Summary: Kiwifruit (Actinidia spp) is a woody, perennial and deciduous vine. In this genus, there are multiple ploidy levels but the main cultivated cultivars are polyploid. Despite the availability of many genomic resources in kiwifruit, SNP genotyping is still a challenge given these different levels of polyploidy. Recent advances in SNP array technologies have offered a high‐throughput genotyping platform for genome‐wide DNA polymorphisms. In this study, we developed a high‐density SNP genotyping array to facilitate genetic studies and breeding applications in kiwifruit. SNP discovery was performed by genome‐wide DNA sequencing of 40 kiwifruit genotypes. The identified SNPs were stringently filtered for sequence quality, predicted conversion performance and distribution over the available Actinidia chinensis genome. A total of 134 729 unique SNPs were put on the array. The array was evaluated by genotyping 400 kiwifruit individuals. We performed a multidimensional scaling analysis to assess the diversity of kiwifruit germplasm, showing that the array was effective to distinguish kiwifruit accessions. Using a tetraploid F1 population, we constructed an integrated linkage map covering 3060.9 cM across 29 linkage groups and performed QTL analysis for the sex locus that has been identified on Linkage Group 3 (LG3) in Actinidia arguta. Finally, our dataset presented evidence of tetrasomic inheritance with partial preferential pairing in A. arguta. In conclusion, we developed and evaluated a 135K SNP genotyping array for kiwifruit. It has the advantage of a comprehensive design that can be an effective tool in genetic studies and breeding applications in this high‐value crop.

Low temperature causes injuries to plants during winter, thereby it affects kiwi fruit quality and yield. However, the changes in metabolites and gene expression during cold acclimation (CA) and deacclimation (DA) in kiwi fruit remain largely unknown. In this study, freezing tolerance, carbohydrate metabolism, and β-amylase gene expression in two Actinidia arguta cv. “CJ-1” and “RB-3” were detected from CA to DA stages. In all acclimation stages, the “CJ-1” was hardier than “RB-3” and possessed lower semi-lethal temperature (LT50). Furthermore, “CJ-1” had a more rapid acclimation speed than “RB-3”. Changes of starch, β-amylase, and soluble sugars were associated with freezing tolerance in both cultivars. Starch contents continued to follow a declining trend, while soluble sugars contents continuously accumulated in both cultivars during CA stages (from October to January). To investigate the possible molecular mechanism underlying cold response in A. arguta, in total, 16 AcBAMs genes for β-amylase were identified in the kiwi fruit genome. We carried out localization of chromosome, gene structure, the conserved motif, and the analysis of events in the duplication of genes from AcBAMs. Finally, a strong candidate gene named AaBAM3 from AcBAMs was cloned in Actinidia arguta (A. arguta), The real-time qPCR showed that AaBAM3 gene expression in seasonal changes was consistent with changes of soluble sugars. These results reveal that AaBAM3 may enhance the freezing tolerance of A. arguta through increasing soluble sugar content.

Kiwifruit coloration is an important agronomic trait used to determine fruit quality, and light plays a vital role in the coloration process. The effect of light on fruit coloration has been studied in many species, but differences in the photoresponse of different fruit parts during fruit coloration is unclear in kiwifruit (Actinidia arguta). In this study, peel and core with bagging and non-bagging treatment at two stages were selected to perform high throughput RNA sequencing. A total of 100,417 unigenes (25,186 unigenes with length beyond 1000 bp) were obtained, of which 37,519 unigenes were annotated in functional databases. GO and KEGG enrichment results showed that ‘plant hormone signal transduction’ and ‘carbon metabolism’ were the key pathways in peel and core coloration, respectively. A total of 27 MYB-related TFs (transcription factors) were differentially expressed in peel and core. An R2R3-MYB typed TF, AaMYB308like, possibly served as a candidate objective, which played a vital role in light-inducible fruit coloration based on bioinformatics analysis. Transient overexpression of AaMYB308like suggested overexpression of AaMYB308like elevated transcription level of NtCHI in Nicotiana tabacum leaves. Integration of all these results imply that AaMYB308like might be served as a light-responsive transcription factor to regulate anthocyanin biosynthesis in A. arguta. Moreover, our study provided important insights into photoreponse mechanisms in A. arguta coloration.

