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Summary Fruit ripening and quality are common botanical phenomena that are closely linked and strictly regulated by transcription factors. It was previously discovered that a banana MADS‐box protein named MuMADS1 interacted with an ovate family protein named MaOFP1 to regulate banana fruit ripening. To further investigate the role of MuMADS1 and MaOFP1 in the regulation of fruit quality, a combination of genetic transformation and transcriptional characterization was used. The results indicated that the co‐expression of MuMADS1 and MaOFP1 in the ovate mutant could compensate for fruit shape and inferior qualities relating to fruit firmness, soluble solids and sugar content. The number of differentially expressed genes (DEGs) was 1395 in WT vs. ovate, with 883 up‐regulated and 512 down‐regulated genes, while the numbers of DEGs gradually decreased with the transformation of MuMADS1 and MaOFP1 into ovate. ‘Starch and sucrose metabolism’ constituted the primary metabolic pathway, and the gene numbers in this pathway were obviously different when MuMADS1 and MaOFP1 were integrated into ovate. A series of metabolic genes involved in cell wall biosynthesis were up‐regulated in the WT vs. ovate, which probably resulted in the firmer texture and lower sugar contents in the ovate fruit. These results demonstrate that MuMADS1 and MaOFP1 are coregulators of fruit quality, facilitating the dissection of the molecular mechanisms underlying fruit quality formation.

Background Abscisic acid (ABA) signaling plays a crucial role in developmental and environmental adaptation processes of plants. However, the PYL-PP2C-SnRK2 families that function as the core components of ABA signaling are not well understood in banana. Results In the present study, 24 PYL, 87 PP2C, and 11 SnRK2 genes were identified from banana, which was further supported by evolutionary relationships, conserved motif and gene structure analyses. The comprehensive transcriptomic analyses showed that banana PYL-PP2C-SnRK2 genes are involved in tissue development, fruit development and ripening, and response to abiotic stress in two cultivated varieties. Moreover, comparative expression analyses of PYL-PP2C-SnRK2 genes between BaXi Jiao (BX) and Fen Jiao (FJ) revealed that PYL-PP2C-SnRK2-mediated ABA signaling might positively regulate banana fruit ripening and tolerance to cold, salt, and osmotic stresses. Finally, interaction networks and co-expression assays demonstrated that the core components of ABA signaling were more active in FJ than in BX in response to abiotic stress, further supporting the crucial role of the genes in tolerance to abiotic stress in banana. Conclusions This study provides new insights into the complicated transcriptional control of PYL-PP2C-SnRK2 genes, improves the understanding of PYL-PP2C-SnRK2-mediated ABA signaling in the regulation of fruit development, ripening, and response to abiotic stress, and identifies some candidate genes for genetic improvement of banana. Electronic supplementary material The online version of this article (doi:10.1186/s12870-017-1093-4) contains supplementary material, which is available to authorized users.

MicroRNAs (miRNAs) are a class of endogenous non-coding small RNAs that regulate targeted mRNAs by post-transcription; thus, miRNAs are considered post-transcription regulators. Although numerous miRNAs have been found in model plants, information on miRNAs in banana is limited. In this study, 139 members in 38 miRNA families were determined by sequencing the small RNA (sRNA) transcriptomes of Fusarium wilt-resistant and susceptible banana varieties, and six out of eight new miRNAs were confirmed by RT-PCR. According to the sRNA transcriptome data and qRT-PCR verification, some miRNAs were differentially expressed between Fusarium wilt-resistant and susceptible banana varieties. A total of 293 and 31 target genes were predicted based on the draft maps of banana A genome and Fusarium oxysporum (FOC1, FOC4) genomes, respectively. Two important pathogenic genes in F. oxysporum genomes, namely, feruloyl esterase gene and proline iminopeptidase gene, were targeted by banana miRNAs. These novel findings may provide a new strategy for preventing and controlling Fusarium wilt in banana.

Plant translation elongation factor 1 alpha (EF1A) is both a protein synthesis factor and an important component of plant signal transduction, immune responses, protein trafficking, and apoptosis. However, its role in plant growth and development remains unclear. Herein, a full-length EF1A gene was isolated from banana (Musa acuminata L.) fruit and termed MaEF1A. We found that MaEF1A shared a high sequence identify with respective genes in other plants and the deduced amino acid sequence contained conserved regions of GTP-EFTU, GTP-EFTU-02, and GTP-EFTU-03, as well as two tRNA binding domains and six GTP-binding sites which represent functional domains for protein biosynthesis. MaEF1A protein is mainly localized to the nucleus. MaEF1A was constitutively expressed in different banana organs including developing fruits, and the highest expression was detected in ovary 4 stage. Arabidopsis thaliana L. (ecotype Columbia) was transformed with MaEF1A and four transgenic lines were obtained. Three transgenic lines were selected for further phenotypic analyses. Our findings indicate that overexpressed MaEF1A could greatly enhance plant height, root length, and both rhachis and silique length by promoting cell expansion and elongation. These experiments suggest an important role for MaEF1A in plant growth and development.

