Twelve newly assembled jasmine chloroplast genomes: unveiling genomic diversity, phylogenetic relationships and evolutionary patterns among Oleaceae and Jasminum species.

BACKGROUND: Jasmine (Jasminum), renowned for its ornamental value and captivating fragrance, has given rise to numerous species and accessions. However, limited knowledge exists regarding the evolutionary relationships among various Jasminum species. RESULTS: In the present study, we sequenced seven distinct Jasminum species, resulting in the assembly of twelve high-quality complete chloroplast (cp) genomes. Our findings revealed that the size of the 12 cp genomes ranged from 159 to 165 kb and encoded 134-135 genes, including 86-88 protein-coding genes, 38-40 tRNA genes, and 8 rRNA genes. J. nudiflorum exhibited a larger genome size compared to other species, mainly attributed to the elevated number of forward repeats (FRs). Despite the typically conservative nature of chloroplasts, variations in the presence or absence of accD have been observed within J. sambac. The calculation of nucleotide diversity (Pi) values for 19 cp genomes indicated that potential mutation hotspots were more likely to be located in LSC regions than in other regions, particularly in genes ycf2, rbcL, atpE, ndhK, and ndhC (Pi > 0.2). Ka/Ks values revealed strong selection pressure on the genes rps2, atpA, rpoA, rpoC1, and rpl33 when comparing J. sambac with the three most closely related species (J. auriculatum, J. multiflorum, and J. dichotomum). Additionally, SNP identification, along with the results of Structure, PCA, and phylogenetic tree analyses, divided the Jasminum cp genomes into six groups. Notably, J. polyanthum showed gene flow signals from both the G5 group (J. nudiflorum) and the G3 group (J. tortuosum and J. fluminense). Phylogenetic tree analysis reflected that most species from the same genus clustered together with robust support in Oleaceae, strongly supporting the monophyletic nature of cp genomes within the genus Jasminum. CONCLUSION: Overall, this study provides comprehensive insights into the genomic composition, variation, and phylogenetic relationships among various Jasminum species. These findings enhance our understanding of the genetic diversity and evolutionary history of Jasminum.

Genomic and cytogenetic analyses reveal satellite repeat signature in allotetraploid okra (Abelmoschus esculentus).

BACKGROUND: Satellite repeats are one of the most rapidly evolving components in eukaryotic genomes and play vital roles in genome regulation, genome evolution, and speciation. As a consequence, the composition, abundance and chromosome distribution of satellite repeats often exhibit variability across various species, genome, and even individual chromosomes. However, we know little about the satellite repeat evolution in allopolyploid genomes. RESULTS: In this study, we investigated the satellite repeat signature in five okra (Abelmoschus esculentus) accessions using genomic and cytogenetic methods. In each of the five accessions, we identified eight satellite repeats, which exhibited a significant level of intraspecific conservation. Through fluorescence in situ hybridization (FISH) experiments, we observed that the satellite repeats generated multiple signals and exhibited variations in copy number across chromosomes. Intriguingly, we found that five satellite repeats were interspersed with centromeric retrotransposons, signifying their involvement in centromeric satellite repeat identity. We confirmed subgenome-biased amplification patterns of these satellite repeats through existing genome assemblies or dual-color FISH, indicating their distinct dynamic evolution in the allotetraploid okra subgenome. Moreover, we observed the presence of multiple chromosomes harboring the 35 S rDNA loci, alongside another chromosomal pair carrying the 5 S rDNA loci in okra using FISH assay. Remarkably, the intensity of 35 S rDNA hybridization signals varied among chromosomes, with the signals predominantly localized within regions of relatively weak DAPI staining, associated with GC-rich heterochromatin regions. Finally, we observed a similar localization pattern between 35 S rDNA and three satellite repeats with high GC content and confirmed their origin in the intergenic spacer region of the 35 S rDNA. CONCLUSIONS: Our findings uncover a unique satellite repeat signature in the allotetraploid okra, contributing to our understanding of the composition, abundance, and chromosomal distribution of satellite repeats in allopolyploid genomes, further enriching our understanding of their evolutionary dynamics in complex allopolyploid genomes.

Microbiota Alters and Its Correlation with Molecular Regulation Underlying Depression in PCOS Patients.

