pCVD011 载体 (V001570)

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。

载体名称:
pCVD011
载体抗性:
Ampicillin
载体长度:
3283 bp
载体类型:
Broad host range vector
复制子:
ori
载体来源:
Taton A, Unglaub F, Wright NE, Zeng WY, Paz-Yepez J, Brahamsha B, Palenik B, Peterson TC, Haerizadeh F, Golden SS, Golden JW.

pCVD011 载体图谱

pCVD0113283 bp6001200180024003000M13 fwdT7 promoterC2G; GC-adaptor for seamless assembly, flanking antibiotic resistance markers, functional devices or custom sequences; syntheticAgeIPveg; veg promoter; derived from Bacillus subtilis, used upstream of nat1_S7942 and nat1_A7120, 3'end carries a RBS and a BamHI siteNrsRtrpA T; trpA terminator from Lorist6 INSD accession X98450, one nt shorter than reference; Lorist6NheIGC; GC-adaptor for seamless assembly, flanking antibiotic resistance markers or Escherichia coli origins to be assembled with a cyanobacterial replicon; syntheticoriAmpRAmpR promoter

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。

pCVD011 载体序列

LOCUS       40924_49067        3283 bp DNA     circular SYN 18-DEC-2018
DEFINITION  Broad host range vector vector pCVD011, complete sequence.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 3283)
  AUTHORS   Taton A, Unglaub F, Wright NE, Zeng WY, Paz-Yepez J, Brahamsha B, 
            Palenik B, Peterson TC, Haerizadeh F, Golden SS, Golden JW.
  TITLE     Broad-host-range vector system for synthetic biology and 
            biotechnology in cyanobacteria
  JOURNAL   Nucleic Acids Res. (2014) In press
  PUBMED    25074377
REFERENCE   2  (bases 1 to 3283)
  AUTHORS   Taton A, Unglaub F, Wright N, Zeng WY, Paz Yepes J, Brahamsha B, 
            Palenik B, Peterson T, Haerizadeh F, Golden SS, Golden JW.
  TITLE     Direct Submission
  JOURNAL   Submitted (17-JUN-2014) Division of Biological Sciences, University 
            of California, San Diego, 9500 Gilman Dr., La Jolla, CA 92093, USA
REFERENCE   3  (bases 1 to 3283)
  TITLE     Direct Submission
REFERENCE   4  (bases 1 to 3283)
  AUTHORS   .
  TITLE     Direct Submission
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Nucleic 
            Acids Res. (2014) In press"
COMMENT     SGRef: number: 2; type: "Journal Article"; journalName: "Submitted 
            (17-JUN-2014) Division of Biological Sciences, University of 
            California, San Diego, 9500 Gilman Dr., La Jolla, CA 92093, USA"
COMMENT     SGRef: number: 3; type: "Journal Article"
FEATURES             Location/Qualifiers
     source          1..3283
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     293..309
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     promoter        320..338
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     misc_feature    370..375
                     /label=EcoRV
                     /note="EcoRV"
     misc_feature    complement(373..393)
                     /note="C2G; GC-adaptor for seamless assembly, flanking 
                     antibiotic resistance markers, functional devices or custom
                     sequences; synthetic"
     misc_feature    394..399
                     /label=AgeI
                     /note="AgeI"
     regulatory      406..667
                     /note="Pveg; veg promoter; derived from Bacillus subtilis,
                     used upstream of nat1_S7942 and nat1_A7120, 3'end carries a
                     RBS and a BamHI site"
                     /regulatory_class="promoter"
     misc_feature    515..522
                     /label=SwaI
                     /note="SwaI"
     CDS             668..1234
                     /label=NrsR
                     /note="nourseothricin acetyltransferase"
     regulatory      1246..1272
                     /note="trpA T; trpA terminator from Lorist6 INSD accession 
                     X98450, one nt shorter than reference; Lorist6"
                     /regulatory_class="terminator"
     misc_feature    1282..1287
                     /label=NheI
                     /note="NheI"
     misc_feature    complement(1288..1308)
                     /note="GC; GC-adaptor for seamless assembly, flanking
                     antibiotic resistance markers or Escherichia coli origins 
                     to be assembled with a cyanobacterial replicon; synthetic"
     misc_feature    1306..1311
                     /label=EcoRV
                     /note="EcoRV"
     rep_origin      complement(1536..2124)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     CDS             complement(2298..3155)
                     /label=AmpR
                     /note="beta-lactamase"
     promoter        complement(3156..3260)
                     /label=AmpR promoter