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Evaluation of a new qPCR test to identify the organisms causing high total bacterial count in bulk tank milk

Journal of Integrative Agriculture 2018年第6期17卷 1241-1245页

作者： Jorgen Katholm Lene Trier Olesen Anders Petersen Snorri Sigurdsson dna diagnostic A/S Risskov 8240Denmark ARLA Foods amba Viby 8260Denmark SEGES Aarhus N 8200Denmark

Milk quality in bulk tank milk （BTM） is measured by flow cytometry technology as total bacterial count （TBC） and somatic cell count （SCC）. To investigate SCC problems, culture or PCR can be used to identify mastitis causing bacteria, e.g., Mastit 4, a commercially available qPCR test. TBC in BTM can be investigated further using culture-based methods such as standard plate count, laboratory pasteurization count, coliform count, and spore counts. To our knowledge, no qPCR addressing the bacteria involved in TBC has been commercially introduced. The aim of this study is to evaluate a recently introduced 3-h qPCR test, TBC 4. The TBC 4 qPCR detects four target groups, Pseudomonas, Streptococci, Enterobacteriacea/ Enterococcus, and Bacillus/Clostridia. These target groups relate to problems on the farm such as cooling, mastitis, environment, and silage. We will continue with new research to compare the TBC 4 qPCR test with traditional culture. For this study, BTM samples from different TBC intervals were selected based on BactoCount results found at routine payment investigation at Eurofins laboratory （Vejen, Denmark）. These samples were analyzed using TBC 4 qPCR assay within 24 h. In total, 346 BTM samples were divided into six different intervals of colony forming units （CFU）. For all four targets in each of the different intervals of CFU, the percent of positive samples, the average Ct-value, the percent of positive samples with Ct〈30 and Ct〈25 were calculated. For Pseudomonas, Streptococci, and Enterobacteriacea/Enterococcus, the number of positive samples with lower Ct-values （high bacteria content） correlated with the CFU mL-1. We found Enterobacteriacea/ Enterococcus, Pseudomonas, and Streptococci in high number of bacteria （Ct〈25） in 25, 19 and 56％ of samples with CFU mL-1 between 50001-100000 and 53, 44, and 39％ in samples with CFU mL-1〉100000. The TBC 4 qPCR test showed to be a strong and fast tool for farmers, advisors and service technicians to address probl

关键词： TBC bulk tank milk qPCR milk quality

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Next-generation sequencing and its clinical application

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Cancer Biology & Medicine 2019年第1期16卷 4-10页

作者： Dahui Qin Molecular diagnostic laboratory Department of Pathology Moffitt Cancer Center

Next-generation sequencing (NGS) is a new technology used for dna and RNA sequencing and variant/mutation detection. NGS can sequence hundreds and thousands of genes or whole genome in a short period of time. The sequence variants/mutations detected by NGS have been widely used for disease diagnosis, prognosis, therapeutic decision, and follow up of patients. The capacity of its massive parallel sequencing offers new opportunities for personalized precision medicine.

关键词： NGS dna capacity

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UFMylation:A Unique & Fashionable Modification for Life

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Genomics, Proteomics & Bioinformatics 2016年第3期14卷 140-146页

作者： Ying Wei Xingzhi Xu Beijing Key laboratory of dna Damage Response and College of Life Sciences Capital Normal University

Ubiquitin-fold modifier 1（UFM1） is one of the newly-identified ubiquitin-like *** to ubiquitin,UFM1 is conjugated to its target proteins by a three-step enzymatic *** UFM1-activating enzyme,ubiquitin-like modifier-activating enzyme 5（UBA5）,serves as the E1 to activate UFM1;UFM1-conjugating enzyme 1（UFC1） acts as the E2 to transfer the activated UFM1 to the active site of the E2;and the UFM1-specific ligase 1（UFL1） acts as the E3 to recognize its substrate,transfer,and ligate the UFM1 from E2 to the *** process is called ***1 chains can be cleaved from its target proteins by UFM1-specific proteases（Uf SPs）,suggesting that the ufmylation modification is ***1 cascade is conserved among nearly all of the eukaryotic organisms,but not in yeast,and associated with several cellular activities including the endoplasmic reticulum stress response and ***,the UFM1 cascade is closely related to a series of human *** this review,we summarize the molecular details of this reversible modification process,the recent progress of its functional studies,as well as its implication in tumorigenesis and potential therapeutic targets for cancer.