Depression is one of the complications in patients with polycystic ovary syndrome (PCOS) that leads to considerable mental health. Accumulating evidence suggests that human gut microbiomes are associated with the progression of PCOS and depression. However, whether microbiota influences depression development in PCOS patients is still uncharacterized. In this study, we employed metagenomic sequencing and transcriptome sequencing (RNA-seq) to profile the composition of the fecal microbiota and gene expression of peripheral blood mononuclear cells in depressed women with PCOS (PCOS-DP, n = 27) in comparison to mentally healthy women with PCOS (PCOS, n = 18) and compared with healthy control (HC, n = 27) and patients with major depressive disorder (MDD, n = 29). Gut microbiota assessment revealed distinct patterns in the relative abundance in the PCOS-DP compared to HC, MDD, and PCOS groups. Several gut microbes exhibited uniquely and significantly higher abundance in the PCOS-DP compared to PCOS patients, inducing EC Ruminococcus torques, Coprococcus comes, Megasphaera elsdenii, Acidaminococcus intestini, and Barnesiella viscericola. Bacteroides eggerthii was a potential gut microbial biomarker for the PCOS-DP. RNA-seq profiling identified that 35 and 37 genes were significantly elevated and downregulated in the PCOS-DP, respectively. The enhanced differential expressed genes (DEGs) in the PCOS-DP were enriched in pathways involved in signal transduction and endocrine and metabolic diseases, whereas several lipid metabolism pathways were downregulated. Intriguingly, genes correlated with the gut microbiota were found to be significantly enriched in pathways of neurodegenerative diseases and the immune system, suggesting that changes in the microbiota may have a systemic impact on the expression of neurodegenerative diseases and immune genes. Gut microbe-related DEGs of CREB3L3 and CCDC173 were possible molecular biomarkers and therapeutic targets of women with PCOS-DP. Our multi-omics data indicate shifts in the gut microbiome and host gene regulation in PCOS patients with depression, which is of possible etiological and diagnostic importance.

The complete mitochondrial genome of Isochrysis galbana harbors a unique repeat structure and a specific trans-spliced cox1 gene.

The haptophyte Isochrysis galbana is considered as a promising source for food supplements due to its rich fucoxanthin and polyunsaturated fatty acids content. Here, the I. galbana mitochondrial genome (mitogenome) was sequenced using a combination of Illumina and PacBio sequencing platforms. This 39,258 bp circular mitogenome has a total of 46 genes, including 20 protein-coding genes, 24 tRNA genes and two rRNA genes. A large block of repeats (~12.7 kb) was segregated in one region of the mitogenome, accounting for almost one third of the total size. A trans-spliced gene cox1 was first identified in I. galbana mitogenome and was verified by RNA-seq and DNA-seq data. The massive expansion of tandem repeat size and cis- to trans-splicing shift could be explained by the high mitogenome rearrangement rates in haptophytes. Strict SNP calling based on deep transcriptome sequencing data suggested the lack of RNA editing in both organelles in this species, consistent with previous studies in other algal lineages. To gain insight into haptophyte mitogenome evolution, a comparative analysis of mitogenomes within haptophytes and among eight main algal lineages was performed. A core gene set of 15 energy and metabolism genes is present in haptophyte mitogenomes, consisting of 1 cob, 3 cox, 7 nad, 2 atp and 2 ribosomal genes. Gene content and order was poorly conserved in this lineage. Haptophyte mitogenomes have lost many functional genes found in many other eukaryotes including rps/rpl, sdh, tat, secY genes, which make it contain the smallest gene set among all algal taxa. All these implied the rapid-evolving and more recently evolved mitogenomes of haptophytes compared to other algal lineages. The phylogenetic tree constructed by cox1 genes of 204 algal mitogenomes yielded well-resolved internal relationships, providing new evidence for red-lineages that contained plastids of red algal secondary endosymbiotic origin. This newly assembled mitogenome will add to our knowledge of general trends in algal mitogenome evolution within haptophytes and among different algal taxa.

Genomic and transcriptomic-based analysis of agronomic traits in sugar beet (Beta vulgaris L.) pure line IMA1.

