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An efficient and safe strategy for germ cell-specific automatic excision of foreign DNA in F1 hybrid transgenic silkworms

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Insect Science 2024年第1期31卷 28-46页

作者： Dingpei Long Rongpeng Liu Yang Huang Anyao Fu Yuli Zhang Zhanzhang Hao Qiang Li Hanfu Xu Zhonghuai Xiang Aichun Zhao State Key Laboratory of Resource Insects Institute of Sericulture and Systems BiologySouthwest UniversityChongqingChina department of Biology Georgia State UniversityAtlantaGeorgiaUSA Guangxi Institute of sericulture Science NanningGuangxiChina department of Mathematics and Statistics Georgia State UniversityAtlantaGeorgiaUSA

The safety of transgenic technology is a major obstacle in the popularization and use of transgenic silkworms and their products.In sericulture,only the first filial generation(F1)hybrid eggs produced by cross-breeding Japanese and Chinese original strains are usually used for the large-scale breeding of silkworms,but this may result in uncontrolled transgene dispersal during the popularization and application of the F1 hybrid transgenic eggs.To address this issue,we developed a safe and efficient strategy using the GAL4/Upstream activating sequence(UAS)system,the FLP/flippase recognition target(FRT)system,and the gonad-specific expression gene promoters(RSHP1p and Nanosp)for the germ cell-specific automatic excision of foreign DNA in the F1 hybrid transgenic silkworms.We established 2 types of activator strains,R1p::GAL4-Gr and Nsp::GAL4-Gr,containing the testis-specific GAL4 gene expression cassettes driven by RSHP1p or Nanosp,respectively,and 1 type of effector strain,UAS::FLP-Rg,containing the UAS-linked FLP gene expression cassette.The FLP recombinase-mediated sperm-specific complete excision of FRT-flanked target DNA in the F1 double-transgenic silkworms resulting from the hybridization of R1p::GAL4-Gr and UAS::FLP-Rg was 100%,whereas the complete excision efficiency resulting from the hybridization of Nsp::GAL4-Gr and UAS::FLP-Rg ranged from 13.73%to 80.3%.Additionally,we identified a gene,sw11114,that is expressed in both testis and ovary of Bombyx mori,and can be used to establish novel gonad-specific expression systems in transgenic silkworms.This strategy has the potential to fundamentally solve the safety issue in the production of F1 transgenic silkworm eggs and provides an important reference for the safety of transgenic technology in other insect species.

关键词： Bombyx mori FLP/FRT system GAL4/UAS system germ cell-specific excision safety of transgenic technology transgenic silkworm

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Mulberry leaf supplementation inhibits skatole deposition by regulating gut microbiota and upregulating liver cytochrome P4501A1 expression in finishing pigs

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Animal Nutrition 2024年第1期16卷 34-44页

作者： Yuqing Sun Xiaoming Men Tianbao Lin Bo Deng Shi Zhong Jinxi Huo Kaipeng Qin Zhiqiang Lv Ziwei Xu Yougui Li Institute of sericulture and Tea Zhejiang Academy of Agricultural Sciences Institute of Animal Husbandry and Veterinary Science Zhejiang Academy of Agricultural Sciences

Skatole, a strong fecal odor substance, is generated through microbial degradation of tryptophan in the animal hindgut. It easily accumulates in adipose tissue and affects meat quality. In this study, the effect of mulberry leaf supplementation on skatole in finishing pigs was studied. In a 35-day trial, 20 finishing pigs (barrows and gilts) were fed with a basal diet or basal diet with 6% mulberry leaves. Growth performance of the pigs (n=10) was automatically recorded by a performance-testing feeder system and 8 pigs in each treatment were slaughtered and sampled for the remaining tests. Skatole and short-chain fatty acids were detected using HPLC and gas chromatography, respectively. Fecal microbiota were analyzed using 16S r RNA gene sequencing. The metabolomics analysis of feces and serum was performed with UHPLC-MS/MS. The major cytochrome P450 (CYP) enzymes that catalyze skatole degradation in the liver were tested by using RT-PCR and Western blot. Effects of major bioactive compounds in mulberry leaves on the CYP genes were verified in the hepatic cell line Hep G2 in an in vitro test (n=3). In finishing pigs, mulberry leaf supplementation had no significant effect on the average daily gain, average daily feed intake, and feed conversion ratio (P>0.05), but reduced skatole levels in feces, serum, and backfat (P<0.05), and increased acetic acid levels in feces (P=0.027). Mulberry leaf supplementation decreased the relative abundance of the skatole-producing bacteria Megasphaera and Olsenella (P<0.05). Indole-3acetic acid, the intermediate that is essential for skatole production, was significantly reduced in feces by mulberry leaf supplementation (P<0.05) and was positively correlated with skatole content in feces (P=0.004). In pigs treated with mulberry leaves, liver CYP1A1 expression was increased (P<0.05) and was negatively correlated with skatole content in backfat (P=0.045). The in vitro test demonstrated that mulberry leaf polyphenols and polysaccharides could directly stimulate CYP1A1 expression in h

