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SLAF marker based QTL mapping of fruit-related traits reveals a major-effect candidate locus ff2.1 for flesh firmness in melon
CHEN Ke-xin, DAI Dong-yang, WANG Ling, YANG Li-min, LI Dan-dan, WANG Chao, JI Peng, SHENG Yun-yan
2023, 22 (11): 3331-3345.   DOI: 10.1016/j.jia.2023.02.014
Abstract166)      PDF in ScienceDirect      

Flesh firmness (FF) is an important and complex trait for melon breeders and consumers.  However, the genetic mechanism underlying FF is unclear.  Here, a soft fruit melon (P5) and a hard fruit melon (P10) were crossed to generate F2, and the FF and fruit-related traits were recorded for two years.  By performing quantitative trait locus (QTL) specific-locus amplified fragment (SLAF) (QTL-SLAF) sequencing and molecular marker-linkage analysis, 112 844 SLAF markers were identified, and 5 919 SNPs were used to construct a genetic linkage map with a total genetic distance of 1 356.49 cM.  Ten FF- and fruit-related QTLs were identified.  Consistent QTLs were detected for fruit length (FL) and fruit diameter (FD) in both years, and QTLs for single fruit weight (SFW) were detected on two separate chromosomes in both years.  For FF, the consistent major locus (ff2.1) was located in a 0.17-Mb candidate region on chromosome 2.  Using 429 F2 individuals derived from a cross between P5 and P10, we refined the ff2.1 locus to a 28.3-kb region harboring three functional genes.  These results provide not only a new candidate QTL for melon FF breeding but also a theoretical foundation for research on the mechanism underlying melon gene function.

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Genome-wide pedigree analysis of elite rice Shuhui 527 reveals key regions for breeding
REN Yun, CHEN Dan, LI Wen-jie, TAO Luo, YUAN Guo-qiang, CAO Ye, LI Xue-mei, DENG Qi-ming, WANG Shi-quan, ZHENG Ai-ping, ZHU Jun, LIU Huai-nian, WANG Ling-xia, LI Ping, LI Shuang-cheng
2021, 20 (1): 35-45.   DOI: 10.1016/S2095-3119(20)63256-7
Abstract226)      PDF in ScienceDirect      
Hybrid rice significantly contributes to the food supply worldwide.  Backbone parents play important roles in elite hybrid rice breeding systems.  In this study, we performed pedigree-based analysis of the elite backbone parent rice variety, namely, Shuhui 527 (SH527, Oryza sativa), to exploit key genome regions during breeding.  Twenty-four cultivars (including SH527, its six progenitors and 17 derived cultivars) were collected and analyzed with high-density single nucleotide polymorphism (SNP) array.  Scanning all these cultivars with genome-wide SNP data indicated the unique contributions of progenitors to the SH527 genome and identified the key genomic regions of SH527 conserved within all its derivatives.  These findings were further supported by known rice yield-related genes or unknown QTLs identified by genome-wide association study.  This study reveals several key regions for SH527 and provides insights into hybrid rice breeding.
 
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Detection of antimicrobial resistance and virulence-related genes in Streptococcus uberis and Streptococcus parauberis isolated from clinical bovine mastitis cases in northwestern China
ZHANG Hang, YANG Feng, LI Xin-pu, LUO Jin-yin, WANG Ling, ZHOU Yu-long, YAN Yong, WANG Xu-rong, LI Hong-sheng
2020, 19 (11): 2784-2791.   DOI: 10.1016/S2095-3119(20)63185-9
Abstract118)      PDF in ScienceDirect      
The objectives of this study were to investigate antimicrobial resistance of Streptococcus uberis and Streptococcus parauberis isolated from cows with bovine clinical mastitis in China and to examine the distribution of resistance- and virulence-related gene patterns.  Antimicrobial susceptibility was determined by the E-test.  Genes encoding antimicrobial resistance and invasiveness factors were examined by PCR.  A total of 27 strains were obtained from 326 mastitis milk samples.  Streptococcus parauberis isolates (n=11) showed high resistance to erythromycin (90.9%), followed by tetracycline (45.5%), chloramphenicol (36.4%) and clindamycin (27.3%).  Streptococcus uberis isolates (n=16) were highly resistant to tetracycline (81.3%) and clindamycin (62.5%).  Both species were susceptible to ampicillin.  The most prevalent resistance gene in S. uberis was tetM (80.0%), followed by blaZ (62.5%) and ermB (62.5%).  However, tetM, blaZ, and ermB genes were only found in 27.3, 45.5, and 27.3%, respectively, of S. parauberis.  In addition, all of the isolates carried at least one selected virulence-related gene.  The most prevalent virulence-associated gene pattern in the current study was sua+pauA/skc+gapC+hasC detected in 22.2% of the strains.  One S. uberis strain carried 7 virulence-associated genes and belonged to the sua+pauA/skc+gapC+cfu+hasA+hasB+hasC pattern.  More than 59.3% of analysed strains carried 4 to 7 virulence-related genes.  Our findings demonstrated that S. parauberis and S. uberis isolated from clinical bovine mastitis cases in China exhibited diverse molecular ecology, and that the strains were highly resistant to antibiotics commonly used in the dairy cow industry.  The data obtained in the current study contribute to a better understanding of the pathogenesis of bacteria in mastitis caused by these pathogens, and the findings are relevant to the development of multivalent vaccines and targeted prevention procedures.
