Rice OsGATA16 is a positive regulator for chlorophyll biosynthesis and chloroplast development.

Chloroplasts are essential organelles in plants that contain chlorophylls and facilitate photosynthesis for growth and development. As photosynthetic efficiency significantly impacts crop productivity, understanding the regulatory mechanisms of chloroplast development has been crucial in increasing grain and biomass production. This study demonstrates the involvement of OsGATA16, an ortholog of Arabidopsis GATA, NITRATE INDUCIBLE, CARBON-METABOLISM INVOLVED (GNC), and GNC-LIKE/CYTOKININ-RESPONSIVE GATA FACTOR 1 (GNL/CGA1), in chlorophyll biosynthesis and chloroplast development in rice (Oryza sativa). The osgata16-1 knockdown mutants produced pale-green leaves, while OsGATA16-overexpressed plants (OsGATA16-OE1) generated dark-green leaves, compared to their parental japonica rice. Reverse transcription and quantitative PCR analysis revealed downregulation of genes related to chloroplast division, chlorophyll biosynthesis, and photosynthesis in the leaves of osgata16-1 and upregulation in those of OsGATA16-OE1. Additionally, in vivo binding assays showed that OsGATA16 directly binds to the promoter regions of OsHEMA, OsCHLH, OsPORA, OsPORB, and OsFtsZ, and upregulates their expression. These findings indicate that OsGATA16 serves as a positive regulator controlling chlorophyll biosynthesis and chloroplast development in rice.

Genes and Regulatory Mechanisms for Ginsenoside Biosynthesis.

Panax ginseng is a medicinal plant belonging to the Araliaceae family. Ginseng is known as the king of oriental medicine, which has been practiced since ancient times in East Asian countries and globally in the modern era. Ginseng is used as an adaptogen, and research shows that it has several pharmacological benefits for various ailments such as cancer, inflammation, diabetes, and neurological symptoms. The pharmacological benefits of ginseng are attributed to the triterpenoid saponin ginsenosides found throughout the Panax ginseng species, which are abundant in its root and are found exclusively in P. ginseng and Panax quinquefolius. Recently, with the completion of the entire ginseng genome sequencing and the construction of the ginseng genome database, it has become possible to access information about many genes newly predicted to be involved in ginsenoside biosynthesis. This review briefly summarizes the current progress in ginseng genome analysis and genes involved in ginsenoside biosynthesis, proposing directions for functional studies of the predicted genes related to ginsenoside production and its regulation.

Genome structure and diversity among Cynanchum wilfordii accessions.

BACKGROUND: Cynanchum wilfordii (Cw) and Cynanchum auriculatum (Ca) have long been used in traditional medicine and as functional food in Korea and China, respectively. They have diverse medicinal functions, and many studies have been conducted, including pharmaceutical efficiency and metabolites. Especially, Cw is regarded as the most famous medicinal herb in Korea due to its menopausal symptoms relieving effect. Despite the high demand for Cw in the market, both species are cultivated using wild resources with rare genomic information. RESULTS: We collected 160 Cw germplasm from local areas of Korea and analyzed their morphological diversity. Five Cw and one Ca of them, which were morphologically diverse, were sequenced, and nuclear ribosomal DNA (nrDNA) and complete plastid genome (plastome) sequences were assembled and annotated. We investigated the genomic characteristics of Cw as well as the genetic diversity of plastomes and nrDNA of Cw and Ca. The Cw haploid nuclear genome was approximately 178 Mbp. Karyotyping revealed the juxtaposition of 45S and 5S nrDNA on one of 11 chromosomes. Plastome sequences revealed 1226 interspecies polymorphisms and 11 Cw intraspecies polymorphisms. The 160 Cw accessions were grouped into 21 haplotypes based on seven plastome markers and into 108 haplotypes based on seven nuclear markers. Nuclear genotypes did not coincide with plastome haplotypes that reflect the frequent natural outcrossing events. CONCLUSIONS: Cw germplasm had a huge morphological diversity, and their wide range of genetic diversity was revealed through the investigation with 14 molecular markers. The morphological and genomic diversity, chromosome structure, and genome size provide fundamental genomic information for breeding of undomesticated Cw plants.

Admixture of divergent genomes facilitates hybridization across species in the family Brassicaceae.

