首页技术支持质粒载体

我们所展示的质粒图谱主要是从文献和开放数据库中收集而来，主要是为了方便研究工作，其中一小部分质粒进行了质量控制，可供科学家使用。

所有的产品都严格仅供科学研究使用，不能应用于临床试验，包括人体摄入、注射或外用的药理学用途。

确保质粒的关键元件正确，但是我们并不能保证实验结果。页面展示的图谱序列为理论序列，可能与测序结果不一致，请自行比对后确定是否满足要求。(如果测过序，本页面一般会提供下载)

开放数据库中的大多数载体序列都没有被完全测序。如果实际序列与参考序列的相似度超过99%，则将其视为正确。

由于科学研究是在探索未知，具有很大的不确定性，在任何情况下，我们都不承担超出质粒本身的额外经济损失或责任。

pDM4 plasmid is a suicidal vector, which needs the strain containing pir gene. It is a derivative of plasmid pNQ705, a chloramphenicol-resistant derivative of the pGP704 vector that contains the oriV replicon originally from the plasmid R6K.

载体名称:
pDM4
载体抗性:
Chloramphenicol
载体长度:
7104 bp
载体类型:
Cloning vector
复制子:
R6K γ ori
载体来源:
Mou X, Spinard EJ, Driscoll MV, Zhao W, Nelson DR.
筛选标记:
SacB
拷贝数:
Low copy number
启动子:
sacB
感受态:
S17-1λpir
培养温度:
37℃

pDM4 载体图谱

pDM47104 bp30060090012001500180021002400270030003300360039004200450048005100540057006000630066006900oriTtraJconjugal transfer relaxase TraIlac operatorcat promoterCmRsacB promoterSacBinsertion element IS1 1/5/6 protein insBbeta-lactamase

质粒操作方法

1. 发货形式：质粒干粉(常温运输，存于-20度，请务必先转化提质粒后使用)

2. 收到质粒干粉后请先5000rpm离心1min，再加入20μl ddH2O溶解质粒;（质粒复测的浓度有时候与标称值差距较大，这可能是因为冻干质粒在管中的位置、复溶效率、测量偏差以及管壁的吸附导致，因此建议先转化提质粒后再使用）

3. 取1支100μl 感受态于冰上解冻10min，加入2μl质粒，再冰浴30min后，42℃热激60s，不要搅动，再冰浴2min；

4. 加入900μl无抗的LB液体培养基，180rpm震荡37℃培养45min (30℃培养1-1.5小时);

5. 6000rpm离心5min，仅留100μl上清液重悬细菌沉淀，并涂布至目标质粒抗性的LB平板上;

6. 将平板倒置37℃培养14h，如果要求是30℃则培养20h; (菌落过多则将质粒稀释后再转化；没有菌落则加入10μl质粒转化；建议不要直接转表达感受态， 要先转克隆感受态，重提质粒后再导入表达感受态);

