similarity coefficient ranged from 0.53 to 0.88. Cluster analysis based on UPGMA illustrated that the wild species and large-flower cultivars were first divided into two clusters, then the large-flower cultivars formed five distinct groups according to petal type, indicated that petal type can be a classification criterion. In the wild species cluster, C. vestitum and C. zawadaskii grouped with A. trilobata, suggested that the Ajania genera was closely related to the Chrysanthemum genera. ‘Hangbaiju’, ‘Gongju’ and ‘Chuju’ were grouped together, and ‘Boju’, O. longilobus and C. mongolicum constituted another branch, showed a correlation with geographic region of origin. Population structure analysis was subsequently performed with K values ranging from 2 to 10, and the most likely estimate for the population structure is five subpopulations, which is nearly consistent with the clustering results. Principle component analysis was further performed to verify the classification results. The results showed that these SSR markers are very powerful for studying genetic relationships and will be useful tools in the identification and classification of chrysanthemum.

Pulping performance of transgenic poplar with depressed Caffeoyl-CoA O-methyltransferase

This paper evaluated pulping performance of 3-year-old field-grown transgenic poplar (Populus tremula × Populus alba). The transgenic poplar with anti-sense CCoAOMT had an about 13% decreased lignin content, in which a slight increment was found in S/G ratio. Chemical analysis showed that the transgenic poplar had significantly less benezene-ethanol extractive than that of control wood, but no significant differences were found in contents of ash, cold water extractive, hot water extractive, 1% NaOH extractive, holocellulose, pentosans and cellulose. Fiber assay demonstrated that down-regulation of CCoAOMT expression improved the fiber quality in transgenic poplar. Kraft pulping showed that lower lignin in transgenic poplar led to remarkable improved pulp quality and increased pulp yield.

Rearrangements of microtubule cytoskeleton in stomatal closure of Arabidopsis induced by nitric oxide

NO (nitric oxide), known as a key signal molecule in plant, plays important roles in regulation of stomatal movement. In this study, microtubule dynamics and its possible mechanism in the NO signal pathway were investigated. The results were as follows: (i) In vivo stomatal aperture assays revealed that both vinblastine (microtubule-disrupting drug) and SNP (exogenous NO donor) prevented stomatal opening in the light, and vinblastine even could enhance the inhibitory effect of SNP, whereas taxol (a microtubule-stabilizing agent) was able to reduce this effect; (ii) microtubules in the opening Arabi- dopsis guard cells expressing GFP:α-tubulin-6 (AtGFP:α-tubulin-6) were organized in parallel, straight and dense bundles, radiating from the ventral side to the dorsal side, and most of them were localized perpendicularly to the ventral wall; (iii) under the same environmental conditions, treated with SNP for 30 min, the radial arrays of microtubules in guard cells began to break down, twisted partially and be- came oblique or exhibited a random pattern; (iv) furthermore, the involvement of cytosolic Ca2+ in this event was tested. Stomatal aperture assays revealed that BAPTA-AM (a chelator of Ca2+) greatly sup- pressed the effect of NO on stomatal closure; however, it did not show the same function on stomatal closure induced by vinblastine. When BAPTA-AM was added to the SNP-pretreated solution, the SNP-induced disordered microtubulue cytoskeleton in guard cells underwent rearrangement in a time-dependent manner. After 30 min of treatment with BAPTA-AM, the cortical microtubules resumed the original radial distribution, almost the same as the control. All this indicates that NO may promote rearrangement of microtubule cytoskeleton via elevation of [Ca2+]cyt (free Ca2+ concentration in the cy- toplasm), finally leading to stomatal closure.

Functional characterization and mapping of two MADS box genes from peach (Prunus persica)

Previously an AGAMOUS gene homologue PpMADS4 and a FRUITFULL gene homologue PpMADS6 were isolated from peach (Prunus persica), and both genes were shown to express in the developing floral and fruits. To gain insight into their function, the two genes were constitutively expressed in Arabidopsis thaliana and their effects on plant growth and floral organ development were studied in this work. The transgenic plants all displayed early flowering and conversion of inflorescence to floral meristem. However, the two genes had different effects on the floral organ structures in A. thaliana. The transgenic plants overexpressing PpMADS4 displayed homeotic conversion of floral organs, and par- ticularly the perianth abscission was inhibited. The plants overexpressing PpMADS6 showed early flowering, produced higher number of carpels, petals, and stamens than nontransgenic plants, and pod shatter was prevented; significantly, the transgenic plants yielded more than one siliques from a single flower. A SSR molecular marker was developed for PpMADS4, and it was then assigned into the G5 linkage group of Prunus sp. Both PpMADS4 and PpMADS6 genes were located at the same region in the G5 linkage group. Our results showed the potential application of these two MADS box genes for crop and fruit tree improvement.