The calcium-dependent protein kinase (CDPK) family is comprised of essential calcium sensors and responders that play important roles in response to various abiotic stresses in plants. In this study we describe the molecular characteristics of eight CDPK genes (MaCDPK1, MaCDPK2, MaCDPK3, MaCDPK4, MaCDPK5, MaCDPK6, MaCDPK7, and MaCDPK8) cloned from banana (Musa acuminate L. AAA group, cv. Cavendish) using a RACE-PCR-based strategy. The predicted molecular masses of these MaCDPKs ranged from 57.78 to 60.46 kDa and their pIs ranged from 5.89 to 8.97. At the amino acid level, they shared high sequence similarity with CDPKs in the banana DH-Pahang (AA group) genome. Phylogenetic analysis showed that the deduced amino acid sequences of MaCDPKs also had high similarity to CPKs of Arabidopsis. Expression analysis by semi-quantitative RT-PCR revealed that these genes were differently expressed in various tissues. In addition, their expression was regulated by various stress conditions, including exposure to signaling molecules, cold, salinity, drought, and Fusarium oxysporum f specialis(f. Sp) cubense Tropical Race 4 (Foc TR4) infection. MaCDPK3 responded to abiotic stresses, such as drought, cold, and salinity, while MaCDPK2 and MaCDPK4 were sensitive to Foc TR4. Our results indicate that MaCDPKs play important roles in both the development and stress responses of banana plantlets.

Starch branching enzyme (SBE), which is one of the key enzymes associated with amylopectin biosynthesis, plays important roles in variable biological processes. Despite its importance, SBE is rarely studied in the banana (Musa acuminata L.) which is a typical starchy fruit. Here, a family of ten SBE proteins (MaSBE) was firstly identified through genome-wide characterization in M. acuminata, which could be clustered into three subfamilies. Systematic transcriptome analysis revealed temporal and spatial expression variations of MaSBE genes and differential response patterns under abiotic and biotic stresses in both banana genotypes, Fen Jiao (FJ) and BaXi Jiao (BX). Moreover, MaSBE2.4 was temporally regulated during fruit development and ripening as well as in response to various abiotic/biotic stresses in both genotypes. Specifically, MaSBE2.3 expression level was higher in FJ than in BX following cold, salt, and drought stress treatments, and it was specifically induced by fungal infection in BX. Characterization of hormone- and stress-related cis-acting elements in the promoters of MaSBE genes suggests their multiple biological functions. In conclusion, our study provides new insights into the complex transcriptional characteristics of the SBE genes, and demonstrates their crucial roles in improving amylopectin biosynthesis and strengthening stress resistance in banana.

The ovate mutation has frequently been used to study changes in fruit shape but not fruit quality. A deterioration in fruit quality associated with the ovate mutation was discovered in this study. To elucidate how ovate influences the quality of fruit, we performed a proteomics analysis of the fruits of the ovate mutant (LA3543) and wild-type (“Ailsa Craig”, LA2838A) using tandem mass tag analysis. The results indicated that the ovate mutation significantly influences fruit quality in a number of ways, including by reducing the expression of 1-aminocyclopropane-1-carboxylic acid oxidase 3 (ACO3) in ethylene biosynthesis, improving firmness by reducing the amount of pectinesterase and polygalacturonase, reducing sugar accumulation by downregulating the abundance of mannan endo-1,4-β-mannosidase 4, β-galactosidase, and β-amylase, and reducing the malic acid content by downregulating the accumulation of malic enzymes and malate synthase. These findings could inform future improvements in fruit quality.

Proteins encoded by MADS-box genes are important transcription factors involved in the regulation of flowering plant growth and development. Currently, no systematic information exists regarding the MADS-box family in the important tropical fruit banana. Ninety-six MADS-box genes were identified from the banana (Pahang) A genome. Phylogenetic analysis indicated that Musa acuminata MCM1-AGAMOUS- DEFICIENS-SRF (MaMADS) could be divided into MIKCc, MIKC*, Mα/β and Mγ groups. MIKCc could be further divided into 11 subfamilies, which was further supported by conserved motif and gene structure analyses. Transcriptome analysis on the Feng Jiao (FJ) and BaXi Jiao (BX) banana cultivars revealed that MaMADS genes are differentially expressed in various organs, at different fruit development and ripening stages, indicating the involvement of these genes in fruit development and ripening processes. Interactive network analysis indicated that MaMADS24 and 49 not only interacted with MaMADS proteins themselves, but also interacted with hormone-response proteins, ethylene signal transduction and biosynthesis-related proteins, starch biosynthesis proteins and metabolism-related proteins. This systematic analysis identified candidate MaMADS genes related to fruit development and ripening for further functional characterization in plants, and also provided new insights into the transcriptional regulation of MaMADS genes, facilitating the future genetic manipulation of MADS-mediated fruit development and ripening.