Sugar beet (Beta vulgaris L.) is an important sugar-producing and energy crop worldwide. The sugar beet pure line IMA1 independently bred by Chinese scientists is a standard diploid parent material that is widely used in hybrid-breeding programs. In this study, a high-quality, chromosome-level genome assembly for IMA1was conducted, and 99.1% of genome sequences were assigned to nine chromosomes. A total of 35,003 protein-coding genes were annotated, with 91.56% functionally annotated by public databases. Compared with previously released sugar beet assemblies, the new genome was larger with at least 1.6 times larger N50 size, thereby substantially improving the completeness and continuity of the sugar beet genome. A Genome-Wide Association Studies analysis identified 10 disease-resistance genes associated with three important beet diseases and five genes associated with sugar yield per hectare, which could be key targets to improve sugar productivity. Nine highly expressed genes associated with pollen fertility of sugar beet were also identified. The results of this study provide valuable information to identify and dissect functional genes affecting sugar beet agronomic traits, which can increase sugar beet production and help screen for excellent sugar beet breeding materials. In addition, information is provided that can precisely incorporate biotechnology tools into breeding efforts.

Multi-omics Analyses Provide Insight into the Biosynthesis Pathways of Fucoxanthin in Isochrysis galbana.

Isochrysis galbana is considered an ideal bait for functional foods and nutraceuticals of humans because of its high fucoxanthin (Fx) content. However, multi-omics analysis of the regulatory networks for Fx biosynthesis in I. galbana has not been reported. In this study, we report a high-quality genome assembly of I. galbana LG007, which has a genome size of 92.73 Mb, with a contig N50 of 6.99 Mb and 14,900 protein-coding genes. Phylogenetic analysis confirmed the monophyly of Haptophyta, with I. galbana sister to Emiliania huxleyi and Chrysochromulina tobinii. Evolutionary analysis revealed an estimated divergence time between I. galbana and E. huxleyi of âˆ¼ 133 million years ago. Gene family analysis indicated that lipid metabolism-related genes exhibited significant expansion, including IgPLMT, IgOAR1, and IgDEGS1. Metabolome analysis showed that the content of carotenoids in I. galbana cultured under green light for 7 days was higher than that under white light, and ß-carotene was the main carotenoid, accounting for 79.09% of the total carotenoids. Comprehensive multi-omics analysis revealed that the content of ß-carotene, antheraxanthin, zeaxanthin, and Fx was increased by green light induction, which was significantly correlated with the expression of IgMYB98, IgZDS, IgPDS, IgLHCX2, IgZEP, IgLCYb, and IgNSY. These findings contribute to the understanding of Fx biosynthesis and its regulation, providing a valuable reference for food and pharmaceutical applications.

Epigenetic silencing of E-cadherin gene induced by lncRNA MALAT-1 in acute myeloid leukaemia.

BACKGROUND: Epigenetic abnormalities in acute myeloid leukaemia provide us with a target for novel therapeutic strategies. The aim of the study was to verify the epigenetic regulatory mechanism of E-cadherin gene silencing induced by long non-coding RNA MALAT-1 in AML. METHODS: Expression of MALAT-1, E-cadherin, EZH2, SUZ12 and EED genes in AML patients was detected by RT-qPCR. After MALAT-1 silencing in AML cell lines, levels of the E-cadherin, EZH2, SUZ12, EED, DNMT1, DNMT3A and DNMT3B genes and encoded proteins were detected by RT-qPCR and Western blotting. The level of CpG island methylation and trimethylation modification of histone H3K27 in the promoter region of E-cadherin was detected by pyrosequencing and ChIP-qPCR. RIP-qPCR was used to detect the interaction between MALAT-1 and proteins. RESULTS: MALAT-1, EZH2 and EED gene expression was markedly increased in AML patients with E-cadherin down-regulation. A positive correlation between EZH2 or SUZ12 and MALAT-1 expression was observed. After MALAT-1 silencing, the expression of E-cadherin was up-regulated, whereas the expression of EZH2, SUZ12, DNMT1, DNMT3A and DNMT3B was down-regulated. Results of Western blotting were consistent with those of RT-qPCR. Methylation levels of E-cadherin in AML patients were higher than that in normal controls, which appeared to increase with age. Methylation of the CpG island and H3K27 trimethylation of E-cadherin were decreased after MALAT-1 silencing. RIP-qPCR suggested that MALAT-1 might be enriched by EZH2 and SUZ12. CONCLUSION: Our findings verified that MALAT-1 might lead to the transcriptional silencing of E-cadherin gene through the trimethylation of H3K27 mediated by recruiting EZH2 and SUZ12.

A survey of transcriptome complexity using full-length isoform sequencing in the tea plant Camellia sinensis.