关键词： Ubiquitin-fold modifier 1 Ufmylation Endoplasmic reticulum stress Cancer Post-translation modification Ubiquitin-like proteins

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Cholera:an overview with reference to the Yemen epidemic

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Frontiers of Medicine 2019年第2期13卷 213-228页

作者： Ali A.Rabaan Molecular diagnostic laboratory Johns Hopkins Aramco HealthcareDhahran 31311Saudi Arabia

Cholera is a secretory diarrhoeal disease caused by infection with Vibrio cholerae, primarily the V. cholerae O1 El Tor biotype. There are approximately 2.9 million cases in 69 endemic countries annually, resulting in 95 000 deaths. Cholera is associated with poor infrastructure and lack of access to sanitation and clean drinking water. The current cholera epidemic in Yemen, linked to spread of V. cholerae O1 (Ogawa serotype), is associated with the ongoing war. This has devastated infrastructure and health services. The World Health Organization had estimated that 172 286 suspected cases arose between 27th April and 19th June 2017, including 1170 deaths. While there are three oral cholera vaccines prequalified by the World Health Organization, there are issues surrounding vaccination campaigns in conflict situations, exacerbated by external factors such as a global vaccine shortage. Major movements of people complicates surveillance and administration of double doses of vaccines. Cholera therapy mainly depends on rehydration, with use of antibiotics in more severe infections. Concerns have arisen about the rise of antibiotic resistance in cholera, due to mobile genetic elements. In this review, we give an overview of cholera epidemiology, virulence, antibiotic resistance, therapy and vaccines, in the light of the ongoing epidemic in Yemen.

关键词： cholera epidemic multi-drug resistant catechin luteolin ToxT CTXФ

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MYPT1 Sustains Centromeric Cohesion and the Spindle-Assembly Checkpoint

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Journal of Genetics and Genomics 2013年第11期40卷 575-578页

作者： Jing Li Xiaoqian Liu Ji Liao Jie Tian Jue Wang Xin Wang Jiezhong Zhang Xingzhi Xu Beijing Key laboratory of dna Damage Response College of Life SciencesCapital Normal University

During mitosis, cohesins hold the sister chromatids together until anaphase when arm cohesins are removed （Peters et al., 2008; Yao and Dai, 2012）. The shugoshin （Sgo） proteins play pivotal roles during this stage. There is only one shu- goshin in the fly and budding yeasts, while there are two in other organisms （including fission yeasts）. The two mamma- lian shugoshins, Sgol and Sgo2, carry out distinct functions： Sgol mainly in mitosis, and Sgo2 mainly in meiosis and perturbed mitosis. Mitotic cyclin-dependent kinase 1 （CDKI） phosphorylates Sgol, and targets the Sgol-protein phospha- tase 2A （PP2A） complex to protect centromeric cohesin （Kitajima et al., 2006; Tang et al., 2006; Liu et al., 2012）,

关键词： MYPT1 Sustains Centromeric Cohesion and the Spindle-Assembly Checkpoint RNAi PP

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Syrosingopine,an anti-hypertensive drug and lactate transporter(MCT1/4)inhibitor,activates hepatic stellate cells and exacerbates liver fibrosis in a mouse model

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Genes & Diseases 2024年第4期11卷 28-30页

作者： Meichun Guo Yannian Gou Xiangyu Dong Jiamin Zhong Aohua Li Ailing Hao Tongchuan He Jiaming Fan Ministry of Education Key laboratory of diagnostic Medicine and the School of Laboratory Diagnostic MedicineChongqing Medical UniversityChongqing 400016China Molecular Oncology laboratory Department of Orthopaedic Surgery and Rehabilitation MedicineThe University of Chicago Medical CenterChicagoIL 60637USA

Syrosingopine is an anti-hypertensive drug and can cause high intracellular lactate levels and end-product inhibition of lactate dehydrogenase by inhibiting the lactate transporters MCT1 and *** studies have shown that syrosingopine plays an essential role in the process of glycolytic blockade,ATP depletion,and cell death in cancer due to high intracellular levels of lactate.