Hybridization and polyploidization are pivotal to plant evolution. Genetic crosses between distantly related species are rare in nature due to reproductive barriers but how such hurdles can be overcome is largely unknown. Here we report the hybrid genome structure of xBrassicoraphanus, a synthetic allotetraploid of Brassica rapa and Raphanus sativus. We performed cytogenetic analysis and de novo genome assembly to examine chromosome behaviors and genome integrity in the hybrid. Transcriptome analysis was conducted to investigate expression of duplicated genes in conjunction with epigenome analysis to address whether genome admixture entails epigenetic reconfiguration. Allotetraploid xBrassicoraphanus retains both parental chromosomes without genome rearrangement. Meiotic synapsis formation and chromosome exchange are avoided between nonhomologous progenitor chromosomes. Reconfiguration of transcription network occurs, and less divergent cis-elements of duplicated genes are associated with convergent expression. Genome-wide DNA methylation asymmetry between progenitors is largely maintained but, notably, B. rapa-originated transposable elements are transcriptionally silenced in xBrassicoraphanus through gain of DNA methylation. Our results demonstrate that hybrid genome stabilization and transcription compatibility necessitate epigenome landscape adjustment and rewiring of cis-trans interactions. Overall, this study suggests that a certain extent of genome divergence facilitates hybridization across species, which may explain the great diversification and expansion of angiosperms during evolution.

Prolonged Exposure to High Temperature Inhibits Shoot Primary and Root Secondary Growth in Panax ginseng.

High temperature is one of the most significant abiotic stresses reducing crop yield and quality by inhibiting plant growth and development. Global warming has recently increased the frequency of heat waves, which negatively impacts agricultural fields. Despite numerous studies on heat stress responses and signal transduction in model plant species, the molecular mechanism underlying thermomorphogenesis in Panax ginseng remains largely unknown. Here, we investigated the high temperature response of ginseng at the phenotypic and molecular levels. Both the primary shoot growth and secondary root growth of ginseng plants were significantly reduced at high temperature. Histological analysis revealed that these decreases in shoot and root growth were caused by decreases in cell elongation and cambium stem cell activity, respectively. Analysis of P. ginseng RNA-seq data revealed that heat-stress-repressed stem and root growth is closely related to changes in photosynthesis, cell wall organization, cell wall loosening, and abscisic acid (ABA) and jasmonic acid (JA) signaling. Reduction in both the light and dark reactions of photosynthesis resulted in defects in starch granule development in the storage parenchymal cells of the main tap root. Thus, by combining bioinformatics and histological analyses, we show that high temperature signaling pathways are integrated with crucial biological processes that repress stem and root growth in ginseng, providing novel insight into the heat stress response mechanism of P. ginseng.

Plastid Genomes of the Early Vascular Plant Genus Selaginella Have Unusual Direct Repeat Structures and Drastically Reduced Gene Numbers.

The early vascular plants in the genus Selaginella, which is the sole genus of the Selaginellaceae family, have an important place in evolutionary history, along with ferns, as such plants are valuable resources for deciphering plant evolution. In this study, we sequenced and assembled the plastid genome (plastome) sequences of two Selaginella tamariscina individuals, as well as Selaginella stauntoniana and Selaginella involvens. Unlike the inverted repeat (IR) structures typically found in plant plastomes, Selaginella species had direct repeat (DR) structures, which were confirmed by Oxford Nanopore long-read sequence assembly. Comparative analyses of 19 lycophytes, including two Huperzia and one Isoetes species, revealed unique phylogenetic relationships between Selaginella species and related lycophytes, reflected by structural rearrangements involving two rounds of large inversions that resulted in dynamic changes between IR and DR blocks in the plastome sequence. Furthermore, we present other uncommon characteristics, including a small genome size, drastic reductions in gene and intron numbers, a high GC content, and extensive RNA editing. Although the 16 Selaginella species examined may not fully represent the genus, our findings suggest that Selaginella plastomes have undergone unique evolutionary events yielding genomic features unparalleled in other lycophytes, ferns, or seed plants.

Identification of a gene responsible for cytoplasmic male-sterility in onions (Allium cepa L.) using comparative analysis of mitochondrial genome sequences of two recently diverged cytoplasms.