Sugars Will Eventually be Exported Transporters (SWEET) are a novel type of sugar transporter that plays crucial roles in multiple biological processes. From banana, for the first time, 25 SWEET genes which could be classified into four subfamilies were identified. Majority of MaSWEETs in each subfamily shared similar gene structures and conserved motifs. Comprehensive transcriptomic analysis of two banana genotypes revealed differential expression patterns of MaSWEETs in different tissues, at various stages of fruit development and ripening, and in response to abiotic and biotic stresses. More than 80% MaSWEETs were highly expressed in BaXi Jiao (BX, Musa acuminata AAA group, cv. Cavendish), in sharp contrast to Fen Jiao (FJ, M. acuminata AAB group) when pseudostem was first emerged. However, MaSWEETs in FJ showed elevated expression under cold, drought, salt, and fungal disease stresses, but not in BX. Interaction networks and co-expression assays further revealed that MaSWEET-mediated networks participate in fruit development signaling and abiotic/biotic stresses, which was strongly activated during early stage of fruit development in BX. This study provides new insights into the complex transcriptional regulation of SWEETs, as well as numerous candidate genes that promote early sugar transport to improve fruit quality and enhance stress resistance in banana.

Three tau class MaGSTs responded to abiotic stress, MaGSTF1 and MaGSTL1 responded to signaling molecules, they may play an important role in the growth of banana plantlet. Glutathione S-transferases (GST) are multifunctional detoxification enzymes that participate in a variety of cellular processes, including stress responses. In this study, we report the molecular characteristics of five GST genes (MaGSTU1, MaGSTU2, MaGSTU3, MaGSTF1 and MaGSTL1) cloned from banana (Musa acuminate L. AAA group, cv. Cavendish) using a RACE-PCR-based strategy. The predicted molecular masses of these GSTs range from 23.4 to 27.7 kDa and their pIs are acidic. At the amino acid level, they share high sequence similarity with GSTs in the banana DH-Pahang (AA group) genome. Phylogenetic analysis showed that the deduced amino acid sequences of MaGSTs also have high similarity to GSTs of other plant species. Expression analysis by semi-quantitative RT-PCR revealed that these genes are differentially expressed in various tissues. In addition, their expression is regulated by various stress conditions, including exposure to signaling molecules, cold, salinity, drought and Fusarium oxysporum f specialis(f. Sp) cubense Tropical Race 4 (Foc TR4) infection. The expression of the tau class MaGSTs (MaGSTU1, MaGSTU2 and MaGSTU3) mainly responded to cold, salinity and drought while MaGSTF1 and MaGSTL1 expressions were upregulated by signaling molecules. Our findings suggest that MaGSTs play a key role in both development and abiotic stress responses.

Plant 14-3-3 proteins act as critical components of various cellular signaling processes and play an important role in regulating multiple physiological processes. However, less information is known about the 14-3-3 gene family in banana. In this study, 25 14-3-3 genes were identified from the banana genome. Based on the evolutionary analysis, banana 14-3-3 proteins were clustered into ε and non-ε groups. Conserved motif analysis showed that all identified banana 14-3-3 genes had the typical 14-3-3 motif. The gene structure of banana 14-3-3 genes showed distinct class-specific divergence between the ε group and the non-ε group. Most banana 14-3-3 genes showed strong transcript accumulation changes during fruit development and postharvest ripening in two banana varieties, indicating that they might be involved in regulating fruit development and ripening. Moreover, some 14-3-3 genes also showed great changes after osmotic, cold, and salt treatments in two banana varieties, suggested their potential role in regulating banana response to abiotic stress. Taken together, this systemic analysis reveals the involvement of banana 14-3-3 genes in fruit development, postharvest ripening, and response to abiotic stress and provides useful information for understanding the functions of 14-3-3 genes in banana.

The leucine zipper (bZIP) transcription factors play important roles in multiple biological processes. However, less information is available regarding the bZIP family in the important fruit crop banana. In this study, 121 bZIP transcription factor genes were identified in the banana genome. Phylogenetic analysis showed that MabZIPs were classified into 11 subfamilies. The majority of MabZIP genes in the same subfamily shared similar gene structures and conserved motifs. The comprehensive transcriptome analysis of two banana genotypes revealed the differential expression patterns of MabZIP genes in different organs, in various stages of fruit development and ripening and in responses to abiotic stresses, including drought, cold and salt. Interaction networks and co-expression assays showed that group A MabZIP-mediated networks participated in various stress signaling, which was strongly activated in Musa ABB Pisang Awak. This study provided new insights into the complicated transcriptional control of MabZIP genes and provided robust tissue-specific, development-dependent and abiotic stress-responsive candidate MabZIP genes for potential applications in the genetic improvement of banana cultivars.