The C-repeat binding factors (CBFs) gene is essential for plants’ cold response, which could not only be induced by the inducer of CBF expression (ICE) genes but also activated the expression of the cold-regulated (COR) gene, thereby participating in the ICE-CBF-COR cold response pathway. However, this gene family and its functions in Actinidia arguta remain unclear. In this study, whole-genome identification and functional analysis of CBF family members in A. arguta were performed. Eighteen CBF genes, which were located on four chromosomes and had five tandem repeats, were identified. The proteins encoded by the genes were predicted to be located in the nucleus and cytoplasm. The results of the promoter cis-acting element analysis revealed light response elements, low-temperature response elements, and hormone (methyl jasmonate, gibberellin, salicylic acid, etc.) response elements. We analyzed collinearity with other kiwifruit genomes, and, interestingly, the number of CBF family members differed across geographic locations of A. arguta. RT-qPCR revealed that the expression of the CBF gene family differed under low-temperature treatment; specifically, we observed differences in the expression of all the genes. Based on phylogenetic relationships and RT-qPCR analysis, the expression of AaCBF4.1 (AaCBF4) was found to be highly upregulated, and the function of this gene in cold resistance was further verified via overexpression in transgenic Arabidopsis. AaCBF4-overexpressing plants showed higher tolerance to cold stress, showing a higher germination rate, higher chlorophyll content and lower relative electrolyte leakage. In addition, compared with the wild-type Arabidopsis, the overexpressing plants exhibited significantly reduced oxidative damage due to the reduction in reactive oxygen species production under cold stress. Therefore, AaCBF4 plays an important role in improving the cold resistance of Actinidia arguta and can be further used to develop kiwifruit germplasm resources with strong cold resistance.

Kiwifruit is a dioecious woody liana fruit tree, and the non-fruitfulness of male plants leads to a great deal of blindness in the selection of male plants in crossbreeding. In this study, we induced the development of male plant ovary by externally applying plant growth regulators (PGRs) and performed histological observation, phytohormone content determination and transcriptome analysis on the abortive ovary of the male kiwifruit (Con), the ovary of the female kiwifruit (Fem) and the PGR-induced developing ovary of the male kiwifruit (PT). Histological analysis showed that the Con ovary was devoid of ovules and the carpels were atrophied, the Fem ovary had ovules and the PT ovary was devoid of ovules, but the carpels developed normally and were not atrophied. Endogenous phytohormone content measurements displayed higher levels of trans-zeatin (tZT) in PT and Fem than Con, and lower levels of gibberellin (GA3) and abscisic acid (ABA) than Con. Transcriptome analysis revealed significant differences in many key genes in the cytokinin and auxin pathways, which were consistent with the results of phytohormone content measurements. Meanwhile, the genes related to carpel development, SPT (DTZ79_04g03580) and SK41 (DTZ79_19g04340), were highly expressed in PT, suggesting that they may play a key role in PGR-induced development of the ovary in male kiwifruit. These results provide information for elucidating the potential regulatory network of PGR-induced ovary development in male flowers and contribute to further identification of valuable target genes.

BackgroundNDR1/HIN1-like (NHL) genes play crucial roles in Psa resistance. Kiwifruit canker, caused by Pseudomonas syringae pv. Actinidiae (Psa) infection is one of the most serious diseases affecting the kiwifruit industry. However, the key NHL has not yet been identified in kiwifruit.ResultsIn this study, we conducted a genome-wide identification of NHL family in kiwifruit (Actinidia eriantha). A total of 33 AeNHLs were divided into five domain-conserved subfamilies, which were mainly assigned into phytohormones and defense responses. The expression of AeNHL genes was analyzed to identify key genes in response to Psa, and we found AeNHL17 was highly expressed upon Psa inoculation. Transgenic tobacco overexpressing AeNHL17 presented higher resistance to Psa than wild-type (WT) tobacco, implying a key role for AeNHL17 in Psa resistance. Finally, we carried out a stable genetic transformation of kiwifruit (A. chinensis), which is sensitive to Psa, and found that the overexpression of AeNHL17 increased resistance to infection. AeNHL17-silenced plants exhibited larger disease lesions than control plants.ConclusionsOur findings revealed the function of AaNHL17 in Psa resistance, providing new data regarding the functional analysis of the NHL gene family in kiwifruit.