Tea is one of the most popular beverages and its leaves are rich in catechins, contributing to the diverse flavor as well as beneficial for human health. However, the study of the post-transcriptional regulatory mechanism affecting the synthesis of catechins remains insufficient. Here, we sequenced the transcriptome using PacBio sequencing technology and obtained 63,111 full-length high-quality isoforms, including 1302 potential novel genes and 583 highly reliable fusion transcripts. We also identified 1204 lncRNAs with high quality, containing 188 known and 1016 novel lncRNAs. In addition, 311 mis-annotated genes were corrected based on the high-quality Isoseq reads. A large number of alternative splicing (AS) events (3784) and alternative polyadenylation (APA) genes (18,714) were analyzed, accounting for 8.84% and 43.7% of the total annotated genes, respectively. We also found that 2884 genes containing AS and APA features exhibited higher expression levels than other genes. These genes are mainly involved in amino acid biosynthesis, carbon fixation in photosynthetic organisms, phenylalanine, tyrosine, tryptophan biosynthesis, and pyruvate metabolism, suggesting that they play an essential role in the catechins content of tea polyphenols. Our results further improved the level of genome annotation and indicated that post-transcriptional regulation plays a crucial part in synthesizing catechins.

SunUp and Sunset genomes revealed impact of particle bombardment mediated transformation and domestication history in papaya.

Transgenic papaya is widely publicized for controlling papaya ringspot virus. However, the impact of particle bombardment on the genome remains unknown. The transgenic SunUp and its progenitor Sunset genomes were assembled into 351.5 and 350.3 Mb in nine chromosomes, respectively. We identified a 1.64 Mb insertion containing three transgenic insertions in SunUp chromosome 5, consisting of 52 nuclear-plastid, 21 nuclear-mitochondrial and 1 nuclear genomic fragments. A 591.9 kb fragment in chromosome 5 was translocated into the 1.64 Mb insertion. We assembled a gapless 9.8 Mb hermaphrodite-specific region of the Yh chromosome and its 6.0 Mb X counterpart. Resequencing 86 genomes revealed three distinct groups, validating their geographic origin and breeding history. We identified 147 selective sweeps and defined the essential role of zeta-carotene desaturase in carotenoid accumulation during domestication. Our findings elucidated the impact of particle bombardment and improved our understanding of sex chromosomes and domestication to expedite papaya improvement.

Genome-Wide Identification and Expression Analysis of the R2R3-MYB Transcription Factor Family Revealed Their Potential Roles in the Flowering Process in Longan (Dimocarpus longan).

Longan (Dimocarpus longan Lour.) is a productive fruit crop with high nutritional and medical value in tropical and subtropical regions. The MYB gene family is one of the most widespread plant transcription factor (TF) families participating in the flowering regulation. However, little is known about the MYB TFs involved in the flowering process in longan and its regulatory network. In this study, a total of 119 DlR2R3-MYB genes were identified in the longan genome and were phylogenetically grouped into 28 subgroups. The groupings were supported by highly conserved gene structures and motif composition of DlR2R3-MYB genes in each subgroup. Collinearity analysis demonstrated that segmental replications played a more crucial role in the expansion of the DlR2R3-MYB gene family compared to tandem duplications, and all tandem/segmental duplication gene pairs have evolved under purifying selection. Interspecies synteny analysis among longan and five representative species implied the occurrence of gene duplication events was one of the reasons contributing to functional differentiation among species. RNA-seq data from various tissues showed DlR2R3-MYB genes displayed tissue-preferential expression patterns. The pathway of flower development was enriched with six DlR2R3-MYB genes. Cis-acting element prediction revealed the putative functions of DlR2R3-MYB genes were related to the plant development, phytohormones, and environmental stresses. Notably, the orthologous counterparts between Arabidopsis and longan R2R3-MYB members tended to play conserved roles in the flowering regulation and stress responses. Transcriptome profiling on off-season flower induction (FI) by KClO3 indicated two up-regulated and four down-regulated DlR2R3-MYB genes involved in the response to KClO3 treatment compared with control groups. Additionally, qRT-PCR confirmed certain genes exhibited high expression in flowers/flower buds. Subcellular localization experiments revealed that three predicted flowering-associated MYB proteins were localized in the nucleus. Future functional studies on these potential candidate genes involved in the flowering development could further the understanding of the flowering regulation mechanism.