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Overexpression of GmBIN2, a soybean glycogen synthase kinase 3 gene, enhances tolerance to salt and drought in transgenic Arabidopsis and soybean hairy roots
WANG Ling-shuang, CHEN Qing-shan, XIN Da-wei, QI Zhao-ming, ZHANG Chao, LI Si-nan, JIN Yang-mei, LI Mo, MEI Hong-yao, SU An-yu, WU Xiao-xia
2018, 17 (09): 1959-1971.   DOI: 10.1016/S2095-3119(17)61863-X
Abstract530)      PDF in ScienceDirect      
Glycogen synthase kinase 3 (GSK3) is a kind of serine/threonine kinase widely found in eukaryotes.  Many plant GSK3 kinases play important roles in regulating stress responses.  This study investigated BRASSINOSTEROID-INSENSITIVE 2 (GmBIN2) gene, a member of the GSK3 protein kinase family in soybean and an orthologue of Arabidopsis BIN2/AtSK21GmBIN2 expression was increased by salt and drought stresses, but was not significantly affected by the ABA treatment.  To examine the function of GmBIN2, transgenic Arabidopsis and transgenic soybean hairy roots were generated.  Overexpression of GmBIN2 in Arabidopsis resulted in increased germination rate and root length compared with wild-type plants under salt and mannitol treatments.  Overexpression of GmBIN2 increased cellular Ca2+ content and reduced Na+ content, enhancing salt tolerance in transgenic Arabidopsis plants.  In the soybean hairy root assay, overexpression of GmBIN2 in transgenic roots also showed significantly higher relative root growth rate than the control when subjected to salt and mannitol treatments.  Measurement of physiological indicators, including proline content, superoxide dismutase (SOD) activity, and relative electrical conductivity, supported this conclusion.  Furthermore, we also found that GmBIN2 could up-regulate the expression of some stress-related genes in transgenic Arabidopsis and soybean hairy roots.  Overall, these results indicated that GmBIN2 improved tolerance to salt and drought in transgenic Arabidopsis and soybean hairy roots.
 
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A study on the pathogen species and physiological races of tomato Fusarium wilt in Shanxi, China
CHANG Yin-dong, DU Bin, WANG Ling, JI Pei, XIE Yu-jie, LI Xin-feng, LI Zhi-gang, WANG Jian-ming
2018, 17 (06): 1380-1390.   DOI: 10.1016/S2095-3119(18)61983-5
Abstract488)      PDF in ScienceDirect      

In order to clarify the main pathogens of tomato Fusarium wilt in Shanxi Province, China, morphological identification, elongation factor 1 alpha (EF-1α) sequence analysis, specific primer amplification and pathogenicity tests were applied to study the isolates which were recovered from diseased plants collected from 17 different districts of Shanxi Province.  The results were as follows: 1) Through morphological and molecular identification, the following 7 species of Fusarium were identified: F. oxysporum, F. solani, F. verticillioidesF. subglutinans, F. chlamydosporum, F. sporotrichioides, and F. semitectum; 2) 56 isolates of F. oxysporum were identified using specific primer amplification, among which, 29, 5 and 6 isolates were respectively identified as F. oxysporum f. sp. lycopersici physiological race 1, race 2, and race 3; 3) pathogenicity test indicated the significant pathogenicity of F. oxysporum, F. solani, F. verticillioides, and F. subglutinans to tomato plant.  Therefore, among these 4 species confirmed as pathogenic to tomato in Shanxi, the highest isolation rate (53.3%) corresponded to F. oxysporum.  Three physiological species, race 1, race 2, and race 3 of F. oxysporum f. sp. lycopersici are detected in Shanxi, among which race 1 is the most widespread pathogen and is also considered as the predominant race.