关键词： Pig Skatole Mulberry leaf Microbiota Cytochrome P450

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Cloning and Alternative Splicing Analysis of Bombyx mori Transformer-2 Gene using Silkworm EST Database

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Acta Biochimica et Biophysica Sinica 2005年第11期37卷 728-736页

作者： Bao-Long NIU Zhi-Qi MENG Yue-Zhi TAO Shun-Lin LU Hong-Biao WENG Li-Hua HE Wei-Feng SHEN Sericultural Research Institute,Zhejiang Academy of Agricultural Sciences,Hangzhou 310021,China department of sericulture and Apiculture,College of Animal Sciences,Zhejiang University,Hangzhou 310029,China Sericultural Research Institute Zhejiang Academy of Agricultural SciencesHangzhou 310021 China department of sericulture and Apiculture College of Animal Sciences Zhejiang UniversityHangzhou 310029 China

We have identified Bombyx mori transformer-2 gene(Bmtra-2)cDNA by blasting the ESTdatabase of B.mori.It was expressed in the whole life of the male and female silkworm and was observed asa band of 1.3 kb by Northern blot analysis.By comparing corresponding ESTs to the Bmtra-2 DNA sequence,it was revealed that there were eight exons and seven introns,and all splice sites of exons/introns conformedto the GT/AG rule.Bmtra-2 pre-mRNA can produce multiple mRNAs encoding six distinct isoforms ofBmTRA-2 protein using an alternative splicing pathway during processing.Six types of Bmtra-2 cDNAclones were identified by reverse transcription-polymerase chain reaction.All isoforms of BmTRA-2 proteincontain two arginine/serine-rich domains and one RNA recognition motif,showing striking organizationalsimilarity to Drosophila TRA-2 proteins.

关键词： EST database transformer-2 gene Bmtra-2 alternative splicing gene clone Bombyxmori

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Possible Effect of 30K Proteins in Embryonic Development of Silkworm Bombyx mori

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Acta Biochimica et Biophysica Sinica 2005年第5期37卷 355-361页

作者： Bo-Xiong ZHONG~1 Jian-Ke LI~(1.2) Jian-Rong LIN~3 Jian-She LIANG~4 Song-Kun SU~1 Hai-Sheng XU~1 Hai-Yan YAN~1 Ping-Bo ZHANG~5 Hiroshi FUJII~5 1 Laboratory of Molecular Biology of Silkworm and Honeybee, department of Bioresource Science, College of Animal Science,Zhejiang University, Hangzhau 310029, China 2 Institute of Apicultural Research, Chinese Academy of Agricultural Science, Beijing 100093, China 3 department of sericulture, South China Agricultural University.Guangzhou 510642, China 4 Biochemistry Co. Ltd. of Zhejiang University, Hangzhou 310029. China 5 Faculty of Agriculture,Kyushu University, Fukuoka 812-81. Japan Laboratory of Molecular Biology of Silkworm and Honeybee Department of Bioresource Science College of Animal Science Zhejiang UniversityHangzhou 310029 China Institute of Agicultural Research Chinese Academy of Agricultural ScienceBeijing 100093 China department of sericulture South China Agricultural UniversityGuangzhou 510642 China Biochemistry Co. Ltd. of Zhejiang UniversityHangzhou 310029 China Faculty of Agriculture Kyushu UniversityFukuoka 812-81 Japan

The silkworm Bombyx mori possesses a 30K protein family of 3×10～4 Da,the biologicalfunctions of which have not been fully identified.The relationship between the 30K protein family and theembryonic development of temperature sensitive sex-linked mutant strain of silkworm was investigated bytwo dimensional polyacrylamide gel electrophoresis(2D-PAGE)and Matrix assisted laser desorption ionization-time of flight mass spectrometry(MALDI-TOF MS).The results show that protein spots 1-5 of the 30Kprotein family,mainly existing in normal strain,are possibly related to embryonic development.The earlyconsumption of a 30K protein named 6G1-30K-1 and the accumulation of 30K proteins named 6G1-30K-3and 6G1-30K-4 are likely caused by the destruction of physiological balance in normal embryonic development,which may lead to lower hatchability of the temperature sensitive strain.The results suggest that reasonablemetabolism of 30K proteins is a prerequisite for the embryo's normal development.