关键词： hypertensive MCT1 drug

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Anticancer Effects of Crude Extract from Melia toosendan *** Zucc on Hepatocellular Carcinoma In Vitro and In Vivo

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Chinese Journal of Integrative Medicine 2016年第5期22卷 362-369页

作者：刘小玲王虹张伶王友良王进王鹏贺潇何於娟 Department of laboratory Medicine Key Laboratory of Diagnostic Medicine(Ministry of Education)Chongqing Medical University Department of laboratory Medicine People's HospitalPengzhouSichuan Province

Objective： To investigate the anti-cancer effects of crude extract from Melia toosendan Sieb. et Zucc and its possible molecular mechanisms in vitro and in vivo. Methods： Transonic alcohol-chloroform extraction method was used to extract toosendanin from the bark of Melia toosendan Sieb. et Zucc, and the content of toosendanin in the crude extract was measured by high performance liquid chromatography （HPLC）. Anti-cancer effects of crude extract from Melia toosendan Sieb. et Zucc were investigated in in vivo and in vitro studies. In the in vitro experiment, human hepatocellular carcinoma cell lines SMMC-7721 and Hep3B were co-incubated with toosendanin crude extract of different concentrations, respectively. In the in vivo experiment, BALB/c mice were subcutaneously inoculated with mouse hepatocellular carcinoma H22 cells and treated with crude extract. Results： HPLC revealed the content of toosendanin was about 15%. Crude extract from Melia toosendan Sieb. et Zucc inhibited cancer cells growth in a dose- and time-dependent manner. The 50% inhibitory concentration （IC50, 72 h） was 0.6 mg/L for SMMC-7721 cells and 0.8 mg/L for Hep3B cells. Both high-dose [0.69 mg/（kg·d）] and low-dose [0.138 mg/（kg·d）] crude extract could markedly suppress cancer growth, and the inhibition rate was greater than 50%. Hematoxylin and eosin staining showed necrotic area in cancers and transmission electron microscopy displayed necrotic and apoptotic cancer cells with apoptotic bodies. Immunohistochemistry showed that the expression of Bax and Fas increased and the expression of Bcl-2 reduced. Conclusions： Toosendanin extract has potent anti-cancer effects via suppressing proliferation and inducing apoptosis of cancer cells in vivo and in vitro. The mechanism of apoptosis involves in mitochondrial pathway and death receptor pathway.

关键词： crude extract Melia toosendan Sieb. et Zucc anti-cancer activity SMMC-7721 cell Hep3B cell murine hepatocellular carcinoma

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Computational Prediction of Rice (Oryza sativa) miRNA Targets

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Genomics, Proteomics & Bioinformatics 2007年第3期5卷 196-206页

作者： Sunil Archak J.Nagaraju laboratory of Molecular Genetics Centre for DNA Fingerprinting and DiagnosticsHyderabad 500076India.

Bioinformatic approaches have complemented experimental efforts to inventorize plant miRNA targets. We carried out global computational analysis of rice （Oryza sativa） transcriptome to generate a comprehensive list of putative miRNA targets. Our predictions （684 unique transcripts） showed that rice miRNAs mediate regulation of diverse functions including transcription （41%）, catalysis （28%）, binding （18%）, and transporter activity （11%）. Among the predicted targets, 61.7% hits were in coding regions and nearly 72% targets had a solitary miRNA hit. The study predicted more than 70 novel targets of 34 miRNAs putatively regulating functions like stress-response, catalysis, and binding. It was observed that more than half （55%） of the targets were conserved between O. sativa indica and O. sativa japonica. Members of 31 miRNA families were found to possess conserved targets between rice and at least one of other grass family members. About 44% of the unique targets were common between two dissimilar miRNA prediction algorithms. Such an extent of cross-species conservation and algorithmic consensus confers confidence in the list of rice miRNA targets predicted in this study.

