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Through analysis of ABO blood group gene typing technology, to assist in the identification of difficult clinical serological specimens.A total of 10 forwardreverse typing ambiguous samples were collected from January 2021 to August 2021 in our hospital.ABO genotypes were analysed by gene sequencing.The genotypes of 10 ABO ambiguous blood group samples were A102/BW11, A102/BW12, O02/O02, A102/B303, A102/B101, BW11/O02, B101/O04, BW11/O01, BW11/O01, A101/O02, respectively. The genotype results of 6 cases was consistent with the serological phenotype, and the serological phenotype of 4 cases were different from the geno sequencing.ABO blood groups genotyping technology combined with serological typing can be used for accurate typing of ambiguous blood group, and better ensure the safety of blood transfusion.10例ABO疑难血型的基因测序分析.利用ABO血型基因分型技术协助临床血清学疑难血型的鉴定.收集2021年1月－2021年8月本院用血清学鉴定ABO正反定型不符的疑难血型标本10例，采用基因测序方法检测ABO基因型.10例疑难血型标本的ABO基因型分别为A102/Bw11、A102/Bw12、O02/O02、A102/B303、A102/B101、Bw11/O02、B101/O04、Bw11/O01、Bw11/O01、A101/O02。其中有6例标本血型基因测序结果和血清学表型一致，其他4例标本的血清学表型和基因测序结果均表现出一定的差异.鉴定ABO疑难血型可利用基因分型技术结合血清学方法，最终准确鉴定血型，更好的保障输血安全.

Low temperature is one of the key environmental stresses, which greatly affects global banana production. However, little is known about the global phosphoproteomes in Musa spp. and their regulatory roles in response to cold stress. In this study, we conducted a comparative phosphoproteomic profiling of cold-sensitive Cavendish Banana and relatively cold tolerant Dajiao under cold stress. Phosphopeptide abundances of five phosphoproteins involved in MKK2 interaction network, including MKK2, HY5, CaSR, STN7 and kinesin-like protein, show a remarkable difference between Cavendish Banana and Dajiao in response to cold stress. Western blotting of MKK2 protein and its T31 phosphorylated peptide verified the phosphoproteomic results of increased T31 phosphopeptide abundance with decreased MKK2 abundance in Daojiao for a time course of cold stress. Meanwhile increased expression of MKK2 with no detectable T31 phosphorylation was found in Cavendish Banana. These results suggest that the MKK2 pathway in Dajiao, along with other cold-specific phosphoproteins, appears to be associated with the molecular mechanisms of high tolerance to cold stress in Dajiao. The results also provide new evidence that the signaling pathway of cellular MKK2 phosphorylation plays an important role in abiotic stress tolerance that likely serves as a universal plant cold tolerance mechanism.

The inhibitory effects of Chinese leek(Allium tuberosum) on Fusarium oxysporum f. sp. cubense (Foc) and on Fusarium wilt incidence were studied in order to identify a potential efficient way to control the disease. Adopting the rotation system of Chinese leek-banana reduced the Fusarium wilt incidence and disease severity index by 88 %-97 % and 91 %-96 %, respectively, improved the crop value by 36 %-86 %, in an area heavily infested by Foc between 2007 and 2009. As a result of inoculation in the greenhouse, Chinese leek treatment reduced disease incidence and the disease severity index by 58 % and 62 %, respectively in the variety Baxi (AAA) and by 79 % and 81 %, respectively in the variety Guangfen NO.1 (ABB). Crude extracts of Chinese leek completely inhibited the growth of Foc race 4 on Petri dishes, suppressed the proliferation of the spores by 91 % and caused 87 % spore mortality. The findings of this study suggest that Chinese leek has the potential to inhibit Foc growth and Fusarium wilt incidence. This potential may be developed into an environmentally friendly treatment to control Fusarium wilt of banana.