The abscisic acid (ABA)-, stress-, and ripening-induced (ASR) protein is a plant-specific hydrophilic transcriptional factor involved in fruit ripening and the abiotic stress response. To date, there have been no studies on the role of ASR genes in delayed flowering time. Here, we found that the ASR from banana, designated as MaASR, was preferentially expressed in the banana female flowers from the eighth, fourth, and first cluster of the inflorescence. MaASR transgenic lines (L14 and L38) had a clear delayed-flowering phenotype. The number of rosette leaves, sepals, and pedicel trichomes in L14 and L38 was greater than in the wild type (WT) under long day (LD) conditions. The period of buds, mid-flowers, and full bloom of L14 and L38 appeared later than the WT. cDNA microarray and quantitative real-time PCR (qRT-PCR) analyses revealed that overexpression of MaASR delays flowering through reduced expression of several genes, including photoperiod pathway genes, vernalization pathway genes, gibberellic acid pathway genes, and floral integrator genes, under short days (SD) for 28 d (from vegetative to reproductive transition stage); however, the expression of the autonomous pathway genes was not affected. This study provides the first evidence of a role for ASR genes in delayed flowering time in plants.

A full-length cDNA isolated from banana (Musa acuminata L. AAA group) fruit was named MuBTB, and it contains an open reading frame encoding 354 amino acids. Sequence analysis showed that MuBTB shares high similarity with broad complex, tramtrack, and bric-a-bracs from other plant species. Reverse transcriptase PCR analysis of MuBTB spatial expression showed that MuBTB was expressed in all organs examined: root, rhizome, leaf, flower, and fruit. Real-time quantitative PCR of postharvest banana revealed that MuBTB exhibited differential expression patterns that are associated with ethylene biosynthesis. In naturally ripened bananas, expression of MuBTB was in accordance with ethylene biosynthesis. In contrast, for 1-methylcyclopropene-treated bananas, MuBTB expression levels were inhibited and remained constant. Following treatment with ethylene, MuBTB expression in banana fruit significantly increased with ethylene biosynthesis and peaked 3 days after harvest, which was 11 days earlier than that for naturally ripened banana fruits. These results suggest that MuBTB expression is induced by ethylene for regulation of postharvest banana ripening. Finally, subcellular localization assays showed that the MuBTB protein localizes to the nucleus.

Efficient transformation of lilies is required for their genetic improvement in ornamental and marketable qualities. Although Lilium longiflorum can be transformed by particle bombardment and Agrobacterium, the transformation frequency is low. In this study, we tested new Agrobacterium-mediated transformation methods using shoot segments combined with two different regeneration systems. Shoot segments were co-cultivated for 2 d with Agrobacterium tumefaciens strain AGL1/pCAS04 harboring a binary vector carrying the neomycin phosphotransferase II driven by a promoter from the maize ubiquitin gene. The effect of different concentrations of 6-benzyladenine (BA) and 2,4-dichlorophenoxyacetic acid (2,4-D) on regeneration was investigated. The results indicated that Murashige and Skoog (MS) medium with 4.4 μM BA and 0.5 μM α-naphthalene acetic acid was optimal for shoot formation, and the nodal stem was the best explant for shoot induction. MS medium with 9.0 μM 2,4-D and 0.4 μM BA was optimal for callus induction. The direct shoot formation method regenerated 187 plantlets per 100 explants, and 74.4% of the regenerants were positive in transgene PCR. The callus regeneration method regenerated 20 plantlets per 100 explants, and 31.5% of them were PCR positive. Southern blotting confirmed the insertion of transgene in the plant host genome. The direct shoot formation method is more than 20-fold more efficient than previously reported transformation method in this species.