The complete chloroplast genome of the Chinese banyan tree Ficus microcarpa.

Banyan tree or Ficus microcarpa is a large perennial plant with extraordinary aerial roots from the Moraceae family. In this study, the complete chloroplast genome sequence of F. microcarpa was assembled using PacBio data. The chloroplast genome size is 141,611 bp, consisting of a large single-copy (LSC) region and a small single-copy (SSC) region of 101,835 bp and 9,676 bp, respectively, which are separated by a pair of 15,050 bp inverted repeat (IR) regions. The genome includes 74 protein-coding genes, 43 tRNA genes, and 8 rRNA genes. A phylogenetic tree reconstructed by 25 complete chloroplast genomes reveals that F. microcarpa is mostly related to Ficus racemosa.

[The Effect of MicroRNA-143 on the Proliferation and Apoptosis of Leukemia Cell Line U-937].

OBJECTIVE: To investigate the expression of microRNA-143 (miR-143) in patients with acute myeloid leukemia (AML), and the effect of miR-143 over-expression on the proliferation and apoptosis of AML cells. And to verify the targeting relationship between miR-143 and long non-coding RNA MALAT-1. So as to explore the possible regulatory mechanism of miR-143 in the pathogenesis of AML. METHODS: The expression of miR-143 in bone marrow cells of AML patients was detected by RT-qPCR. After miR-143 was over-expressed in U-937 cell lines, the proliferation of U-937 cell lines was detected by CCK-8, clone formation assay, and flow cytometry. In addition, cell apoptosis was detected by Hoechst 33258 fluorescence staining and flow cytometry. At the same time, bioinformatics, RT-qPCR and dual luciferase reporter gene assay were used to predicted and verified the targeting relationship between miR-143 and MALAT-1. RESULTS: Expression of miR-143 in AML patients was significantly lower than those in normal controls. Over-expressed miR-143 could inhibit the proliferation of U-937 cells and promote the apoptosis of the cell. The miR-143 binding site was located on the MALAT-1 RNA sequence, and MALAT-1 was down-regulated in U-937 cells after over-expressed miR-143. However, the expression of miR-143 showed no significantly changed after MALAT-1 silencing. CONCLUSION: Expression of miR-143 in AML patients is lower than that in normal controls. Over-expression of miR-143 can inhibit the proliferation of U-937 cells and promote its apoptosis. And its mechanism may be related to its targe regulation of MALAT-1.

Auxin and cytokinin mediated regulation involved in vitro organogenesis of papaya.

In vitro organogenesis is a multistep process which is largely controlled by the balance between auxin and cytokinin. Previous studies revealed a complex network regulating in vitro organogenesis in Arabidopsis thaliana; however, our knowledge of the molecular mechanisms underlying de novo shoot formation in papaya (Carica papaya) remains limited. Here, we optimized multiple factors to achieve an efficient and reproducible protocol for the induction of papaya callus formation and shoot regeneration. Subsequently, we analyzed the dynamic transcriptome profiles of samples undergoing this process, identified 5381, 642, 4047, and 2386 differentially expressed genes (DEGs), including 447, 66, 350, and 263 encoding transcription factors (TFs), in four stage comparisons. The DEGs were mainly involved in phytohormone modulation and transduction processes, particularly for auxin and cytokinin. Of these, 21 and 7 candidate genes involved in the auxin and cytokinin pathways, respectively, had distinct expression patterns throughout in vitro organogenesis. Furthermore, we found two genes encoding key TFs, CpLBD19 and CpESR1, were sharply induced on callus induction medium and shoot induction medium, indicating these two TFs may serve as proxies for callus induction and shoot formation in papaya. We therefore report a regulatory network of auxin and cytokinin signaling in papaya according to the one previously modeled for Arabidopsis. Our comprehensive analyses provide insight into the early molecular regulation of callus initiation and shoot formation in papaya, and are useful for the further identification of the regulators governing in vitro organogenesis.

Comparative Analysis of the MADS-Box Genes Revealed Their Potential Functions for Flower and Fruit Development in Longan (Dimocarpus longan).