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Prevalence and characteristics of extended spectrum β-lactamaseproducing Escherichia coli from bovine mastitis cases in China
YANG Feng, ZHANG Shi-dong, SHANG Xiao-fei, WANG Xu-rong, WANG Ling, YAN Zuo-ting, LI Hong-sheng
2018, 17 (06): 1246-1251.   DOI: 10.1016/S2095-3119(17)61830-6
Abstract524)      PDF in ScienceDirect      
The aim of the study was to investigate the prevalence and characterization of extended-spectrum β-lactamase (ESBL)-producing Escherichia coli isolated from bovine mastitis cases in China.  ChromID ESBL agar was used to confirm ESBL-producing E. coli.  PCR and DNA sequencing were employed to characterize the genotype of ESBL-producers.  Antimicrobial susceptibility was measured by disc diffusion.  Overall, 73 of 318 E. coli isolates (22.96%) were identified as ESBL-producers.  Of these ESBL-producing E. coli, the prevalence of blaCTX-M and blaTEM-1 was 97.26 and 71.23%, respectively.  The predominant CTX-M-type ESBL was CTX-M-15 (65.75%), followed by CTX-M-14 (10.96%), CTX-M-55 (9.59%), CTX-M-64 (5.48%), CTX-M-65 (4.11%) and CTX-M-3 (1.37%).  This study is the first report of CTX-M-64 and CTX-M-65 in E. coli isolated from bovine mastitis.  Furthermore, 72 ESBL-producing E. coli isolates (98.63%) were found to be multidrug-resistance.  This study noted high prevalence and rates of antimicrobial resistance of ESBL-producing E. coli isolates from bovine mastitis cases in China.
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Penicillin-resistant characterization of Staphylococcus aureus isolated from bovine mastitis in Gansu, China
YANG Feng, LIU Long-hai, WANG Ling, WANG Xu-rong, LI Xin-pu, LUO Jin-yin, ZHANG Zhe, ZHANG Shi-dong, YAN Zuo-ting, LI Hong-sheng
2017, 16 (08): 1874-1878.   DOI: 10.1016/S2095-3119(16)61531-9
Abstract1414)      PDF in ScienceDirect      
    Bovine mastitis caused by Staphylococcus aureus is difficult to treat because of increasing resistance against antibiotics, especially penicillin. β-Lactamase and biofilm are responsible for penicillin resistance of S. aureus. The aim of this study was to investigate the β-lactamase activity and biofilm formation capacity of 37 penicillin-resistant S. aureus strains (35 were blaZ positive and 2 were blaZ negative) from bovine mastitis in Gansu Province, China, as well as to measure the intercellular adhesion genes icaA and icaD of these strains. β-Lactamase Test Kit was used to determine the β-lactamase activity, biofilm formation was tested by semi-quantitative adherence assay method. Moreover, the presence of icaA and icaD were measured by PCR. A total of 32 penicillin-resistant S. aureus strains, including the two blaZ-negative strains, were identified as β-lactamase producers. All tested S. aureus isolates produced biofilm in the microtiter plate assay. Meanwhile, all these strains were PCR-positive for the ica locus, icaA and icaD. The study indicated high prevalence of β-lactamase activity, biofilm-forming capacity, and the ica genes among the penicillin-resistant S. aureus isolates, and implied that S. aureus resistant to penicillin was attributed to multiple mechanisms.
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Molecular characterization and tissue expression profile of the Dnmts gene family in pig
LUO Zong-gang, ZHANG Kai, CHEN Lei, YANG Yuan-xin, FU Peng-hui, WANG Ke-tian, WANG Ling, LI Ming-zhou, LI Xue-wei, ZUO Fu-yuan, WANG Jin-yong
2017, 16 (06): 1367-1374.   DOI: 10.1016/S2095-3119(16)61512-5
Abstract803)      PDF in ScienceDirect      
DNA methyltransferases (Dnmts) comprise a family of proteins which involved in the establishment and maintenance of DNA methylation patterns.  In pig, the molecular characterization and tissue expression profile of Dnmt gene family are not clear.  To solve this problem, reverse transcriptase PCR and rapid amplification of cDNA ends were used to clone the sequences of the porcine Dnmt2 and Dnmt3b genes.  Furthermore, the mRNA expression profiles of Dnmt1, Dnmt2, Dnmt3a and Dnmt3b genes from 54 adult tissues and 2 entire fetuses of Rongchang pig were analyzed by quantitative real-time PCR (qRT-PCR).  As a result, the lengths of porcine Dnmt2 and Dnmt3b gene cDNAs were 1 227 and 2 559 bp with cytosine-C5 specific DNA methylase domain, respectively.  The four Dnmt genes were highly expressed in longissimus dorsi muscle (P<0.01).  Dnmt1 is highly expressed in heart (P<0.01) and Dnmt 2 shows its preference in liver and seminal vesicle tissue (P<0.01).  Dnmt3a and Dnmt3b are highly expressed in the two fetus stages (P<0.01).  All these results suggested that each gene has its specific expression profile, and deeper study is required to dig more details between the methylation level and Dnmt family mRNA expressions in different tissues.