7. 挑取单菌落至LB液体培养基中，加入对应抗生素，220rpm震荡培养14h，根据实验需要和质粒提取试剂盒说明书提取质粒。

pDM4 载体序列

LOCUS       40924_14860        7104 bp DNA     circular SYN 18-DEC-2018
DEFINITION  Cloning vector pDM4, complete sequence.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7104)
  AUTHORS   Mou X, Spinard EJ, Driscoll MV, Zhao W, Nelson DR.
  TITLE     H-NS Is a Negative Regulator of the Two Hemolysin/Cytotoxin Gene 
            Clusters in Vibrio anguillarum
  JOURNAL   Infect. Immun. 81 (10), 3566-3576 (2013)
  PUBMED    23836825
REFERENCE   2  (bases 1 to 7104)
  AUTHORS   Mou X, Spinard EJ, Driscoll MV, Zhao W, Nelson DR.
  TITLE     Direct Submission
  JOURNAL   Submitted (19-MAR-2013) Cell and Molecular Biology, University of 
            Rhode Island, 120 Flagg Rd, Kingston, RI 02881, USA
REFERENCE   3  (bases 1 to 7104)
  TITLE     Direct Submission
REFERENCE   4  (bases 1 to 7104)
  AUTHORS   .
  TITLE     Direct Submission
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Infect. 
            Immun."; date: "2013"; volume: "81"; issue: "10"; pages: "3566-3576"
COMMENT     SGRef: number: 2; type: "Journal Article"; journalName: "Submitted 
            (19-MAR-2013) Cell and Molecular Biology, University of Rhode 
            Island, 120 Flagg Rd, Kingston, RI 02881, USA"
COMMENT     SGRef: number: 3; type: "Journal Article"
COMMENT     ##Assembly-Data-START##
            Sequencing Technology :: Sanger dideoxy sequencing 
            ##Assembly-Data-END##
FEATURES             Location/Qualifiers
     source          1..7104
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     oriT            623..732
                     /label=oriT
                     /note="incP origin of transfer"
     CDS             765..1133
                     /codon_start=1
                     /label=traJ
                     /note="oriT-recognizing protein"
                     /translation="MADETKPTRKGSPPIKVYCLPDERRAIEEKAAAAGMSLSAYLLAV
                     GQGYKITGVVDYEHVRELARINGDLGRLGGLLKLWLTDDPRTARFGDATILALLAKIEE
                     KQDELGKVMMGVVRPRAEP"
     CDS             1192..1818
                     /codon_start=1
                     /product="conjugal transfer relaxase TraI"
                     /label=conjugal transfer relaxase TraI
                     /protein_id="AGO64135.1"
                     /translation="MRSIKKSDFAELVKYITDEQGKTERLGHVRVTNCEANTLPAVMAE
                     VMATQHGNTRSEADKTYHLLVSFRAGEKPDAETLRAIEDRICAGLGFAEHQRVSAVHHD
                     TDNLHIHIAINKIHPTRNTIHEPYRAYRALADLCATLERDYGLERDNHETRQRVSENRA
                     NDMERHAGVESLVGWIRPRCVRRRGSEDQQFNLLIVRTKLSCFTY"
     rep_origin      1754..2142
                     /label=R6K gamma ori
                     /note="gamma replication origin from E. coli plasmid R6K;
                     requires the R6K initiator protein pi for replication"
     protein_bind    2177..2193
                     /label=lac operator
                     /note="The lac repressor binds to the lac operator to
                     inhibit transcription in E. coli. This inhibition can be 
                     relieved by adding lactose or 
                     isopropyl-beta-D-thiogalactopyranoside (IPTG)."
     promoter        2574..2676
                     /label=cat promoter
                     /note="promoter of the E. coli cat gene encoding
                     chloramphenicol acetyltransferase"
     CDS             2677..3333
                     /codon_start=1
                     /label=CmR
                     /note="chloramphenicol acetyltransferase"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA"
     promoter        3753..4198
                     /label=sacB promoter
                     /note="sacB promoter and control region"
     CDS             4199..5617
                     /codon_start=1
                     /label=SacB
                     /note="secreted levansucrase that renders bacterial growth 
                     sensitive to sucrose"
                     /translation="MNIKKFAKQATVLTFTTALLAGGATQAFAKETNQKPYKETYGISH
                     ITRHDMLQIPEQQKNEKYQVPEFDSSTIKNISSAKGLDVWDSWPLQNADGTVANYHGYH
                     IVFALAGDPKNADDTSIYMFYQKVGETSIDSWKNAGRVFKDSDKFDANDSILKDQTQEW
                     SGSATFTSDGKIRLFYTDFSGKHYGKQTLTTAQVNVSASDSSLNINGVEDYKSIFDGDG
                     KTYQNVQQFIDEGNYSSGDNHTLRDPHYVEDKGHKYLVFEANTGTEDGYQGEESLFNKA
                     YYGKSTSFFRQESQKLLQSDKKRTAELANGALGMIELNDDYTLKKVMKPLIASNTVTDE
                     IERANVFKMNGKWYLFTDSRGSKMTIDGITSNDIYMLGYVSNSLTGPYKPLNKTGLVLK
                     MDLDPNDVTFTYSHFAVPQAKGNNVVITSYMTNRGFYADKQSTFAPSFLLNIKGKKTSV
                     VKDSILEQGQLTVNK"
     CDS             complement(5758..6135)
                     /codon_start=1
                     /product="insertion element IS1 1/5/6 protein insB"
                     /label=insertion element IS1 1/5/6 protein insB
                     /protein_id="AGO64138.1"
                     /translation="MDEQWGYVGAKSRQRWLFYAYDSLRKTVVAHVFGERTMATLGRLM
                     SLLSPFDVVIWMTDGWPLYESRLKGKLHVISKRYTQRIERHNLNLRQHLARLGRKSLSF
                     SKSVELHDKVIGHYLNIKHYQ"
     CDS             6413..6649
                     /codon_start=1
                     /product="beta-lactamase"
                     /EC_number="3.5.2.6"
                     /label=beta-lactamase
                     /protein_id="AGO64139.1"
                     /translation="MEADKVAGPLLRSALPAGWFIADKSGAGERGSRGIIAALGPDGKP
                     SRIVVIYTTGSQATMDERNRQIAEIGASLIKHW"