KEY MESSAGE: Almost identical mitochondrial genome sequences of two recently diverged male-fertile normal and male-sterile CMS-T-like cytoplasms were obtained in onions. A chimeric gene, orf725 , was found to be a CMS-inducing gene. In onions (Allium cepa L.), cytoplasmic male-sterility (CMS) has been widely used in hybrid seed production. Two types of CMS (CMS-S and CMS-T) have been reported in onions. A complete mitochondrial genome sequence of the CMS-S cytoplasm has been reported in our previous study. Draft mitochondrial genome sequences of male-fertile normal and CMS-T-like cytoplasms are reported in this study. Raw reads obtained from normal and CMS-T-like cytoplasms were assembled into eight and nine almost identical contigs, respectively. After connection and reorganization of contigs by PCR amplification and genome walking, four scaffold sequences with total length of 339 and 180 bp were produced for the normal cytoplasm. A mitochondrial genome sequence of the CMS-T-like cytoplasm was obtained by mapping trimmed reads of CMS-T onto scaffold sequences of the normal cytoplasm. Compared with the CMS-S mitochondrial genome, the normal mitochondrial genome was highly rearranged with 31 syntenic blocks. A total of 499 single nucleotide polymorphisms (SNPs) or insertions/deletions were identified in these syntenic regions. On the other hand, normal and CMS-T-like mitochondrial genome sequences were almost identical except for orf725, a chimeric gene consisting of cox1 with other sequences. Only three SNPs were identified between normal and CMS-T-like syntenic sequences. These results indicate that orf725 is likely to be the casual gene for CMS induction in onions and that CMS-T-like cytoplasm has recently diverged from the normal cytoplasm by introduction of orf725.

Dynamic Chloroplast Genome Rearrangement and DNA Barcoding for Three Apiaceae Species Known as the Medicinal Herb "Bang-Poong".

Three Apiaceae species Ledebouriella seseloides, Peucedanum japonicum, and Glehnia littoralis are used as Asian herbal medicines, with the confusingly similar common name "Bang-poong". We characterized the complete chloroplast (cp) genomes and 45S nuclear ribosomal DNA (45S nrDNA) sequences of two accessions for each species. The complete cp genomes of G. littoralis, L. seseloides, and P. japonicum were 147,467, 147,830, and 164,633 bp, respectively. Compared to the other species, the P. japonicum cp genome had a huge inverted repeat expansion and a segmental inversion. The 45S nrDNA cistron sequences of the three species were almost identical in size and structure. Despite the structural variation in the P. japonicum cp genome, phylogenetic analysis revealed that G. littoralis diverged 5-6 million years ago (Mya), while P. japonicum diverged from L. seseloides only 2-3 Mya. Abundant copy number variations including tandem repeats, insertion/deletions, and single nucleotide polymorphisms, were found at the interspecies level. Intraspecies-level polymorphism was also found for L. seseloides and G. littoralis. We developed nine PCR barcode markers to authenticate all three species. This study characterizes the genomic differences between L. seseloides, P. japonicum, and G. littoralis; provides a method of species identification; and sheds light on the evolutionary history of these three species.

Ginseng Genome Database: an open-access platform for genomics of Panax ginseng.

BACKGROUND: The ginseng (Panax ginseng C.A. Meyer) is a perennial herbaceous plant that has been used in traditional oriental medicine for thousands of years. Ginsenosides, which have significant pharmacological effects on human health, are the foremost bioactive constituents in this plant. Having realized the importance of this plant to humans, an integrated omics resource becomes indispensable to facilitate genomic research, molecular breeding and pharmacological study of this herb. DESCRIPTION: The first draft genome sequences of P. ginseng cultivar "Chunpoong" were reported recently. Here, using the draft genome, transcriptome, and functional annotation datasets of P. ginseng, we have constructed the Ginseng Genome Database http://ginsengdb.snu.ac.kr /, the first open-access platform to provide comprehensive genomic resources of P. ginseng. The current version of this database provides the most up-to-date draft genome sequence (of approximately 3000 Mbp of scaffold sequences) along with the structural and functional annotations for 59,352 genes and digital expression of genes based on transcriptome data from different tissues, growth stages and treatments. In addition, tools for visualization and the genomic data from various analyses are provided. All data in the database were manually curated and integrated within a user-friendly query page. CONCLUSION: This database provides valuable resources for a range of research fields related to P. ginseng and other species belonging to the Apiales order as well as for plant research communities in general. Ginseng genome database can be accessed at http://ginsengdb.snu.ac.kr /.

Genome and evolution of the shade-requiring medicinal herb Panax ginseng.