Auxin signaling regulates various auxin-responsive genes via two types of transcriptional regulators, Auxin Response Factors (ARF) and Aux/IAA. ARF transcription factors act as critical components of auxin signaling that play important roles in modulating various biological processes. However, limited information about this gene family in fruit crops is currently available. Herein, 47 ARF genes were identified in banana based on its genome sequence. Phylogenetic analysis of the ARFs from banana, rice, and Arabidopsis suggested that the ARFs could be divided into four subgroups, among which most ARFs from the banana showed a closer relationship with those from rice than those from Arabidopsis. Conserved motif analysis showed that all identified MaARFs had typical DNA-binding and ARF domains, but 12 members lacked the dimerization domain. Gene structure analysis showed that the number of exons in MaARF genes ranged from 5 to 21, suggesting large variation amongst banana ARF genes. The comprehensive expression profiles of MaARF genes yielded useful information about their involvement in diverse tissues, different stages of fruit development and ripening, and responses to abiotic stresses in different varieties. Interaction networks and co-expression assays indicated the strong transcriptional response of banana ARFs and ARF-mediated networks in early fruit development for different varieties. Our systematic analysis of MaARFs revealed robust tissue-specific, development-dependent, and abiotic stress-responsive candidate MaARF genes for further functional assays in planta. These findings could lead to potential applications in the genetic improvement of banana cultivars, and yield new insights into the complexity of the control of MaARF gene expression at the transcriptional level. Finally, they support the hypothesis that ARFs are a crucial component of the auxin signaling pathway, which regulates a wide range of physiological processes.

Aquaporins (AQPs) function to selectively control the flow of water and other small molecules through biological membranes, playing crucial roles in various biological processes. However, little information is available on the AQP gene family in bananas. In this study, we identified 47 banana AQP genes based on the banana genome sequence. Evolutionary analysis of AQPs from banana, Arabidopsis, poplar, and rice indicated that banana AQPs (MaAQPs) were clustered into four subfamilies. Conserved motif analysis showed that all banana AQPs contained the typical AQP-like or major intrinsic protein (MIP) domain. Gene structure analysis suggested the majority of MaAQPs had two to four introns with a highly specific number and length for each subfamily. Expression analysis of MaAQP genes during fruit development and postharvest ripening showed that some MaAQP genes exhibited high expression levels during these stages, indicating the involvement of MaAQP genes in banana fruit development and ripening. Additionally, some MaAQP genes showed strong induction after stress treatment and therefore, may represent potential candidates for improving banana resistance to abiotic stress. Taken together, this study identified some excellent tissue-specific, fruit development- and ripening-dependent, and abiotic stress-responsive candidate MaAQP genes, which could lay a solid foundation for genetic improvement of banana cultivars.

Mitogen-activated protein kinases (MAPKs) play important roles in signal transduction pathways responding to various stresses. In this study, we described the molecular characteristics of six MAPK genes (MaMAPK1, MaMAPK2, MaMAPK3, MaMAPK4, MaMAPK5 and MaMAPK6) cloned from banana (Musa acuminate L. AAA group, cv. Cavendish) using a RACE-PCR-based strategy. The predicted molecular masses of these MAPKs ranged from 43.0 to 70.1 kDa and their pIs ranged from 5.67 to 9.32. At the amino acid level, they shared high sequence similarity with MAPKs in the banana DH-Pahang (AA group) genome. Phylogenetic analysis showed that the deduced amino acid sequences of MaMAPKs also had high similarity with MAPKs of other plant species. These genes were expressed in nearly all tissues using semi-quantitative RT-PCR. All of the six MaMAPKs were induced by signal molecules in banana seedlings, such as ABA, Ethephon, MeJA and SA. The gene expressions of MaMAPK1, MaMAPK2, MaMAPK3, MaMAPK5 and MaMAPK6 were induced in resistant cultivar of banana after being inoculated with Fusarium Oxysporum f specialis (f. Sp) cubense Tropical Race 4 (Foc TR4). Our results suggest that MaMAPKs play a key role in both the development of banana seedlings and banana resistance to Foc TR4.

The ovate family protein named MaOFP1 was identified in banana (Musa acuminata L.AAA) fruit by a yeast two-hybrid (Y2H) method using the banana MADS-box gene MuMADS1 as bait and a 2 day postharvest (DPH) banana fruit cDNA library as prey. The interaction between MuMADS1 and MaOFP1 was further confirmed by Y2H and Bimolecular Fluorescence Complementation (BiFC) methods, which showed that the MuMADS1 K domain interacted with MaOFP1. Real-time quantitative PCR evaluation of MuMADS1 and MaOFP1 expression patterns in banana showed that they are highly expressed in 0 DPH fruit, but present in low levels in the stem, which suggests that simultaneous but different expression patterns exist for both MuMADS1 and MaOFP1 in different tissues and developing fruits. Meanwhile, MuMADS1 and MaOFP1 expression was highly stimulated and greatly suppressed, respectively, by exogenous ethylene. In contrast, MaOFP1 expression was highly stimulated while MuMADS1 was greatly suppressed by the ethylene competitor 1-methylcyclopropene (1-MCP). These results indicate that MuMADS1 and MaOFP1 are antagonistically regulated by ethylene and might play important roles in postharvest banana fruit ripening.

