Complete T2T genome assembly of sweet cherry. Chromosome doubled sweet cherry. Mutations that affecting fruit color related genes. Sweet cherry (Prunus avium L. cv. Tieton) is a famous commercial variety for its high nutritional and healthcare value fruit, with the draft genome been assembled. At present, fundamental research on sweet cherry are mainly based on the draft genomes [1-3]. However, genome evolution based on high-quality genome assembly that provides large collinear regions has never been conducted. Meanwhile, the research on genomic and phenotypic changes of chromosome doubled variety remain to be carried out. In view of the lack of high-quality genomic information to promote sweet cherry genomic research, we used Tieton to assemble the complete 341.62 Mb telomere-to-telomere (T2T) genome after high-throughput chromosome conformation capture (Hi-C) correction. We used advanced sequencing technology third−generation circular consensus sequencing (CCS) and Hi-C to assemble a high-quality genome. Furthermore, the new characteristics of evolution and fruit color change-related genes of chromosome-doubled sweet cherry were investigated using this T2T genome. We analyzed repetitive sequences and coding genes, and studied the evolutionary relationship and genetic variations between sweet cherries and other Rosaceae plants. On the basis of RNA-seq and resequencing analysis, we were able to identify differentially expressed genes (DEGs) in ripening fruits and mutations that occurred when the chromosome of sweet cherry were doubled, revealing the changes of potential fruit color change genes after chromosome doubling. Taken together, our study presents the latest up-to-date complete T2T genome of Tieton, shedding new insights into genomic evolution and alterations during chromosome doubling, and potential changes of fruit color genes. To assembly the genome, 18.46 Gb (50.02×) MGISEQ reads, 20.94 Gb (61.32×) PacBioHiFi reads, and 39.73 Gb (96.79×) Hi−C data were used (Table S1). After removing the contaminants, organelle sequences, and duplicated contigs, a final assembly with a total size of 341,620,392 bp and a N50 length of 39.81 Mb was generated by integrating the published genome and Hi−C data. This assembly consists of eight contigs, named Chr1−Chr8 in descending order of reported information. The BUSCO evaluation showed that the genome integrity was 98.40% (Figure 1A,B and Table S2), with about 59.29% repetitive sequences (including TEs [transposons]) (Tables S3−S4). A total of 58,204 protein-coding genes were predicted with different pipelines, all distributed in eight chromosomes, with 56,822 (97.63%) having functional annotations (Table S5). Telomere sequences were identified in all eight chromosomes, ranging from 1448 to 3297 bp (Figure S2 and Table S6). The centromeres were predicted based on the long tandem repeat sequence and Hi−C data (Figures S2−S3). In addition, the centromere region usually lacks genes, and the type of repeats varied in different centromeres, such as LTR (long terminal repeats)/Copia or Gypsy repeats (Figures S4−S5). All the previous reported 2152 gaps were well closed in this T2T genome (Figure 1A,F). Further, the new genome found 205 new DEGs, 512 increased structure variants (SVs) and 9.55 increased LAI (LTR assembly index value) index, comparing with the previous published genome (Figure 1A) [2], indicating the high quality of this new genome (Table S2). With this high-quality genome, a rooted tree was constructed (Figure 1C). Based on molecular clock analysis, Prunus diverged from Malus domestica in 64.04 million years ago (Mya) and other species evaluated gradually from Prunus avium approximately 48.39 Mya (Figure 1C). This large time span (95% confidence interval) was owing to the low quality of other genomes and no fossil calibration information, and we inferred a relatively recent divergence when comparing with other species [3]. We further conducted expansion and contraction analysis and discovered 3836 expanded and 2087 contracted families in Prunus avium (Figures S6−S7) compared with 1506 expanded and 4054 contracted families in Prunus armeniaca (Figure 1C). No recent whole genome duplication events were found in the genome of Prunus avium, which is consistent with the 4DTv curves, while the slightly upper curve may due to the gene family expansion (Figure S8−S9) [3]. The gene family distribution of the 15 analyzed species (Figure 1C) was shown in Figures S10−S11 and we found 322 unique gene families in the new genome. Collinearity analysis showed that the percentage of collinear genes was high, indicating the close relationship in Prunus. Several collinear genes that played roles in fruit ripening were detected in 10 Prunus species in Figure 1C (Figure S12), thus we further analyzed the fruit ripening-related genes. One of the most urgent tasks in sweet cherry cultivation is to create tetraploid sweet cherry germplasm, by which the disease resistance, flavor, and color genes can be introduced from diverse homoploid Chinese cherry resources [3]. Therefore, studying chromosome doubled genomic