Pilot-scale experiments on multilevel contact oxidation treatment of poultry farm wastewater using saran lock carriers under different operation model

A pilot-scale multilevel contact oxidation reactors system, coupled with saran lock carriers,was applied for the treatment of poultry farm wastewater. The removal efficiencies of CODcr, ammonia, and the total nitrogen as well as the elimination performance of CODcr and total nitrogen along the three-level contact oxidation tanks under six designed operational models were investigated. Based on the performance of the nitrogen removal of the saran lock carriers and the distribution of anoxic–aerobic interspace under the suitable operation model, the mechanism of nitrogen removal of the system was also explored. The results revealed that the intermittent aeration under parallel model is the most suitable operation model, while the removal efficiencies of CODcr, ammonia, and the total nitrogen were 86.86%, 84.04%, and 80.96%, respectively. The effluent concentration of CODcr,ammonia, and the total nitrogen were 55.6 mg/L, 8.3 mg/L, and 12.0 mg/L, which satisfy both the discharge standard of pollutants for livestock and poultry breeding industry(GB18596–2001) and the first grade of the integrated wastewater discharge standard(GB 8978–1996). Moreover, the mechanism for the nitrogen removal should be attributed to the plenty of anoxic–aerobic interspaces of the biofilm and the three-dimensional spiral structure of the saran lock carriers, where the oxygen-deficient distribution was suitable for the happening of the simultaneous nitrification and denitrification process. Therefore, the multilevel contact oxidation tanks system is an effective pathway for the treatment of the poultry farm wastewater on the strength of a suitable operation model and novel carriers.

Virus-induced silencing of a tobacco deoxyhypusine synthase gene

A cDNA fragment corresponding to deoxyhy-pusine synthase gene NbDHS was isolated and cloned into potato virus X (PVX) vector for functional analysis in Nico-tiana benthamiana by using virus-induced gene silencing (VIGS). Plants agroinfected with recombinant virus vector PVX-NbDHS exhibited an increase in leaf biomass, delay in natural leaf senescence and flowering time, and decrease in leaf chlorophyll content. Semi-quantitative RT-PCR and Northern analysis showed that the transcript level of DHS was significantly lower in PVX-NbDHS infected plants. At the same time, the expression for eIF-5A, the target proteins of DHS in N. benthamiana, was concomitantly suppressed by semi-quantitative RT-PCR and Western analysis. From the phenotypic feature of the infected plants and the reduced expression abundance of DHS and eIF-5A, we concluded that NbDHS plays important roles in plant growth, development and senescence. The possible application of DHS gene in ge-netic modification of crops and horticultural plants was dis-cussed.

The size distribution of airborne bacteria and human pathogenic bacteria in a commercial composting plant

Composting plants are regarded as one of the important sources of environmental bioaerosols.However,limitations in the size distribution of airborne bacteria have prevented our comprehensive understanding of their risk to human health and their dispersal behavior.In this study,different sizes of airborne bacteria were collected using an eight-stage impactor from a full-scale composting facility.Size-related abundance and communities of airborne bacteria as well as human pathogenic bacteria(HPB)were investigated using 16S rRNA gene sequencing coupled with droplet digital PCR.Our results indicate that the bacterial concentrations from the eight stages were approximately 10^(4)-105copies/m^(3).Although no statistical correlation was detected between the particle size and the Shannon index,the influence of size on bacterial lineages was observed in both composting and packaging areas.For airborne bacteria from different stages,the dominant phyla were Firmicutes,Proteobacteria,and Actinobacteria,and the dominant genera was Bacillus.Seven out of eight HPB with a small geometric mean aerodynamic diameter had a high concentration in composting areas.Based on diameters of 2.42 to 5.09μm,most HPB in the composting areas were expected to be deposited on the bronchus and secondary bronchus.However,in the packaging areas,the deposition of HPB(diameters 3.70 to 8.96μm)occurred in the upper part of the respiratory tract.Our results on the size distribution,abundance,and diversity of these bacteria offer important information for the systematic evaluation of bacterial pathogenicity and the potential health impacts on workers in composting plants and the surrounding residents.

Elucidation of the MicroRNA Transcriptome in Western Corn Rootworm Reveals Its Dynamic and Evolutionary Complexity

Diabrotica virgifera virgifera(western corn rootworm,WCR)is one of the most destructive agricultural insect pests in North America.It is highly adaptive to environmental stimuli and crop protection technologies.However,little is known about the underlying genetic basis of WCR behavior and adaptation.More specifically,the involvement of small RNAs(s RNAs),especially micro RNAs(mi RNAs),a class of endogenous small non-coding RNAs that regulate various biological processes,has not been examined,and the datasets of putative s RNA sequences have not previously been generated for WCR.To achieve a comprehensive collection of s RNA transcriptomes in WCR,we constructed,sequenced,and analyzed s RNA libraries from different life stages of WCR and northern corn rootworm(NCR),and identified 101 conserved precursor mi RNAs(pre-mi RNAs)in WCR and other Arthropoda.We also identified 277 corn rootworm specific pre-mi RNAs.Systematic analyses of s RNA populations in WCR revealed that its s RNA transcriptome,which includes PIWI-interacting RNAs(pi RNAs)and mi RNAs,undergoes a dynamic change throughout insect development.Phylogenetic analysis of mi RNA datasets from model species reveals that a large pool of species-specific mi RNAs exists in corn rootworm;these are potentially evolutionarily transient.Comparisons of WCR mi RNA clusters to other insect species highlight conserved mi RNA-regulated processes that are common to insects.Parallel Analysis of RNA Ends(PARE)also uncovered potential mi RNA-guided cleavage sites in WCR.Overall,this study provides a new resource for studying the s RNA transcriptome and mi RNA-mediated gene regulation in WCR and other Coleopteran insects.