ADP-glucose pyrophosphorylase (AGPase) is the first rate-limiting enzyme in starch biosynthesis and plays crucial roles in multiple biological processes. Despite its importance, AGPase is poorly studied in starchy fruit crop banana (Musa acuminata L.). In this study, eight MaAGPase genes have been identified genome-wide in M. acuminata, which could be clustered into the large (APL) and small (APS) subunits. Comprehensive transcriptomic analysis revealed temporal and spatial expression variations of MaAPLs and MaAPSs and their differential responses to abiotic/biotic stresses in two banana genotypes, Fen Jiao (FJ) and BaXi Jiao (BX). MaAPS1 showed generally high expression at various developmental and ripening stages and in response to abiotic/biotic stresses in both genotypes. MaAPL-3 and -2a were specifically induced by abiotic stresses including cold, salt, and drought, as well as by fungal infection in FJ, but not in BX. The presence of hormone-related and stress-relevant cis-acting elements in the promoters of MaAGPase genes suggests that MaAGPases may play an important role in multiple biological processes. Taken together, this study provides new insights into the complex transcriptional regulation of AGPases, underlying their key roles in promoting starch biosynthesis and enhancing stress tolerance in banana.

Fusarium wilt caused by the fungus Fusarium oxysporum f. sp. cubens (Foc) is the most serious disease that attacks banana plants. Salicylic acid (SA) can play a key role in plant-microbe interactions. Our study is the first to examine the role of SA in conferring resistance to Foc TR4 in banana (Musa acuminata L. AAA group, cv. Cavendish), which is the greatest commercial importance cultivar in Musa. We used quantitative real-time reverse polymerase chain reaction (qRT-PCR) to analyze the expression profiles of 45 genes related to SA biosynthesis and downstream signaling pathways in a susceptible banana cultivar (cv. Cavendish) and a resistant banana cultivar (cv. Nongke No. 1) inoculated with Foc TR4. The expression of genes involved in SA biosynthesis and downstream signaling pathways was suppressed in a susceptible cultivar and activated in a resistant cultivar. The SA levels in each treatment arm were measured using high-performance liquid chromatography. SA levels were decreased in the susceptible cultivar and increased in the resistant cultivar. Finally, we examined the contribution of exogenous SA to Foc TR4 resistance in susceptible banana plants. The expression of genes involved in SA biosynthesis and signal transduction pathways as well as SA levels were significantly increased. The results suggest that one reason for banana susceptibility to Foc TR4 is that expression of genes involved in SA biosynthesis and SA levels are suppressed and that the induced resistance observed in banana against Foc TR4 might be a case of salicylic acid-dependent systemic acquired resistance.

Background Bananas and plantains (Musa spp.) are among the most important crops in the world due to their nutritional and export value. However, banana production has been devastated by fungal infestations caused by Fusarium oxysporum f. sp. cubense (Foc), which cannot be effectively prevented or controlled. Since there is very little known about the molecular mechanism of Foc infections; therefore, we aimed to investigate the transcriptional changes induced by Foc in banana roots. Results We generated a cDNA library from total RNA isolated from banana roots infected with Foc Tropical Race 4 (Foc TR 4) at days 0, 2, 4, and 6. We generated over 26 million high-quality reads from the cDNA library using deep sequencing and assembled 25,158 distinct gene sequences by de novo assembly and gap-filling. The average distinct gene sequence length was 1,439 base pairs. A total of 21,622 (85.94%) unique sequences were annotated and 11,611 were assigned to specific metabolic pathways using the Kyoto Encyclopedia of Genes and Genomes database. We used digital gene expression (DGE) profiling to investigate the transcriptional changes in the banana root upon Foc TR4 infection. The expression of genes in the Phenylalanine metabolism, phenylpropanoid biosynthesis and alpha-linolenic acid metabolism pathways was affected by Foc TR4 infection. Conclusion The combination of RNA-Seq and DGE analysis provides a powerful method for analyzing the banana root transcriptome and investigating the transcriptional changes during the response of banana genes to Foc TR4 infection. The assembled banana transcriptome provides an important resource for future investigations about the banana crop as well as the diseases that plague this valuable staple food.

Cold stress is an environmental factor affecting plant development and production. Recently, microRNAs (miRNAs) have been found to be involved in several plant processes such as growth regulation and stress responses. Although miRNAs and their targets have been identified in several banana species, their participation during cold accumulation in banana remains unknown. In this study, two small RNA libraries were generated from micropropagated plantlets of Musa balbisiana grown at normal and low temperature (5°C). A total of 69 known miRNAs and 32 putative novel miRNAs were detected in the libraries by Solexa sequencing. Sixty-four cold-inducible miRNAs were identified through differentially expressed miRNAs analysis. Among 43 miRNAs belonging to 26 conserved miRNA families with altered expression, 18 were upregulated and 25 downregulated under cold stress. Of 21 putative novel miRNAs with altered expression, four were downregulated and 17 upregulated. Furthermore, eight miRNAs were validated by stem-loop qRT-PCR and their dynamic differential expression was analyzed. In addition, 393 target genes of 58 identified cold-responsive miRNAs were predicted and categorized by function. These results provide important information for further characterization and functional analysis of cold-responsive miRNAs in banana.