changes not only helps to identify trait-related genes but also has important practical significance for creating new sweet cherry germplasm. We first obtained resequencing and transcriptomic data for Tieton diploid (T2X) and tetraploid (T4X) individuals, which were created and observed for more than 10 years (Figures 1D,E and S1; Table S1). Using this data set and our new gap-free genome (Figure 1F), 3046 DEGs (1718 down and 1328 up) were found between fruits of the T2X and T4X (Figures 1A and S13−S14). The key pathways that these DEGs are involved were further investigated by the time series, WGCNA, and KEGG analysis (Figures S15−S21). To explore to what extent the tetraploid mutations influence the found flavonoids and anthocyanin pathways, 1,121,762 high-quality SNPs, 250,919 INDELs and 6948 SVs were identified when comparing T4X data with the new T2X genome (Figure 1G and Table S7), with influenced (silenced) 85.71% of the flavonoids and anthocyanin DEGs observed by chromosome doubling site spectrum (Figure 1H and Table S8). These newly found mutations that influenced coexpressed transcription factors (Table S9) will provide potentially new targets for further elucidating color change mechanism of chromosome-doubled sweet cherries and fill the blank of genomic change mechanism of chromosome-doubled plants [4]. Xin Zhang: Conceptualization; writing original draft, methodology, writing review and editing; visualization, investigation. Xiaoming Zhang, Guohua Yan, and Yu Zhou: Visualization; writing review and editing, project administration, investigation. Kaichun Zhang, Xuwei Duan, Jing Wang, and Chuanbao Wu: Visualization; writing review and editing, project administration, investigation. This work was supported by National Natural Science Foundation of China (Grant nos. 32202424; 32372664), Financial Special Foundation (Grant no. KJCX20240408), Technological Innovation Ability project (Grant nos. KJCX20230805; KJCX20240326; KJCX20240403), International science and technology cooperation platform construction project (Grant no. 2024-11), Innovation Platform Construction project (Grant no. PT2024-10), and Promotion and Innovation project (Grant nos. KJCX20200114; KJCX201910) from Beijing Academy of Agriculture and Forestry Sciences. The authors declare no conflict of interest. No animals or humans were involved in this study. All the data for this project is in the CNGBdb under BioProject accession: https://db.cngb.org/search/project/CNP0004619/ and http://www.cherries.org.cn. Supplementary materials (methods, figures, tables, graphical abstract, slides, videos, Chinese translated version and update materials) may be found in the online DOI or iMeta Science http://www.imeta.science/imetaomics/. Figure S1: Flow cytometry identification of induced tetraploid sweet cherry. Figure S2: Telomeres detection map. Figure S3: Hi-C heatmap of chromosome interactions. Figure S4: The centromeres detection map. Figure S5: Density map of LTR/Copis, LTR/Gypsy, and protein coding genes along chromosomes. Figure S6: The KEGG analysis of the expanded genes families. Figure S7: The KEGG analysis of the contracted genes families. Figure S8: Synonymous substitutions per site (Ks). Figure S9: Fourfold synonymous third-codon transversion rate (4DTv) distribution. Figure S10: Genes families distribution and unique genes families in the 15 species. Figure S11: The KEGG analysis of the unique genes families. Figure S12: Collinearity diagram including the 10 Prunus species. Figure S13: Genes expression MA map. Figure S14: The KEGG analysis of the differential genes. Figure S15: The time series (MFUZZ) analysis of the genes in the T2X samples. Figure S16: The time series (MFUZZ) analysis of the genes in the T4X samples. Figure S17: The WGCNA analysis of the genes. Figure S18: The analysis of the modules in the WGCNA analysis results. Figure S19: The KEGG analysis of the biseque4 genes in the WGCNA analysis results. Figure S20: The KEGG pathways analysis of the flavonoid DEGs. Figure S21: The KEGG pathways analysis of the anthocyanin DEGs. Table S1: Statistics of sequencing data for sweet cherry (Prunusaviumcv. Tieton). Table S2: Quality statistics of genome assembly and annotation. Table S3: Statistics of the TE repetitive sequences annotated in sweet cherry (Prunusavium cv. Tieton) T2T genome. Table S4: Statistics of the intact TE repetitive sequences and genes annotated in chromosome. Table S5: Statistics of the functional annotated genes in the genome. Table S6: Statistics of the telomeres and centromeres sequences in the genome. Table S7: Statistics of the mutations types in the T4X genome. Table S8: Statistics of the mutations that affecting the flavonoid and anthocyanidin genes in the T4X. Table S9: Statistics of the mutations influenced transcription factors that co-expressed with the flavonoid and anthocyanidin genes in the T4X. Please note: The publisher is not responsible for the content or functionality of any supporting information supplied by the authors. Any queries (other than missing content) should be directed to the corresponding author for the article.