Panax ginseng C. A. Meyer, reputed as the king of medicinal herbs, has slow growth, long generation time, low seed production and complicated genome structure that hamper its study. Here, we unveil the genomic architecture of tetraploid P. ginseng by de novo genome assembly, representing 2.98 Gbp with 59 352 annotated genes. Resequencing data indicated that diploid Panax species diverged in association with global warming in Southern Asia, and two North American species evolved via two intercontinental migrations. Two whole genome duplications (WGD) occurred in the family Araliaceae (including Panax) after divergence with the Apiaceae, the more recent one contributing to the ability of P. ginseng to overwinter, enabling it to spread broadly through the Northern Hemisphere. Functional and evolutionary analyses suggest that production of pharmacologically important dammarane-type ginsenosides originated in Panax and are produced largely in shoot tissues and transported to roots; that newly evolved P. ginseng fatty acid desaturases increase freezing tolerance; and that unprecedented retention of chlorophyll a/b binding protein genes enables efficient photosynthesis under low light. A genome-scale metabolic network provides a holistic view of Panax ginsenoside biosynthesis. This study provides valuable resources for improving medicinal values of ginseng either through genomics-assisted breeding or metabolic engineering.

Completion of the mitochondrial genome sequence of onion (Allium cepa L.) containing the CMS-S male-sterile cytoplasm and identification of an independent event of the ccmF N gene split.

Cytoplasmic male-sterility (CMS) conferred by the CMS-S cytoplasm has been most commonly used for onion (Allium cepa L.) F1 hybrid seed production. We first report the complete mitochondrial genome sequence containing CMS-S cytoplasm in this study. Initially, seven contigs were de novo assembled from 150-bp paired-end raw reads produced from the total genomic DNA using the Illumina NextSeq500 platform. These contigs were connected into a single circular genome consisting of 316,363 bp (GenBank accession: KU318712) by PCR amplification. Although all 24 core protein-coding genes were present, no ribosomal protein-coding genes, except rps12, were identified in the onion mitochondrial genome. Unusual trans-splicing of the cox2 gene was verified, and the cox1 gene was identified as part of the chimeric orf725 gene, which is a candidate gene responsible for inducing CMS. In addition to orf725, two small chimeric genes were identified, but no transcripts were detected for these two open reading frames. Thirteen chloroplast-derived sequences, with sizes of 126-13,986 bp, were identified in the intergenic regions. Almost 10 % of the onion mitochondrial genome was composed of repeat sequences. The vast majority of repeats were short repeats of <100 base pairs. Interestingly, the gene encoding ccmFN was split into two genes. The ccmF N gene split is first identified outside the Brassicaceae family. The breakpoint in the onion ccmF N gene was different from that of other Brassicaceae species. This split of the ccmF N gene was also present in 30 other Allium species. The complete onion mitochondrial genome sequence reported in this study would be fundamental information for elucidation of onion CMS evolution.

Major repeat components covering one-third of the ginseng (Panax ginseng C.A. Meyer) genome and evidence for allotetraploidy.

Ginseng (Panax ginseng) is a famous medicinal herb, but the composition and structure of its genome are largely unknown. Here we characterized the major repeat components and inspected their distribution in the ginseng genome. By analyzing three repeat-rich bacterial artificial chromosome (BAC) sequences from ginseng, we identified complex insertion patterns of 34 long terminal repeat retrotransposons (LTR-RTs) and 11 LTR-RT derivatives accounting for more than 80% of the BAC sequences. The LTR-RTs were classified into three Ty3/gypsy (PgDel, PgTat and PgAthila) and two Ty1/Copia (PgTork and PgOryco) families. Mapping of 30-Gbp Illumina whole-genome shotgun reads to the BAC sequences revealed that these five LTR-RT families occupy at least 34% of the ginseng genome. The Ty3/Gypsy families were predominant, comprising 74 and 33% of the BAC sequences and the genome, respectively. In particular, the PgDel family accounted for 29% of the genome and presumably played major roles in enlargement of the size of the ginseng genome. Fluorescence in situ hybridization (FISH) revealed that the PgDel1 elements are distributed throughout the chromosomes along dispersed heterochromatic regions except for ribosomal DNA blocks. The intensity of the PgDel2 FISH signals was biased toward 24 out of 48 chromosomes. Unique gene probes showed two pairs of signals with different locations, one pair in subtelomeric regions on PgDel2-rich chromosomes and the other in interstitial regions on PgDel2-poor chromosomes, demonstrating allotetraploidy in ginseng. Our findings promote understanding of the evolution of the ginseng genome and of that of related species in the Araliaceae.