关键词： silkworm Bombyx mori 30K protein embryonic development 2D-PAGE MALDI-TOF MS

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ULTRASTRUCTURE ANALYSIS OF MITOCHONDRIA OF HIND SILK GLAND IN Bombyx mori BY MEANS OF PSEUDOCOLOUR ENHANCEMENT TECHNIQUES WITH A MINICOMPUTER

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Chinese Science Bulletin 1986年第1期31卷 69-70页

作者：唐任天朱敏慧曹阳唐维六 Institute of Microbiology Academia Sinica Beijing Institute of Electronics Academia Sinica Beijing department of sericulture South China Agricultural University Guangzhou department of sericulture

The resolution of a black-white image is not as good as a colour image to the naked eyes. However, it is possible to obtain additional information from a black-white image by means of pseudocolour enhancement techniqu... 详细信息

关键词： naked transformed Bombyx colour photograph sparse irregular monitor finger sometimes

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Bioinspired Hemostatic Strategy via Pulse Ejections for Severe Bleeding Wounds

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Research 2024年第1期2023卷 281-299页

作者： Bitao Lu Enling Hu Weiwei Ding Wenyi Wang Ruiqi Xie Kun Yu Fei Lu Guangqian Lan Fangyin Dai State Key Laboratory of Silkworm Genome Biology College of SericultureTextile and Biomass SciencesSouthwest UniversityChongqing 400715China Chongqing Engineering Research Center of Biomaterial Fiber and Modern Textile Chongqing 400715China Division of Trauma and Surgical Intensive Care Unit Research Institute of General SurgeryJinling HospitalMedical School of Nanjing UniversityNanjing210002 Jiangsu ProvinceChina department of Applied Biology and Chemical Technology The Hong Kong Polytechnic UniversityHong KongChina

Efficient hemostasis during emergency trauma with massive bleeding remains a critical challenge in prehospital settings.Thus,multiple hemostatic strategies are critical for treating large bleeding wounds.In this study,inspired by bombardier beetles to eject toxic spray for defense,a shape-memory aerogel with an aligned microchannel structure was proposed,employing thrombin-carrying microparticles loaded as a built-in engine to generate pulse ejections for enhanced drug permeation.Bioinspired aerogels,after contact with blood,can rapidly expand inside the wound,offering robust physical barrier blocking,sealing the bleeding wound,and generating a spontaneous local chemical reaction causing an explosivelike generation of CO_(2) microbubbles,which provide propulsion thrust to accelerate burst ejection from arrays of microchannels for deeper and faster drug diffusion.The ejection behavior,drug release kinetics,and permeation capacity were evaluated using a theoretical model and experimentally demonstrated.This novel aerogel showed remarkable hemostatic performance in severely bleeding wounds in a swine model and demonstrated good degradability and biocompatibility,displaying great potential for clinical application in humans.

关键词： wound sealing offering

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SOME INDUCIBLE ANTIBACTERIAL SUBSTANCES DEVELOPED FROM THE HEAMOLYMPH OF OAK SILKWORM, Antheraea pernyi PUPAE BY ULTRASONIC TREATMENT

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Chinese Science Bulletin 1984年第5期29卷 670-674页

作者：祁国荣周奇屈贤铭黄自然 Shanghai Institute of Biochemistry Academia Sinica department of sericulture South China Agricultural College Guangzhou

Insects have higher adaptability and defence functions compared with mammal animals. The lysozyme and antibacterial substances can be induced from the heamolymph by injecting bacteria into the insects, such as Bombyx ... 详细信息

关键词： Antheraea pernyi Bombyx antibacterial adaptability lysozyme mammal sterile larvae belongs

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Mulberry Biomass-Derived Nanomedicines Mitigate Colitis through Improved Inflamed Mucosa Accumulation and Intestinal Microenvironment Modulation

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Research 2024年第1期6卷 791-807页