Longan (Dimocarpus longan Lour.) is an important economic crop widely planted in tropical and subtropical regions, and flower and fruit development play decisive effects on the longan yield and fruit quality formation. MCM1, AGAMOUS, DEFICIENS, Serum Response Factor (MADS)-box transcription factor family plays important roles for the flowering time, floral organ identity, and fruit development in plants. However, there is no systematic information of MADS-box family in longan. In this study, 114 MADS-box genes were identified from the longan genome, phylogenetic analysis divided them into type I (Mα, Mß, Mγ) and type II (MIKC*, MIKC C ) groups, and MIKC C genes were further clustered into 12 subfamilies. Comparative genomic analysis of 12 representative plant species revealed the conservation of type II in Sapindaceae and analysis of cis-elements revealed that Dof transcription factors might directly regulate the MIKC C genes. An ABCDE model was proposed for longan based on the phylogenetic analysis and expression patterns of MADS-box genes. Transcriptome analysis revealed that MIKC C genes showed wide expression spectrums, particularly in reproductive organs. From 35 days after KClO3 treatment, 11 MIKC genes were up-regulated, suggesting a crucial role in off-season flower induction, while DlFLC, DlSOC1, DlSVP, and DlSVP-LIKE may act as the inhibitors. The gene expression patterns of longan fruit development indicated that DlSTK, DlSEP1/2, and DlMADS53 could be involved in fruit growth and ripening. This paper carried out the whole genome identification and analysis of the longan MADS-box family for the first time, which provides new insights for further understanding its function in flowers and fruit.

The evolutionary origin and domestication history of goldfish (Carassius auratus).

Goldfish have been subjected to over 1,000 y of intensive domestication and selective breeding. In this report, we describe a high-quality goldfish genome (2n = 100), anchoring 95.75% of contigs into 50 pseudochromosomes. Comparative genomics enabled us to disentangle the two subgenomes that resulted from an ancient hybridization event. Resequencing 185 representative goldfish variants and 16 wild crucian carp revealed the origin of goldfish and identified genomic regions that have been shaped by selective sweeps linked to its domestication. Our comprehensive collection of goldfish varieties enabled us to associate genetic variations with a number of well-known anatomical features, including features that distinguish traditional goldfish clades. Additionally, we identified a tyrosine-protein kinase receptor as a candidate causal gene for the first well-known case of Mendelian inheritance in goldfish-the transparent mutant. The goldfish genome and diversity data offer unique resources to make goldfish a promising model for functional genomics, as well as domestication.

Genomic variation between PRSV resistant transgenic SunUp and its progenitor cultivar Sunset.

BACKGROUND: The safety of genetically transformed plants remains a subject of scrutiny. Genomic variants in PRSV resistant transgenic papaya will provide evidence to rationally address such concerns. RESULTS: In this study, a total of more than 74 million Illumina reads for progenitor 'Sunset' were mapped onto transgenic papaya 'SunUp' reference genome. 310,364 single nucleotide polymorphisms (SNPs) and 34,071 small Inserts/deletions (InDels) were detected between 'Sunset' and 'SunUp'. Those variations have an uneven distribution across nine chromosomes in papaya. Only 0.27% of mutations were predicted to be high-impact mutations. ATP-related categories were highly enriched among these high-impact genes. The SNP mutation rate was about 8.4 × 10- 4 per site, comparable with the rate induced by spontaneous mutation over numerous generations. The transition-to-transversion ratio was 1.439 and the predominant mutations were C/G to T/A transitions. A total of 3430 nuclear plastid DNA (NUPT) and 2764 nuclear mitochondrial DNA (NUMT) junction sites have been found in 'SunUp', which is proportionally higher than the predicted total NUPT and NUMT junction sites in 'Sunset' (3346 and 2745, respectively). Among all nuclear organelle DNA (norgDNA) junction sites, 96% of junction sites were shared by 'SunUp' and 'Sunset'. The average identity between 'SunUp' specific norgDNA and corresponding organelle genomes was higher than that of norgDNA shared by 'SunUp' and 'Sunset'. Six 'SunUp' organelle-like borders of transgenic insertions were nearly identical to corresponding sequences in organelle genomes (98.18 ~ 100%). None of the paired-end spans of mapped 'Sunset' reads were elongated by any 'SunUp' transformation plasmid derived inserts. Significant amounts of DNA were transferred from organelles to the nuclear genome during bombardment, including the six flanking sequences of the three transgenic insertions. CONCLUSIONS: Comparative whole-genome analyses between 'SunUp' and 'Sunset' provide a reliable estimate of genome-wide variations and evidence of organelle-to-nucleus transfer of DNA associated with biolistic transformation.

