Cytoplasmic male sterility (CMS) is important for exploiting heterosis in crop plants and also serves as a model for investigating nuclear–cytoplasmic interaction. The molecular mechanism of cytoplasmic male sterility and fertility restoration was investigated in several important economic crops but remains poorly understood in the Welsh onion. Therefore, we compared the differences between the CMS line 64-2 and its maintainer line 64-1 using transcriptome sequencing with the aim of determining critical genes and pathways associated with male sterility. This study combined two years of RNA-seq data; there were 1504 unigenes (in May 2013) and 2928 unigenes (in May 2014) that were differentially expressed between the CMS and cytoplasmic male maintainer Welsh onion varieties. Known CMS-related genes were found in the set of differentially expressed genes and checked by qPCR. These genes included F-type ATPase, NADH dehydrogenase, cytochrome c oxidase, etc. Overall, this study demonstrated that the CMS regulatory genes and pathways may be associated with the mitochondria and nucleus in the Welsh onion. We believe that this transcriptome dataset will accelerate the research on CMS gene clones and other functional genomics research on A. fistulosum L.

Chinese chestnut (Castanea mollissima Blume) is one of the earliest domesticated plants of high nutritional and ecological value, yet mechanisms of C. mollissima underlying its growth and development are poorly understood. Although individual chestnut species differ greatly, the molecular basis of the formation of their characteristic traits remains unknown. Though the draft genomes of chestnut have been previously released, the pan-genome of different variety needs to be studied. We report the genome sequence of three cultivated varieties of chestnut herein, namely Hei-Shan-Zhai-7 (H7, drought-resistant variety), Yan-Hong (YH, easy-pruning variety), and Yan-Shan-Zao-Sheng (ZS, early-maturing variety), to expedite convenience and efficiency in its genetics-based breeding. We obtained three chromosome-level chestnut genome assemblies through a combination of Oxford Nanopore technology, Illumina HiSeq X, and Hi-C mapping. The final genome assemblies are 671.99 Mb (YH), 790.99 Mb (ZS), and 678.90 Mb (H7), across 12 chromosomes, with scaffold N50 sizes of 50.50 Mb (YH), 65.05 Mb (ZS), and 52.16 Mb (H7). Through the identification of homologous genes and the cluster analysis of gene families, we found that H7, YH and ZS had 159, 131, and 91 unique gene families, respectively, and there were 13,248 single-copy direct homologous genes in the three chestnut varieties. For the convenience of research, the chestnut genome database¹ was constructed. Based on the results of gene family identification, the presence/absence variations (PAVs) information of the three sample genes was calculated, and a total of 2,364, 2,232, and 1,475 unique genes were identified in H7, YH and ZS, respectively. Our results suggest that the GBSS II-b gene family underwent expansion in chestnut (relative to nearest source species). Overall, we developed high-quality and well-annotated genome sequences of three C. mollissima varieties, which will facilitate clarifying the molecular mechanisms underlying important traits, and shortening the breeding process.

Coryloideae is a subfamily in the family Betulaceae consisting of four extant genera: Carpinus, Corylus, Ostrya, and Ostryopsis. We sequenced the plastomes of six species of Corylus and one species of Ostryopsis for comparative and phylogenetic analyses. The plastomes are 159–160 kb long and possess typical quadripartite cp architecture. The plastomes show moderate divergence and conserved arrangement. Five mutational hotspots were identified by comparing the plastomes of seven species of Coryloideae: trnG-atpA, trnF-ndhJ, accD-psaI, ndhF-ccsA, and ycf1. We assembled the most complete phylogenomic tree for the family Betulaceae using 68 plastomes. Our cp genomic sequence phylogenetic analyses placed Carpinus, Ostrya, and Ostryopsis in a clade together and left Corylus in a separate clade. Within the genus Corylus, these analyses indicate the existence of five subclades reflecting the phylogeographical relationships among the species. The data offer significant genetic information for the identification of species of the Coryloideae, taxonomic and phylogenetic studies, and molecular breeding.

Methyl-CpG-binding domain (MBD) proteins have diverse molecular and biological functions in plants. Most studies of MBD proteins in plants have focused on the model plant Arabidopsis thaliana L. Here we cloned SvMBD5 from the willow Salix viminalis L. by reverse transcription-polymerase chain reaction (RT-PCR) and analyzed the structure of SvMBD5 and its evolutionary relationships with proteins in other species. The coding sequence of SvMBD5 is 645 bp long, encoding a 214 amino acid protein with a methyl-CpG-binding domain. SvMBD5 belongs to the same subfamily as AtMBD5 and AtMBD6 from Arabidopsis. Subcellular localization analysis showed that SvMBD5 is only expressed in the nucleus. We transformed Arabidopsis plants with a 35S::SvMBD5 expression construct to examine SvMBD5 function. The Arabidopsis SvMBD5-expressing line flowered earlier than the wild type. In the transgenic plants, the expression of FLOWERING LOCUS T and CONSTANS significantly increased, while the expression of FLOWERING LOCUS C greatly decreased. In addition, heterologously expressing SvMBD5 in Arabidopsis significantly inhibited the establishment and maintenance of methylation of CHROMOMETHYLASE 3 and METHYLTRANSFERASE 1, as well as their expression, and significantly increased the expression of the demethylation-related genes REPRESSOR OF SILENCING1 and DEMETER-LIKE PROTEIN3. Our findings suggest that SvMBD5 participates in the flowering process by regulating the methylation levels of flowering genes, laying the foundation for further studying the role of SvMBD5 in regulating DNA demethylation.

Chinese chestnut (Castanea mollissima) is an economically and ecologically important woody nut crop. In C. mollissima, flowering is fundamental for nut yield. The MADS-box gene APETALA 1 (AP1) plays essential roles in floral initiation and floral organ development in many plants. However, the function of the AP1 gene CmAP1 in C. mollissima is still unclear. Here, we cloned the coding sequence (CDS) and promoter of CmAP1 and analyzed the function of this gene. The CDS of CmAP1 is 741 bp and encodes a 246–amino acid protein. Subcellular analysis revealed that CmAP1 localizes to the nucleus. GUS driven by the CmAP1 promoter was expressed in seedlings and in leaf margins, petals, and carpels of transgenic Arabidopsis (Arabidopsis thaliana). RNA in situ hybridization indicated that CmAP1 was mainly expressed in the inflorescence meristem, floral primordia, sepal primordia, petal primordia, stamen primordia, and carpel primordia during the early stage of flower development. An auxin response element (TGA element), jasmonic acid response element (TGACG motif), and WRKY binding site (W-box element) were identified in the CmAP1 promoter. Heterologous expression of CmAP1 in wild-type and ap1-11 Arabidopsis resulted in early flowering. Notably, the expression of CmAP1 rescued the loss of the petal whorl in the ap1-11 mutant. AtAP1, AtSEP1, AtSEP2, AtSEP3, and AtSEP4 were upregulated in CmAP1-expressing Arabidopsis plants. These findings suggest that CmAP1 promotes flowering and plays a key role in petal development. Our findings help reveal the regulatory mechanism of flowering and flower development in C. mollissima, providing a practical basis for increasing yield in Chinese chestnut.