关键词： miRNA, target prediction conservation consensus, rice

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Trash to treasure:lactate and protein lactylation in maize root impacts response to drought

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Science China(Life Sciences) 2023年第8期66卷 1903-1914页

作者： Zi Shi Miaoyi Zhou Wei Song Ya Liu Ronghuan Wang Yuandong Wang Ruyang Zhang Jiuran Zhao Wen Ren Beijing Key laboratory of Maize dna Fingerprinting and Molecular Breeding Beijing Academy of Agriculture and Forestry SciencesBeijing 100097China

Lactate,protein lactylation(Kla),and specifically histone lactylation have recently been shown to regulate antipathogenic immune responses in ***,after we confirmed the presence and accumulation of lactate in maize roots under drought conditions,a lactylome profiling analysis revealed that Kla modifications were invariably present in maize roots,that there were obvious differences in the lactylomes of drought-sensitive(B73)***-tolerant(Jing2416)lines,and that growing Jing2416 under drought conditions caused significant decreases in the lactylation of multiple enzymes responsible for fatty acid *** by findings of histone-Kla based epigenetic regulation of immune functions in animals,we initially discovered 37 Kla sites on 16 histones in the maize genome,and again detected obvious differential histone Klamediated trends between two lines by Ch ***,only 2.7%of genes with differential histone Kla peaks detected during drought stress were commonly present in both lines,a finding demonstrating that abiotic stress triggers distinct epigenetic activities in diverse germplasm while also strongly supporting that a histone Kla layer of regulation is associated with physiological responses to drought ***,exogenous application of spermidine improved the drought tolerance of B73and substantially altered the levels of lactate,protein lactylation,and histone Kla ***,beyond extending the known domain of Kla-based biochemical and epigenetic regulation from animal immunity to plant stress physiology,our study suggests the physiological,biochemical,and genetic function of“the best-known metabolic waste”,lactate.

关键词： lactate lactylome drought spermidine maize

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Discriminated sgRNAs-Based SurroGate System Greatly Enhances the Screening Efficiency of Plant Base-Edited Cells

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Molecular Plant 2020年第1期13卷 169-180页

作者： Wen Xu Yongxing Yang Ya Liu Guiting Kang Feipeng Wang Lu Li Xinxin Lv Si Zhao Shuang Yuan Jinling Song Ying Wu Feng Feng Xiaoqing He Chengwei Zhang Wei Song Jiuran Zhao Jinxiao Yang Beijing Key laboratory of Maize dna Fingerprinting and Molecular Breeding Beijing Academy of Agriculture&Forestry SciencesBeijing 100080China

The development of CRISPR/Cas9-mediated base editing has made genomic modification more efficient. However, selection of genetically modified cells from millions of treated cells, especially plant cells, is still challenging. In this study, an efficient surrogate reporter system based on a defective hygromycin resistance gene was established in rice to enrich base-edited cells. After step-by-step optimization, the Discriminated sgRNAs-based SurroGate system (DisSUGs) was established by artificially differentiating the editing abilities of a wild-type single guide RNA (sgRNA) targeting the surrogate reporter gene and an enhanced sgRNA targeting endogenous sites. The DisSUGs enhanced the efficiency of screening base-edited cells by 3- to 5-fold for a PmCDA1-based cytosine-to-tyrosine base editor (PCBE), and 2.5- to 6.5-fold for an adenine base editor (ABE) at endogenous targets. These targets showed editing efficiencies of <25% in the conventional systems. The DisSUGs greatly enhanced the frequency of homozygous substitutions and expanded the activity window slightly for both a PCBE and an ABE. Analyses of the total number of single-nucleotide variants from whole-genome sequencing revealed that, compared with the no-enrichment PCBE strategy, the DisSUGs did not alter the frequency of genome-wide sgRNA-independent off-target mutations, but slightly increased the frequency of target-dependent off-target mutations. Collectively, the DisSUGs developed in this study greatly enhances the efficiency of screening plant base-edited cells and will be a useful system in future applications.

关键词： cell enrichment surrogate reporter cytosine base editors adenine base editors

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