Improved bioethanol production using CRISPR/Cas9 to disrupt the ADH2 gene in Saccharomyces cerevisiae.

Bioethanol, as a form of renewable and clean energy, has become increasingly important to the energy supply. One major obstacle in ethanol production is developing a high-capacity system. Existing approaches for regulating the ethanol production pathway are relatively insufficient, with nonspecific genetic manipulation. Here, we used CRISPR/Cas9 technology to disrupt the alcohol dehydrogenase (ADH) 2 gene via complete deletion of the gene and introduction of a frameshift mutation in the ADH2 locus. Sequencing demonstrated the accurate knockout of the target gene with 91.4% and near 100% targeting efficiency. We also utilized genome resequencing to validate the mutations in the ADH2 mutants targeted by various single-guide RNAs. This extensive analysis indicated the mutations in the CRISPR/Cas9-engineered strains were homozygous. We applied the engineered Saccharomyces cerevisiae strains for bioethanol production. Results showed that the ethanol yield improved by up to 74.7% compared with the yield obtained using the native strain. This work illustrates the applicability of this highly efficient and specific genome engineering approach to promote the improvement of bioethanol production in S. cerevisiae via metabolic engineering. Importantly, this study is the first report of the disruption of a target gene, ADH2, in S. cerevisiae using CRISPR/Cas9 technology to improve bioethanol yield.

Genotype-Corrector: improved genotype calls for genetic mapping in F2 and RIL populations.

F2 and recombinant inbred lines (RILs) populations are very commonly used in plant genetic mapping studies. Although genome-wide genetic markers like single nucleotide polymorphisms (SNPs) can be readily identified by a wide array of methods, accurate genotype calling remains challenging, especially for heterozygous loci and missing data due to low sequencing coverage per individual. Therefore, we developed Genotype-Corrector, a program that corrects genotype calls and imputes missing data to improve the accuracy of genetic mapping. Genotype-Corrector can be applied in a wide variety of genetic mapping studies that are based on low coverage whole genome sequencing (WGS) or Genotyping-by-Sequencing (GBS) related techniques. Our results show that Genotype-Corrector achieves high accuracy when applied to both synthetic and real genotype data. Compared with using raw or only imputed genotype calls, the linkage groups built by corrected genotype data show much less noise and significant distortions can be corrected. Additionally, Genotype-Corrector compares favorably to the popular imputation software LinkImpute and Beagle in both F2 and RIL populations. Genotype-Corrector is publicly available on GitHub at https://github.com/freemao/Genotype-Corrector .

Transcriptome Profiling Revealed Stress-Induced and Disease Resistance Genes Up-Regulated in PRSV Resistant Transgenic Papaya.

Papaya is a productive and nutritious tropical fruit. Papaya Ringspot Virus (PRSV) is the most devastating pathogen threatening papaya production worldwide. Development of transgenic resistant varieties is the most effective strategy to control this disease. However, little is known about the genome-wide functional changes induced by particle bombardment transformation. We conducted transcriptome sequencing of PRSV resistant transgenic papaya SunUp and its PRSV susceptible progenitor Sunset to compare the transcriptional changes in young healthy leaves prior to infection with PRSV. In total, 20,700 transcripts were identified, and 842 differentially expressed genes (DEGs) randomly distributed among papaya chromosomes. Gene ontology (GO) category analysis revealed that microtubule-related categories were highly enriched among these DEGs. Numerous DEGs related to various transcription factors, transporters and hormone biosynthesis showed clear differences between the two cultivars, and most were up-regulated in transgenic papaya. Many known and novel stress-induced and disease-resistance genes were most highly expressed in SunUp, including MYB, WRKY, ERF, NAC, nitrate and zinc transporters, and genes involved in the abscisic acid, salicylic acid, and ethylene signaling pathways. We also identified 67,686 alternative splicing (AS) events in Sunset and 68,455 AS events in SunUp, mapping to 10,994 and 10,995 papaya annotated genes, respectively. GO enrichment for the genes displaying AS events exclusively in Sunset was significantly different from those in SunUp. Transcriptomes in Sunset and transgenic SunUp are very similar with noteworthy differences, which increased PRSV-resistance in transgenic papaya. No detrimental pathways and allergenic or toxic proteins were induced on a genome-wide scale in transgenic SunUp. Our results provide a foundation for unraveling the mechanism of PRSV resistance in transgenic papaya.