To attempt to introduce genetic information of disease resistance from Musa acuminata cv. Mas (AA) to Musa silk cv. Guoshanxiang (AAB) and obtain somatic hybrids, we developed an asymmetric protoplast fusion with 20% (w/v) polyethylene glycol (PEG). The protoplasts derived from embryogenic suspension cultural cells of cv. Guoshanxiang (AAB) and cv. Mas (AA) were, respectively treated with 1.5 mM iodoacetamide (IOA) and with ultraviolet light (UV) at an intensity of 50 W/m2 for 120 s. A total of 47 regenerated green plants were obtained and eight of which were survived in greenhouse. Six of the survived plants were identified as hybrids by RAPD analysis and only three hybrids were retained vigorously in field. The hybrid nature of the three plants was further confirmed according to their ISSR (inter-simple sequence repeat) patterns and the results indicated that they were true somatic hybrids. Chromosome analysis revealed that the three hybrids possessed an aneuploid chromosome number (2n = 34).

Cold is an environmental factor that limits the growing season of banana and adversely affects fruit quality and productivity. C-repeat/dehydration responsive element binding factor 1 (CBF1) play important roles in the responses of plants to low-temperature and cold. To test the function and potential use of cold-induced factor CBF1 in improving the chill tolerance of banana, AtCBF1 driven by a cauliflower mosaic virus 35S promoter was introduced into banana plants by Agrobacterium -mediated transformation of embryogenic cell suspensions (ECSs) of banana cultivar Furenzhi (AA). The presence of AtCBF1 transgenes in regenerated plants was confirmed by polymerase chain reaction (PCR) and Southern blots. s-glucoronidase (GUS) histochemical assays reverse transcription-polymerase chain reaction (RT-PCR) and real-time PCR analysis demonstrated that foreign genes were stably expressed in regenerated plants. Transgenic banana showed growth retardation, thicker leaves and higher chlorophyll content than non-transformed plants. While relative electrolyte leakage (REL) and malondialdehyde (MDA) content were significantly lower in transgenic banana plants than in non-transformed plants under low temperature stress. An obviously higher cold tolerance was observed among the transgenic plants in the cold detection. These results suggest that over-expression of AtCBF 1 in transgenic banana plants plays an important role in improving tolerance to low-temperature.

BACKGROUND: Fusarium wilt, caused by the fungal pathogen Fusarium oxysporum f. sp. cubense (Foc), is one of the most destructive diseases of banana. Toxins produced by Foc have been proposed to play an important role during the pathogenic process. The objectives of this study were to investigate the contamination of banana with toxins produced by Foc, and to elucidate their role in pathogenesis. METHODOLOGY/PRINCIPAL FINDINGS: Twenty isolates of Foc representing races 1 and 4 were isolated from diseased bananas in five Chinese provinces. Two toxins were consistently associated with Foc, fusaric acid (FA) and beauvericin (BEA). Cytotoxicity of the two toxins on banana protoplast was determined using the Alamar Blue assay. The virulence of 20 Foc isolates was further tested by inoculating tissue culture banana plantlets, and the contents of toxins determined in banana roots, pseudostems and leaves. Virulence of Foc isolates correlated well with toxin deposition in the host plant. To determine the natural occurrence of the two toxins in banana plants with Fusarium wilt symptoms, samples were collected before harvest from the pseudostems, fruit and leaves from 10 Pisang Awak 'Guangfen #1' and 10 Cavendish 'Brazilian' plants. Fusaric acid and BEA were detected in all the tissues, including the fruits. CONCLUSIONS/SIGNFICANCE: The current study provides the first investigation of toxins produced by Foc in banana. The toxins produced by Foc, and their levels of contamination of banana fruits, however, were too low to be of concern to human and animal health. Rather, these toxins appear to contribute to the pathogenicity of the fungus during infection of banana plants.