Genome-wide characterization of long intergenic non-coding RNAs (lincRNAs) provides new insight into viral diseases in honey bees Apis cerana and Apis mellifera.

BACKGROUND: Long non-coding RNAs (lncRNAs) are a class of RNAs that do not encode proteins. Recently, lncRNAs have gained special attention for their roles in various biological process and diseases. RESULTS: In an attempt to identify long intergenic non-coding RNAs (lincRNAs) and their possible involvement in honey bee development and diseases, we analyzed RNA-seq datasets generated from Asian honey bee (Apis cerana) and western honey bee (Apis mellifera). We identified 2470 lincRNAs with an average length of 1011 bp from A. cerana and 1514 lincRNAs with an average length of 790 bp in A. mellifera. Comparative analysis revealed that 5 % of the total lincRNAs derived from both species are unique in each species. Our comparative digital gene expression analysis revealed a high degree of tissue-specific expression among the seven major tissues of honey bee, different from mRNA expression patterns. A total of 863 (57 %) and 464 (18 %) lincRNAs showed tissue-dependent expression in A. mellifera and A. cerana, respectively, most preferentially in ovary and fat body tissues. Importantly, we identified 11 lincRNAs that are specifically regulated upon viral infection in honey bees, and 10 of them appear to play roles during infection with various viruses. CONCLUSIONS: This study provides the first comprehensive set of lincRNAs for honey bees and opens the door to discover lincRNAs associated with biological and hormone signaling pathways as well as various diseases of honey bee.

Uncovering the novel characteristics of Asian honey bee, Apis cerana, by whole genome sequencing.

BACKGROUND: The honey bee is an important model system for increasing understanding of molecular and neural mechanisms underlying social behaviors relevant to the agricultural industry and basic science. The western honey bee, Apis mellifera, has served as a model species, and its genome sequence has been published. In contrast, the genome of the Asian honey bee, Apis cerana, has not yet been sequenced. A. cerana has been raised in Asian countries for thousands of years and has brought considerable economic benefits to the apicultural industry. A cerana has divergent biological traits compared to A. mellifera and it has played a key role in maintaining biodiversity in eastern and southern Asia. Here we report the first whole genome sequence of A. cerana. RESULTS: Using de novo assembly methods, we produced a 238 Mbp draft of the A. cerana genome and generated 10,651 genes. A.cerana-specific genes were analyzed to better understand the novel characteristics of this honey bee species. Seventy-two percent of the A. cerana-specific genes had more than one GO term, and 1,696 enzymes were categorized into 125 pathways. Genes involved in chemoreception and immunity were carefully identified and compared to those from other sequenced insect models. These included 10 gustatory receptors, 119 odorant receptors, 10 ionotropic receptors, and 160 immune-related genes. CONCLUSIONS: This first report of the whole genome sequence of A. cerana provides resources for comparative sociogenomics, especially in the field of social insect communication. These important tools will contribute to a better understanding of the complex behaviors and natural biology of the Asian honey bee and to anticipate its future evolutionary trajectory.

Transcriptome analysis reveals in vitro cultured Withania somnifera leaf and root tissues as a promising source for targeted withanolide biosynthesis.

BACKGROUND: The production of metabolites via in vitro culture is promoted by the availability of fully defined metabolic pathways. Withanolides, the major bioactive phytochemicals of Withania somnifera, have been well studied for their pharmacological activities. However, only a few attempts have been made to identify key candidate genes involved in withanolide biosynthesis. Understanding the steps involved in withanolide biosynthesis is essential for metabolic engineering of this plant to increase withanolide production. RESULTS: Transcriptome sequencing was performed on in vitro adventitious root and leaf tissues using the Illumina platform. We obtained a total of 177,156 assembled transcripts with an average unigene length of 1,033 bp. About 13% of the transcripts were unique to in vitro adventitious roots but no unique transcripts were observed in in vitro-grown leaves. A putative withanolide biosynthetic pathway was deduced by mapping the assembled transcripts to the KEGG database, and the expression of candidate withanolide biosynthesis genes -were validated by qRT PCR. The accumulation pattern of withaferin A and withanolide A varied according to the type of tissue and the culture period. Further, we demonstrated that in vitro leaf extracts exhibit anticancer activity against human gastric adenocarcinoma cell lines at sub G1 phase. CONCLUSIONS: We report here a validated large-scale transcriptome data set and the potential biological activity of in vitro cultures of W. somnifera. This study provides important information to enhance tissue-specific expression and accumulation of secondary metabolites, paving the way for industrialization of in vitro cultures of W. somnifera.