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Cytochemical localization of H2O2 in pigment glands of cotton (Gossypium hirsutum L.)
WANG Ling-li, ZHENG Shuang-shuang, TONG Pan-pan, CHEN Yan, LIU Wen-zhe
2016, 15 (7): 1490-1498.   DOI: 10.1016/S2095-3119(15)61210-2
Abstract1507)      PDF in ScienceDirect      
    Programmed cell death (PCD) plays a critical role in the development of plant pigment glands, while H2O2, which is a kind of reactive oxygen species (ROS) produced by the aerobic metabolism of cells, acts as an important signal in this process. Here, we investigated the temporal and spatial dynamics of accumulated H2O2 in pigment glands of Gossypium hirsutum L. with 3,3-diaminobenzidine (DAB) staining, 2’,7’-dichlorodihydrofluorescein diacetate (DCFH2)-DA fluorescent labeling and CeCl3 cytochemical localization techniques. The results showed that the pigment glands of G. hirsutum could generate H2O2, and the amount and localization of H2O2 varied at different developmental stages. At the early developmental stage, a small amount of HH2O2 accumulated in the vacuole membrane of pigment gland cells. At the intermediate stage, a large number of H2O2 appeared in the vacuole membrane, while cell walls started to accumulate a small amount of H2O2. When pigment gland cell degraded, H2O2 mainly accumulated on the chloroplast envelope membrane of inner sheath cells. With the degradation of the sheath cells, H2O2 was detected in cell wall and the membrane of secretory vesicles which contains the preliminary contents of pigment gland. With the pigment glands completely maturation, H2O2 would disappeared. The accumulation sites of H2O2 are consistent with the process of PCD of individual gland cells, which started from the degradation of intracellular membrane and ended with the degradation of cell walls. Thus H2O2 probably plays an important role in the development of pigment glands. In addition, the development of pigment glands and the generation of H2O2 are not associated with the light, and no H2O2 was detected in the secretions of pigment glands.
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Genetic characterization of antimicrobial resistance in Staphylococcus aureus isolated from bovine mastitis cases in Northwest China
YANG Feng, WANG Qi, WANG Xu-rong, WANG Ling, LI Xin-pu, LUO Jin-yin, ZHANG Shi-dong, LI Hong-sheng
2016, 15 (12): 2842-2847.   DOI: 10.1016/S2095-3119(16)61368-0
Abstract1119)      PDF in ScienceDirect      
    Staphylococcus aureus is the most common etiological pathogen of bovine mastitis. The resistant strains make the disease difficult to cure. The aim of this study was to characterize the genetic nature of the antimicrobial resistance in S. aureus cultured from bovine mastitis in Northwest China in 2014. A total of 44 S. aureus were isolated for antimicrobial resistance and resistance-related genes. Antimicrobial resistance was determined by disc diffusion and the corresponding resistance genes were detected by PCR. Phenotype indicated that S. aureus isolates were resistant to penicillin (84.09%), erythromycin (20.45%), tetracycline (15.91%), gentamicin (9.09%), tobramycin (6.82%), kanamycin (6.82%) and methicillin (2.27%). 9.09% of the S. aureus isolates were classified as multidrug resistant. In addition, genotypes showed that the isolates were resistant to rifampicin (100%, rpoB), penicillin (95.45%, blaZ), tetracycline (22.73%, tetK, tetM, alone or in combination), erythromycin (22.73%, ermB or ermC), gentamicin/tobramycin/kanamycin (2.27%, aacA-aphD), methicillin (2.27%, mecA) and vancomycin (2.27%, vanA). Resistance to tetracycline was attributed to the genes tetK and tetM (r=0.558, P<0.001). This study noted high-level geno- and phenotypic antimicrobial resistance in S. aureus isolates from bovine mastitis cases in Northwest China.