β-amylase (BAM) is an important enzyme involved in conversion of starch to maltose in multiple biological processes in plants. However, there is currently insufficient information on the BAM gene family in the important fruit crop banana. This study identified 16 BAM genes in the banana genome. Phylogenetic analysis showed that MaBAMs were classified into four subfamilies. Most MaBAMs in each subfamily shared similar gene structures. Conserved motif analysis showed that all identified MaBAM proteins had the typical glyco hydro 14 domains. Comprehensive transcriptomic analysis of two banana genotypes revealed the expression patterns of MaBAMs in different tissues, at various stages of fruit development and ripening, and in responses to abiotic stresses. Most MaBAMs showed strong transcript accumulation changes during fruit development and late-stage ripening. Some MaBAMs showed significant changes under cold, salt, and osmotic stresses. This finding indicated that MaBAMs might be involved in regulating fruit development, ripening, and responses to abiotic stress. Analysis of five hormone-related and seven stress-relevant elements in the promoters of MaBAMs further revealed that BAMs participated in various biological processes. This systemic analysis provides new insights into the transcriptional characteristics of the BAM genes in banana and may serve as a basis for further functional studies of such genes.

The main function of endochitinase is believed to be pathogenesis related protein. However, more and more scientists reported the roles of endochitinase in plant growth and development. In order to investigate the role of endochitinase in postharvest banana fruit ripening, an endochitinase gene known as MaECHI1 had been isolated from a suppression subtractive hybridization (SSH) complementary deoxyribonucleic acid (cDNA) library. MaECHI1 was mainly expressed in banana fruit and flowers. Ethylene biosynthesis, gene expression and chitinase activities in different stages of postharvest banana fruit with or without ethylene and 1-methylcycle–propene (1-MCP) treatments were investigated. The results show that under ethylene treatment, banana ethylene production, gene expression, and chitinase activities increased markedly at the onset of banana ripening. Moreover, banana ethylene production and MaECHI1 gene expression peaks appeared earlier with ethylene treatment than with other treatment. MaECHI1 gene expression was markedly responsive to the fruit ripening process and to exogenous ethylene treatment. Keywords: Banana ( Musa acuminata L.AAA), endochitinase gene expression, ethylene production fruit ripening

An ascorbate peroxidase (APX) cDNA, designated MaAPX1, was isolated from banana fruit by suppression subtractive hybridization (SSH). MaAPX1 shares an extensive sequence identity (79 to 83%) with other plant APX homologues. Southern blot analysis revealed only two copies of the APX gene in the banana genome. Reverse-transcriptase PCR analysis of MaAPX1 expression confirmed its expression in the root, leaf, flower and fruit, with higher levels detected in the leaf compared to other organs. Real-time quantitative polymerase chain reaction was used to explore expression patterns of MaAPX1 in banana postharvest. In naturally ripened banana fruits, MaAPX1 expression gradually peaked at day 6 after harvest, and subsequently decreased. In ethylene-treated fruits, MaAPX1 expression increased to a maximum at day 3 and then decreased. Meanwhile, in banana treated with abscisic acid, MaAPX1 levels were suppressed from day 0 to 8. These data suggest that MaAPX1 may play distinct roles in the multiple mechanisms that underlie banana fruit ripening. Key words : Banana, postharvest ripening, ascorbate peroxidase, gene expression, ethylene, abscisic acid.

KEY MESSAGE : The interaction of MuMADS1 and MuUBA in banana was reported, which will help us to understand the mechanism of the MADS-box gene in regulating banana fruit development and ripening. The ubiquitin-activating enzyme E1 gene fragment MuUBA was obtained from banana (Musa acuminata L.AAA) fruit by the yeast two-hybrid method using the banana MADS-box gene MuMADS1 as bait and 2-day post-harvest banana fruit cDNA library as prey. MuMADS1 interacted with MuUBA. The interaction of MuMADS1 and MuUBA in vivo was further proved by bimolecular fluorescence complementation assay. Real-time quantitative PCR evaluation of MuMADS1 and MuUBA expression patterns in banana showed that they are highly expressed in the ovule 4 stage, but present in low levels in the stem, which suggests a simultaneously differential expression action exists for both MuMADS1 and MuUBA in different tissues and developmental fruits. MuMADS1 and MuUBA expression was highly stimulated by exogenous ethylene and suppressed by 1-methylcyclopropene. These results indicated that MuMADS1 and MuUBA were co-regulated by ethylene and might play an important role in post-harvest banana fruit ripening.