Comprehensive analysis of Panax ginseng root transcriptomes.

BACKGROUND: Korean ginseng (Panax ginseng C.A. Meyer) is a highly effective medicinal plant containing ginsenosides with various pharmacological activities, whose roots are produced commercially for crude drugs. RESULTS: Here, we used the Illumina platform to generate over 232 million RNA sequencing reads from four root samples, including whole roots from one-year-old plants and three types of root tissue from six-year-old plants (i.e., main root bodies, rhizomes, and lateral roots). Through de novo assembly and reference-assisted selection, we obtained a non-redundant unigene set consisting of 55,949 transcripts with an average length of 1,250 bp. Among transcripts in the unigene set, 94 % were functionally annotated via similarity searches against protein databases. Approximately 28.6 % of the transcripts represent novel gene sequences that have not previously been reported for P. ginseng. Digital expression profiling revealed 364 genes showing differential expression patterns among the four root samples. Additionally, 32 were uniquely expressed in one-year-old roots, while seven were uniquely expressed in six-year-old root tissues. We identified 38 transcripts encoding enzymes involved in ginsenoside biosynthesis pathways and 189 encoding UDP-glycosyltransferases. CONCLUSION: Our analysis provides new insights into the role of the root transcriptome in development and secondary metabolite biosynthesis in P. ginseng.

Genome-wide SNP identification and QTL mapping for black rot resistance in cabbage.

BACKGROUND: Black rot is a destructive bacterial disease causing large yield and quality losses in Brassica oleracea. To detect quantitative trait loci (QTL) for black rot resistance, we performed whole-genome resequencing of two cabbage parental lines and genome-wide SNP identification using the recently published B. oleracea genome sequences as reference. RESULTS: Approximately 11.5 Gb of sequencing data was produced from each parental line. Reference genome-guided mapping and SNP calling revealed 674,521 SNPs between the two cabbage lines, with an average of one SNP per 662.5 bp. Among 167 dCAPS markers derived from candidate SNPs, 117 (70.1%) were validated as bona fide SNPs showing polymorphism between the parental lines. We then improved the resolution of a previous genetic map by adding 103 markers including 87 SNP-based dCAPS markers. The new map composed of 368 markers and covers 1467.3 cM with an average interval of 3.88 cM between adjacent markers. We evaluated black rot resistance in the mapping population in three independent inoculation tests using F2:3 progenies and identified one major QTL and three minor QTLs. CONCLUSION: We report successful utilization of whole-genome resequencing for large-scale SNP identification and development of molecular markers for genetic map construction. In addition, we identified novel QTLs for black rot resistance. The high-density genetic map will promote QTL analysis for other important agricultural traits and marker-assisted breeding of B. oleracea.

Characterization of three active transposable elements recently inserted in three independent DFR-A alleles and one high-copy DNA transposon isolated from the Pink allele of the ANS gene in onion (Allium cepa L.).

Intact retrotransposon and DNA transposons inserted in a single gene were characterized in onions (Allium cepa) and their transcription and copy numbers were estimated in this study. While analyzing diverse onion germplasm, large insertions in the DFR-A gene encoding dihydroflavonol 4-reductase (DFR) involved in the anthocyanin biosynthesis pathway were found in two accessions. A 5,070-bp long terminal repeat (LTR) retrotransposon inserted in the active DFR-A (R4) allele was identified from one of the large insertions and designated AcCOPIA1. An intact ORF encoded typical domains of copia-like LTR retrotransposons. However, AcCOPIA1 contained atypical 'TG' and 'TA' dinucleotides at the ends of the LTRs. A 4,615-bp DNA transposon was identified in the other large insertion. This DNA transposon, designated AcCACTA1, contained an ORF coding for a transposase showing homology with the CACTA superfamily transposable elements (TEs). Another 5,073-bp DNA transposon was identified from the DFR-A (TRN) allele. This DNA transposon, designated AchAT1, belonged to the hAT superfamily with short 4-bp terminal inverted repeats (TIRs). Finally, a 6,258-bp non-autonomous DNA transposon, designated AcPINK, was identified in the ANS-p allele encoding anthocyanidin synthase, the next downstream enzyme to DFR in the anthocyanin biosynthesis pathway. AcPINK also possessed very short 3-bp TIRs. Active transcription of AcCOPIA1, AcCACTA1, and AchAT1 was observed through RNA-Seq analysis and RT-PCR. The copy numbers of AcPINK estimated by mapping the genomic DNA reads produced by NextSeq 500 were predominantly high compared with the other TEs. A series of evidence indicated that these TEs might have transposed in these onion genes very recently, providing a stepping stone for elucidation of enormously large-sized onion genome structure.