Banana and its close relative, plantain are globally important crops and there is considerable interest in optimizing their cultivation. Plantain has superior cold tolerance compared with banana and a thorough understanding of the molecular mechanisms and responses of plantain to cold stress has great potential value for developing cold tolerant banana cultivars. In this study, we used iTRAQ-based comparative proteomic analysis to investigate the temporal responses of plantain to cold stress. Plantain seedlings were exposed for 0, 6, and 24 h of cold stress at 8 °C and subsequently allowed to recover for 24 h at 28 °C. A total of 3477 plantain proteins were identified, of which 809 showed differential expression from the three treatments. The majority of differentially expressed proteins were predicted to be involved in oxidation-reduction, including oxylipin biosynthesis, whereas others were associated with photosynthesis, photorespiration, and several primary metabolic processes, such as carbohydrate metabolic process and fatty acid beta-oxidation. Western blot analysis and enzyme activity assays were performed on seven differentially expressed, cold-response candidate plantain proteins to validate the proteomics data. Similar analyses of the seven candidate proteins were performed in cold-sensitive banana to examine possible functional conservation, and to compare the results to equivalent responses between the two species. Consistent results were achieved by Western blot and enzyme activity assays, demonstrating that the quantitative proteomics data collected in this study are reliable. Our results suggest that an increase of antioxidant capacity through adapted ROS scavenging capability, reduced production of ROS, and decreased lipid peroxidation contribute to molecular mechanisms for the increased cold tolerance in plantain. To the best of our knowledge, this is the first report of a global investigation on molecular responses of plantain to cold stress by proteomic analysis.

Background Fusarium wilt, caused by the fungal pathogen Fusarium oxysporum f. sp. cubense tropical race 4 (Foc TR4), is considered the most lethal disease of Cavendish bananas in the world. The disease can be managed in the field by planting resistant Cavendish plants generated by somaclonal variation. However, little information is available on the genetic basis of plant resistance to Foc TR4. To a better understand the defense response of resistant banana plants to the Fusarium wilt pathogen, the transcriptome profiles in roots of resistant and susceptible Cavendish banana challenged with Foc TR4 were compared. Results RNA-seq analysis generated more than 103 million 90-bp clean pair end (PE) reads, which were assembled into 88,161 unigenes (mean size = 554 bp). Based on sequence similarity searches, 61,706 (69.99%) genes were identified, among which 21,273 and 50,410 unigenes were assigned to gene ontology (GO) categories and clusters of orthologous groups (COG), respectively. Searches in the Kyoto Encyclopedia of Genes and Genomes Pathway database (KEGG) mapped 33,243 (37.71%) unigenes to 119 KEGG pathways. A total of 5,008 genes were assigned to plant-pathogen interactions, including disease defense and signal transduction. Digital gene expression (DGE) analysis revealed large differences in the transcriptome profiles of the Foc TR4-resistant somaclonal variant and its susceptible wild-type. Expression patterns of genes involved in pathogen-associated molecular pattern (PAMP) recognition, activation of effector-triggered immunity (ETI), ion influx, and biosynthesis of hormones as well as pathogenesis-related (PR) genes, transcription factors, signaling/regulatory genes, cell wall modification genes and genes with other functions were analyzed and compared. The results indicated that basal defense mechanisms are involved in the recognition of PAMPs, and that high levels of defense-related transcripts may contribute to Foc TR4 resistance in banana. Conclusions This study generated a substantial amount of banana transcript sequences and compared the defense responses against Foc TR4 between resistant and susceptible Cavendish bananas. The results contribute to the identification of candidate genes related to plant resistance in a non-model organism, banana, and help to improve the current understanding of host-pathogen interactions.