To investigate the regulation of 14-3-3 proteins in banana (Musa acuminata L. AAA group, cv. Brazilian) fruit postharvest ripening, four cDNAs encoding 14-3-3 proteins were isolated from banana and designated as Ma-14-3-3a, Ma-14-3-3c, Ma-14-3-3e, and Ma-14-3-3i, respectively. Amino acid sequence alignment showed that the four 14-3-3 proteins shared a highly conserved core structure and variable C-terminal as well as N-terminal regions with 14-3-3 proteins from other plant species. Phylogenetic analysis revealed that the four 14-3-3 genes belong to the non-e groups. They were differentially and specifically expressed in various tissues. Real-time RT-PCR analysis indicated that these four genes function differentially during banana fruit postharvest ripening. Three genes, Ma-14-3-3a, Ma-14-3-3c, and Ma-14-3-3e, were significantly induced by exogenous ethylene treatment. However, gene function differed in naturally ripened fruits. Ethylene could induce Ma-14-3-3c expression during postharvest ripening, but expression patterns of Ma-14-3-3a and Ma-14-3-3e suggest that these two genes appear to be involved in regulating ethylene biosynthesis during fruit ripening. No obvious relationship emerged between Ma-14-3-3i expression in naturally ripened and 1-MCP (1-methylcyclopropene)-treated fruit groups during fruit ripening. These results indicate that the 14-3-3 proteins might be involved in various regulatory processes of banana fruit ripening. Further studies will mainly focus on revealing the detailed biological mechanisms of these four 14-3-3 genes in regulating banana fruit postharvest ripening.

To investigate the regulation of MADS-box genes in banana (Musa acuminata L. AAA group cv. Brazilian) fruit development and postharvest ripening, we isolated from banana fruit a MADS-box gene designated MuMADS1. Amino acid alignment indicated MuMADS1 belongs to the AGAMOUS subfamily, and phylogenetic analysis indicates that this gene is most similar to class D MADS-box genes. Reverse transcriptase-polymerase chain reaction (RT-PCR) analysis showed that MuMADS1 is expressed in the stamen and pistil of male and female flowers and in the rhizome, the vegetative reproductive organ of the banana plant. In preharvest banana fruit, MuMADS1 is likely expressed throughout banana fruit development. In postharvest banana ripening, MuMADS1 is associated with ethylene biosynthesis. Expression patterns of MuMADS1 during postharvest ripening as determined by real-time RT-PCR suggest that differential expression of MuMADS1 may not only be induced by ethylene biosynthesis associated with postharvest banana ripening, but also may be induced by exogenous ethylene.

Article history:Received 7 June 2014Accepted 13 August 2014Available online 18 September 2014Keywords:ASRBananaExpression patternsSalt resistance Background:Abscisicacid(ABA)-,stress-andripenin g-inducedprotein(ASR)isplant-speci fichydrophilictranscriptionalregulatorsinvolvedinsucrosestr essandwoundinginbanana.However,itisnotknownwhetherbanana ASR genesconfersalt stress tolerance. The contexts of the study was to analysis the sequence characterization of banana ASR1,andidentifyits expression patterns and function under salt stress using quantitative real-time PCR (qPCR) and overexpression inArabidopsis . The purpose was to evaluate the role of banana ASR1 to salt stress tolerance employed by plants.Results: Afull-lengthcDNAisolatedfrombananafruitwasnamed MaASR1,andithada432bpopenreadingframe(ORF)encoding 143 amino acids. MaASR1 was preferential expression in roots and leaves compared to low expression in fruits,rhizomes and flowers. Under salt stress, the expression of MaASR1 quickly increased and highest expression level wasdetected in roots and leaves at 4 h, and then gradually decreased. These results suggested that MaASR1 expression wasinduced under salt stress. MaASR1 protein was localized in the nucleus and plasma membrane. MaASR1 wastransformed to Arabidopsis and veri fied by southern and northern analysis, tran sgenic lines L14 and L38 integrated oneand two copies of MaASR1, respectively, while overexpression in tran sgenic lines provided evidence for the role ofMaASR1 to salt stress tolerance.Conclusions: This study demonstrated that overexpression of MaASR1 in Arabidopsis confers salt stress tolerance byreducing the expression of ABA/stress -responsive genes, but does not affect the expression of the ABA-independentpathway and biosynth esis pathway genes.©2014Pontificia Universidad Catolica de Valparaiso. Production and hosting by Elsevier B.V. All rights reserved.