The nondestructive method for evaluating soluble solid content(SSC)of intact peach using diffuse near infrared reflectance was investigated.The prediction model was established by modified partial least squares(modified PLS)analysis with spectral and constituent measurement of calibration sample of 150 peach fruit.The results of validation with 20 peach fruit showed that the first derivative spectra with modified PLS provided the better predication of SSC of peach fruit with the bias of 0.381 between predicted and measured values,the standard error of prediction of 0.427 and the correlation coefficient of 0.701.It suggested that the diffuse near infrared reflectance technique be feasible for nondestructive detection of soluble solid content of peach fruit in the wave number range of 600~1848nm.
Summary Molecular stacking enables multiple traits to be effectively engineered in crops using a single vector. However, the co‐existence of distinct plant promoters in the same transgenic unit might, like their mammalian counterparts, interfere with one another. In this study, we devised a novel approach to investigate enhancer–promoter and promoter–promoter interactions in transgenic plants and demonstrated that three of four flower‐specific enhancer/promoters were capable of distantly activating a pollen‐ and stigma‐specific Pps promoter (fused to the cytotoxic DT‐A gene) in other tissues, as revealed by novel tissue ablation phenotypes in transgenic plants. The NtAGI1 enhancer exclusively activated stamen‐ and carpel‐specific DT‐A expression, thus resulting in tissue ablation in an orientation‐independent manner; this activation was completely abolished by the insertion of an enhancer‐blocking insulator ( EXOB ) between the NtAGI1 enhancer and Pps promoter. Similarly, AGL8 and AP1Lb1 , but not AP1La , promoters also activated distinct tissue‐specific DT‐A expression and ablation, with the former causing global growth retardation and the latter ablating apical inflorescences. While the tissue specificity of the enhancer/promoters generally defined their activation specificities, the strength of their activity in particular tissues or developmental stages appeared to determine whether activation actually occurred. Our findings provide the first evidence that plant‐derived enhancer/promoters can distantly interact/interfere with one another, which could pose potential problems for the tissue‐specific engineering of multiple traits using a single‐vector stacking approach. Therefore, our work highlights the importance of adopting enhancer‐blocking insulators in transformation vectors to minimize promoter–promoter interactions. The practical and fundamental significance of these findings will be discussed.
In order to discover weight loss of goose egg and hatching results during incubation,three breeds of geese were involved.The results showed that the average weight loss of Rhine Geese was higher than that of Zi Geese and Huo Geese(p0.05) in 28 days,but it was not significant beween ZiGeese and HuoGeese(p0.05).The Rhines fertilization rate was the highest(p0.05),the ZiGeeses fertilization hatchability rate was the highest(p0.05).Egg weight was positively correlated with that in 28th day.Percentage of weight loss decreased,while egg hatching weight increased.The bigger the egg weight was,the bigger the birth weight.