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YGL9, encoding the putative chloroplast signal recognition particle 43 kDa protein in rice, is involved in chloroplast development
WANG Zhong-wei, ZHANG Tian-quan, XING Ya-di, ZENG Xiao-qin, WANG Ling, LIU Zhong-xian, SHI Jun-qiong, ZHU Xiao-yan, MA Ling, LI Yun-feng, LING Ying-hua, SANG Xian-chun, HE Guang-hua
2016, 15 (05): 944-953.   DOI: 10.1016/S2095-3119(15)61310-7
Abstract1656)      PDF in ScienceDirect      
    The nuclear-encoded light-harvesting chlorophyll a/b-binding proteins (LHCPs) are specifically translocated from the stroma into the thylakoid membrane through the chloroplast signal recognition particle (cpSRP) pathway. The cpSRP is composed of a cpSRP43 protein and a cpSRP54 protein, and it forms a soluble transit complex with LHCP in the chloroplast stroma. Here, we identified the YGL9 gene that is predicted to encode the probable rice cpSRP43 protein from a rice yellow-green leaf mutant. A phylogenetic tree showed that an important conserved protein family, cpSRP43, is present in almost all green photosynthetic organisms such as higher plants and green algae. Sequence analysis showed that YGL9 comprises a chloroplast transit peptide, three chromodomains and four ankyrin repeats, and the chromodomains and ankyrin repeats are probably involved in protein-protein interactions. Subcellular localization showed that YGL9 is localized in the chloroplast. Expression pattern analysis indicated that YGL9 is mainly expressed in green leaf sheaths and leaves. Quantitative real-time PCR analysis showed that the expression levels of genes associated with pigment metabolism, chloroplast development and photosynthesis were distinctly affected in the ygl9 mutant. These results indicated that YGL9 is possibly involved in pigment metabolism, chloroplast development and photosynthesis in rice.
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Development of lepidopteran pest-resistant transgenic japonica rice harboring a synthetic cry2A* gene
JIN Yong-mei, MA Rui, YU Zhi-jing, WANG Ling, JIANG Wen-zhu, LIN Xiu-feng
2015, 14 (3): 423-429.   DOI: 10.1016/S2095-3119(14)60897-2
Abstract1504)      PDF in ScienceDirect      
A synthetic cry2A* gene encoding Bacillus thuringiensis (Bt) δ-endotoxin that resistance to lepidopteran pest was transformed into japonica rice variety Jijing 88, which is the most widely cultivated variety in Jilin Province, Northeast China, by Agrobacterium-mediated transformation. A total of 106 independent transformants overexpressing cry2A* gene driven by ubiquitin (Ubi) promoter was produced. Three single-copy homozygous transgenic lines were finally selected based on the results of PCR analysis, segregation ratio of Basta resistance, and Southern hybridization analyses. RT-PCR and enzyme linked immune sorbent assay (ELISA) revealed that cry2A* transcripts and protein were highly expressed in these lines. The high level of Cry2A* protein expression resulted in high resistance to rice striped stem borer as evidenced by insect feeding bioassays. Our results demonstrate that cry2A* transgenic japonica rice confers resistance to the rice striped stem borer in the laboratory conditions.
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Preparation, Characterization and Nematicidal Activity of Lansiumamide B Nano-Capsules
YIN Yan-hua, GUO Qing-ming, HAN Yun, WANG Ling-jing, WAN Shu-qing
2012, 12 (7): 1151-1158.   DOI: 10.1016/S1671-2927(00)8641
Abstract1792)      PDF in ScienceDirect      
In this study, nano-capsules of lansiumamide B (NCLB) was prepared by the microemulsion polymerization method to improve the nematicidal efficacy of lansiumamide B (LB). An optimal formulation was gained by orthogonal experiment design based on the encapsulation efficiency (En, %) value. The optimized NCLB were spherical and uniform under transmission electron microscopy (TEM). The mean particle size, zeta potential and En were (38.50±0.64) nm, (-70.5±0.76) mV and (95.13±1.16)%, respectively. The release profile indicated that the accumulated release of LB in NCLB reached up to 82% within 96 h. Effects of NCLB against Bursaphelenehus xylophilus and J2 of Meloidogyne incognita were reported in this paper. The nematicidal activity of NCLB has been remarkably increased, with LC50 values of 2.1407 mg L-1 and 19.3608 mg L-1, respectively, at 24 h after treatment. The disease progression and the average number of root knots of Ipomoea aquatica were 1.50 and 7.25, respectively, in the treatment of NCLB, at concentration of 200 mg L-1, significantly lower than the treatment of LB and ethoprophos. Compared to control, the treatments of NCLB, LB and ethoprophos leaded the disease progression to drop 68.42, 36.84 and 26.32%, respectively, and caused the average number of root knots to fall 83.94, 78.03 and 63.66%. These results suggested that NCLB, as a novel nematicides formulation, performed more efficient and longer effective maintenance against plant parasitic nematodes.
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