Identification and expression analysis of WRKY family genes under biotic and abiotic stresses in Brassica rapa.

WRKY proteins constitute one of the largest transcription factor families in higher plants, and they are involved in multiple biological processes such as plant development, metabolism, and responses to biotic and abiotic stresses. Genes of this family have been well documented in response to many abiotic and biotic stresses in many plant species, but not yet against Pectobacterium carotovorum subsp. carotovorum and Fusarium oxysporum f.sp. conglutinans in any of the plants. Moreover, potentiality of a specific gene may vary depending on stress conditions and genotypes. To identify stress resistance-related potential WRKY genes of Brassica rapa, we analyzed their expressions against above-mentioned pathogens and cold, salt, and drought stresses in B. rapa. Stress resistance-related functions of all Brassica rapa WRKY (BrWRKY) genes were firstly analyzed through homology study with existing biotic and abiotic stress resistance-related WRKY genes of other plant species and found a high degree of homology. We then identified all BrWRKY genes in a Br135K microarray dataset, which was created by applying low-temperature stresses to two contrasting Chinese cabbage doubled haploid (DH) lines, Chiifu and Kenshin, and selected 41 BrWRKY genes with high and differential transcript abundance levels. These selected genes were further investigated under cold, salt, and drought stresses as well as after infection with P. carotovorum subsp. carotovorum and F. oxysporum f.sp. conglutinans in B. rapa. The selected genes showed an organ-specific expression, and 22 BrWRKY genes were differentially expressed in Chiifu compared to Kenshin under cold and drought stresses. Six BrWRKY genes were more responsive in Kenshin compared to Chiffu under salt stress. In addition, eight BrWRKY genes showed differential expression after P. carotovorum subsp. carotovorum infection and five genes after F. oxysporum f.sp. conglutinans infection in B. rapa. Thus, the differentially expressed BrWRKY genes might be potential resources for molecular breeding of Brassica crops against abiotic and biotic stresses and several genes, which showed differential expressions commonly in response to several stresses, might be useful for multiple stress resistance. These findings would also be helpful in resolving the complex regulatory mechanism of WRKY genes in stress resistance and for this further functional genomics study of these potential genes in different Brassica crops is essential.

Exceptional reduction of the plastid genome of saguaro cactus (Carnegiea gigantea): Loss of the ndh gene suite and inverted repeat.

UNLABELLED: • PREMISE OF THE STUDY: Land-plant plastid genomes have only rarely undergone significant changes in gene content and order. Thus, discovery of additional examples adds power to tests for causes of such genome-scale structural changes.• METHODS: Using next-generation sequence data, we assembled the plastid genome of saguaro cactus and probed the nuclear genome for transferred plastid genes and functionally related nuclear genes. We combined these results with available data across Cactaceae and seed plants more broadly to infer the history of gene loss and to assess the strength of phylogenetic association between gene loss and loss of the inverted repeat (IR).• KEY RESULTS: The saguaro plastid genome is the smallest known for an obligately photosynthetic angiosperm (∼113 kb), having lost the IR and plastid ndh genes. This loss supports a statistically strong association across seed plants between the loss of ndh genes and the loss of the IR. Many nonplastid copies of plastid ndh genes were found in the nuclear genome, but none had intact reading frames; nor did three related nuclear-encoded subunits. However, nuclear pgr5, which functions in a partially redundant pathway, was intact.• CONCLUSIONS: The existence of an alternative pathway redundant with the function of the plastid NADH dehydrogenase-like complex (NDH) complex may permit loss of the plastid ndh gene suite in photoautotrophs like saguaro. Loss of these genes may be a recurring mechanism for overall plastid genome size reduction, especially in combination with loss of the IR.