Recently,sweet cherry crinkle leaf disease seriously hurts production and qualities of sweet cherry.However,detailed disease observations and effective prevention and control measures have not been reported in china.Two years' successive and systematic investigations were caried on 248 adult cherry variety Hongdeng in a sweet cherry orchard near the Fragrant Hill,Beijing.We find that sweet cherry crinkle leaf disease mainly affects leaf shape,fruit development shape,fruit quality and production.Leaves become rough,narrow and with abnormal distorted edge.Fruits develop slowly with abnormal fruit and crinkled leaf being in the same branch.The crinkle leaf disease has made the fruits shorter,thinner and lighter.Their maturing date is postponed and yields reduced remarkably.Crinkle leaf disease can be found in several leaves,a branch or the whole tree.The incidence situation is different every year.However,the serious disease affected trees show stable performance.The best periods for observing crinkle leaf disease are from frondescing stage to pit-hardening stage in spring and in autumn,respectively.Grafting test was conducted in serious crinkle leaf disease affected trees,but no crinkle leaf disease is observed in scion.
<p><em>Eichhornia crassipes</em> is an aquatic plant native to the Amazon River Basin. It has become a serious weed in freshwater habitats in rivers, lakes and reservoirs both in tropical and warm temperate areas worldwide. Some research has stated that it can be used for water phytoremediation, due to its strong assimilation of nitrogen and phosphorus, and the accumulation of heavy metals, and its growth and spread may play an important role in environmental ecology. In order to explore the molecular mechanism of <em>E. crassipes</em> to responses to nitrogen deficiency, we constructed forward and reversed subtracted cDNA libraries for <em>E. crassipes</em> roots under nitrogen deficient condition using a suppressive subtractive hybridization (SSH) method. The forward subtraction included 2 100 clones, and the reversed included 2 650 clones. One thousand clones were randomly selected from each library for sequencing. About 737 (527 unigenes) clones from the forward library and 757 (483 unigenes) clones from the reversed library were informative. Sequence BlastX analysis showed that there were more transporters and adenosylhomocysteinase-like proteins in <em>E. crassipes</em> cultured in nitrogen deficient medium; while, those cultured in nitrogen replete medium had more proteins such as UBR4-like e3 ubiquitin-protein ligase and fasciclin-like arabinogalactan protein 8-like, as well as more cytoskeletal proteins, including actin and tubulin. Cluster of Orthologous Group (COG) analysis also demonstrated that in the forward library, the most ESTs were involved in coenzyme transportation and metabolism. In the reversed library, cytoskeletal ESTs were the most abundant. Gene Ontology (GO) analysis categories demonstrated that unigenes involved in binding, cellular process and electron carrier were the most differentially expressed unigenes between the forward and reversed libraries. All these results suggest that <em>E. crassipes</em> can respond to different nitrogen status by efficiently regulating and controlling some transporter gene expressions, certain metabolism processes, specific signal transduction pathways and cytoskeletal construction. </p>
Recent studies have shown that loss of pollen-S function in S4' pollen from sweet cherry (Prunus avium) is associated with a mutation in an S haplotype-specific F-box4 (SFB4) gene. However, how this mutation leads to self-compatibility is unclear. Here, we examined this mechanism by analyzing several self-compatible sweet cherry varieties. We determined that mutated SFB4 (SFB4') in S4' pollen (pollen harboring the SFB4' gene) is approximately 6 kD shorter than wild-type SFB4 due to a premature termination caused by a four-nucleotide deletion. SFB4' did not interact with S-RNase. However, a protein in S4' pollen ubiquitinated S-RNase, resulting in its degradation via the 26S proteasome pathway, indicating that factors in S4' pollen other than SFB4 participate in S-RNase recognition and degradation. To identify these factors, we used S4-RNase as a bait to screen S4' pollen proteins. Our screen identified the protein encoded by S4-SLFL2, a low-polymorphic gene that is closely linked to the S-locus. Further investigations indicate that SLFL2 ubiquitinates S-RNase, leading to its degradation. Subcellular localization analysis showed that SFB4 is primarily localized to the pollen tube tip, whereas SLFL2 is not. When S4-SLFL2 expression was suppressed by antisense oligonucleotide treatment in wild-type pollen tubes, pollen still had the capacity to ubiquitinate S-RNase; however, this ubiquitin-labeled S-RNase was not degraded via the 26S proteasome pathway, suggesting that SFB4 does not participate in the degradation of S-RNase. When SFB4 loses its function, S4-SLFL2 might mediate the ubiquitination and degradation of S-RNase, which is consistent with the self-compatibility of S4' pollen.
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