作者： Wenjing Yang Ya Ma Haiting Xu Zhenhua Zhu Jiaxue Wu Cheng Xu Wei Sun Erhu Zhao Min Wang Rui L.Reis Subhas C.Kundu Xiaoxiao Shi Bo Xiao State Key Laboratory of Silkworm Genome Biology College of SericultureTextileand Biomass SciencesSouthwest UniversityChongqing 400715China Chongqing Key Laboratory of Soft-Matter Material Chemistry and Function Manufacturing Faculty of Materials and EnergySouthwest UniversityChongqing 400715China department of Gastroenterology The First Affiliated Hospital of Nanchang UniversityNanchang 330006China Bs Research Group I3Bs—Research Institute on BiomaterialsBiodegradables and BiomimeticsUniversity of MinhoHeadquarters of the European Institute of Excellence on Tissue Engineering and Regenerative MedicineAveParkBarco 4805-017GuimaraesPortugal ICVS/3B’s-PT Government Associate Laboratory BragaGuimarãesPortugal

The therapeutic outcomes of conventional oral medications against ulcerative colitis(UC)are restricted by inefficient drug delivery to the colitis mucosa and weak capacity to modulate the inflammatory microenvironment.Herein,a fluorinated pluronic(FP127)was synthesized and employed to functionalize the surface of mulberry leaf-derived nanoparticles(MLNs)loading with resveratrol nanocrystals(RNs).The obtained FP127@RN-MLNs possessed exosome-like morphologies,desirable particle sizes(around 171.4 nm),and negatively charged surfaces(−14.8 mV).The introduction of FP127 to RN-MLNs greatly improved their stability in the colon and promoted their mucus infiltration and mucosal penetration capacities due to the unique fluorine effect.

关键词： fluorine sizes Biomass

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Analysis of culturable and unculturable microbial community in bensulfuron-methyl contaminated paddy soils

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Journal of Environmental Sciences 2008年第12期20卷 1494-1500页

作者： M.L.Umashankara department of sericulture College of YuvarajasMysore University

To investigate the influence of bensulfuron-methyl(BSM)on culturable microbial quantities and unculturable microbial community structures,conventional and molecular biological methods were employed in five BSM treated soils with three replications,respectively. The results obtained with traditional culture-dependent methods showed that a low-level of BSM had slight and transient effects on culturable microorganisms;nevertheless,high concentration of BSM resulted in a dramatic decrease in bacterial colony fo...

关键词： bensulfuron-methyl（BSM） microbial quantities community structures DGGE

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BmTBP upregulates the transcription of BmSuc1 in silkworm (Bombyx mori) by binding to BmTfΙΙA-S

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Insect Science 2023年第5期30卷 1405-1419页

作者： Xinwei Zhang Liangli Yang Quan Gan Song Jiang Dan Liang Junshan Gao Yan Meng School of Life Sciences Anhui Agricultural UniversityHefeiChina department of Pathology Henan Provincial People's HospitalZhengzhouChina Anhui International Joint Research and Development Center of sericulture Resources Utilization HefeiChina Anhui Academy of Agricultural Sciences HefeiChina

The BmSuc1 gene,which encodes a novel animal-typeβ-fructofuranosidase(EC 3.***.1.26),was first cloned and identified in silkworm(Bombyx mori).As an essential sucrase,the activity of BmSUC1 is unaffected by alkaloidal sugar mimics in mulberry leaves.This enzyme may also directly regulate the degree of sucrose hydrolysis in the silkworm midgut.In addition,BmSUC1 is involved in the synthesis of sericin 1 in the silk gland tissue.However,the mechanism underlying the regulation of BmSuc1 transcription remains unclear.In this study,we analyzed the BmSuc1 promoter activity using a dual-luciferase reporter assay and identified 4 regions that are critical for transcriptional activation.The gene encoding a predicted transcription factor(TATA-box-binding protein;BmTBP)capable of binding to the core promoter regions was cloned.A quantitative real-time polymerase chain reaction analysis indicated the gene was highly expressed in the midgut.Downregulating BmTBP expression via RNA interference decreased the expression of BmSuc1 at the transcript and protein levels.An electrophoretic mobility shift analysis and chromatin immunoprecipitation indicated that BmTBP can bind to the TATA-box cis-regulatory element in the BmSuc1 promoter.Furthermore,a bioinformatics-based analysis and a far-western blot revealed the interaction between BmTBP and another transcription factor(BmTfIIA-S).The luciferase reporter gene assay results confirmed that the BmTBP–BmTfIIA-S complex increases the BmSuc1 promoter activity.Considered together,these findings suggest that BmTBP regulates BmSuc1 expression through its interaction with BmTfIIA-S.

关键词： BmSucl BmTBP Bombyx mori transcriptional regulation

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