Conventional breeding for dual resistance of disease and pest of Musa cultivars remains a difficult endeavor, as the plant is polyploidic and high in sterility. Biotechnological techniques, eg., genetic engineering, in vitro mutation breeding, or protoplast fusion, may overcome the difficulties and improve the germplasm. Establishment of a stable embryogenic cell suspension (ECS) is a prerequisite for any of the biotechnological breeding methods. In this study an embryogenic cell suspension was established from immature male flower of Musa acuminata cv. Mas (AA), a popular commercial variety of banana in the South-East Asian region. After culture for 5-6 months on callus induction media, which consisted of MS salts, different concentrations of 2,4-dichlorophenoxyacetic acid (2,4-D), 4.1 micromol/L biotin, 5.7 micromol/L indoleacetic acid (IAA), 5.4 micromol/L naphthaleneacetic acid (NAA), other vitamins, 87 mmol/L sucrose, and solidified with 7 g/L agarose, meristematic globules and yellow, friable embryogenic cultures were induced from the explants of 1-15th row young floral hands of immature male flowers. Of the four treatments of 2,4-D, 9 micromol/L was the most effective on the callus induction, it transformed 40.96% and 7.45% of the cultivated male floral hands into callus and embryogenic callus respectively. The explants to produce highest frequency of the embryogenic calli were floral hands of 6 to 12th rows, which generated 5.79% of the embryogenic calli. Suspension cultures were initiated from these embryogenic calli in liquid medium supplemented with 4.5 micromol/L 2, 4-D. After sieving selection of the cultures using a stainless steel metallic strainer with pore sizes of 154 microm at 15 day intervals for 3 months, homogeneous and yellow embryogenic cell suspensions, composed of single cells and small cell aggregates, were established. Based upon the growth quantity and growth rate of ECS, it was determined that the appropriate inoculum was 2.0 mL PCV ECS/30 mL medium in 100 mL flask, and the appropriate subculture cycle was 15 days. Planting of 6 months old ECS on semi-solid medium of somatic embryo induction and development (MSD) resulted in approximately 280 x 10(3) somatic embryos/mL PCV ECS. MSD contained SH macronutrients, micro-nutrients, Fe-EDTA and MS vitamins supplemented with 4.5 micromol/L biotin, 680 micromol/L glutamine, 2 mmol/L proline, 100 mg/L malt extract, 1.1 micromol/L NAA, 0.2 micromol/L zeatin, 0.5 micromol/L kinetin, 0.7 micromol/L N6-(2-isopentenyl) adenine, 29 mmol/L lactose, 130 mmol/L sucrose and solidified with 2g/L gelrite. After 3 months of maturity on MSD, 17.28% of the somatic embryos were germinated on germination media (MG), consisted of MS salt, Morel and Wetmore vitamins, 0.2 micromol/L 6-BA, 1.1 micromol/L IAA, 87 micromol/L sucrose and solidified with 2 g/L gelrite; and 14.16% of the somatic embryos could develop into normal plantlets on rooting media contained the same composition as that of MG but without auxin and cytokinin.

The histological origin of organogenesis was studied during plant regeneration of banana (Musa AAA cv. Williams) via direct organogenesis from micro-cross-sections of the rhizome in modified Murashige–Skoog (MS) medium (including 10 µm 6-BAP, 1 µm IAA and 50 µm KN). The frequency of the organogenesis was dependent on the position from where the explant was cut. Generally, more adventitious shoots were obtained from explants cut from regions close to apical meristem. The origin of adventitious shoots was multicellular, from cell layers with active cell division in the epidermis of the rhizome axis. After 24 h of culture, a layer of epidermal cells close to the leaf sheath base was actively dividing, and as the time in culture increased, regions containing actively dividing cells became enlarged. By the fifth day of culture, regions of actively dividing cells covered the whole peripheral zone of cortex. Several meristemoid structures were formed from the peripheral zone of cortex after 7 days of culture. These structures developed into adventitious shoots by the ninth day of culture. Vigorous plantlets could be regenerated from the shoots when they were transferred to rooting medium containing MS basal salts supplemented with 30 g L–1 sucrose, 6.5 g L–1 agar, a vitamin/amino acid mixture, 1 µm NAA and 1 µm 6-BAP.

Fusarium wilt is one of the most serious threats to banana production worldwide. The disease is caused by the soil-borne fungal pathogen Fusarium oxysporum f. sp. cubense (Foc), especially tropical race 4 (Foc TR4). It has been reported that crop rotation and intercropping of Chinese leek (Allium tuberosum) with banana can effectively reduce incidence of the disease in the field. In this study, we investigated the toxic mechanism and bioactive compounds of Chinese leek root exudates (CLREs) acting against Foc TR4. In vitro experiments showed that CLREs inhibit the germination and growth of Foc TR4, initiate the accumulation of reactive oxygen species (ROS), and trigger a decrease in the mitochondrial transmembrane potential (ΔΨm). In addition, it induced decreases in the expression of ergosterol biosynthesis genes, and up-regulated the expression of genes associated with autophagy. Dimethyl trisulfide, dimethyl disulfide, 2-propenyl methyl disulfide, 2-propenyl methyl trisulfide, and 2-methoxy-4-vinylphenol are major components of CLREs volatiles, and strongly inhibit Foc TR4 development. These results suggest that CLREs induce cell death in Foc TR4 by inducing oxidative bursts, mitochondrial impairment, and plasma membrane depolarization. In addition, sulfur and phenolic compounds contribute to its antifungal activity.