pCtB1 载体 (V008125)

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:
pCtB1
载体抗性:
Kanamycin
载体长度:
5457 bp
载体类型:
Yeast dual vector
复制子:
ori
宿主:
Yeast
载体来源:
Lee WY, Chung S-C., Song J-Y., Cho KM.
启动子:
URA3

pCtB1 载体图谱

pCtB15457 bp600120018002400300036004200480054002u oriM13 fwdT7 promoterMCSHATPS1 terminator; interiorURA3 promoterURA3CYC1 terminatorTPS1 terminator; lateralT3 promoterM13 revlac operatorlac promoterCAP binding siteoriNeoR/KanRAmpR 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。

pCtB1 载体序列

LOCUS       40924_13755        5457 bp DNA     circular SYN 17-DEC-2018
DEFINITION  Yeast dual vector pCtB1, complete sequence.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5457)
  AUTHORS   Lee WY, Chung S-C., Song J-Y., Cho KM.
  TITLE     Direct Submission
  JOURNAL   Submitted (02-JUN-2014) BioMaterial Lab, Samsung Advanced Institute 
            of Technology, 130 Samsung-ro, Suwon 443803, Republic of Korea
REFERENCE   2  (bases 1 to 5457)
  TITLE     Direct Submission
REFERENCE   3  (bases 1 to 5457)
  AUTHORS   .
  TITLE     Direct Submission
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Submitted 
            (02-JUN-2014) BioMaterial Lab, Samsung Advanced Institute of 
            Technology, 130 Samsung-ro, Suwon 443803, Republic of Korea"
COMMENT     SGRef: number: 2; type: "Journal Article"
COMMENT     ##Assembly-Data-START##
            Sequencing Technology :: Sanger dideoxy sequencing 
            ##Assembly-Data-END##
FEATURES             Location/Qualifiers
     source          1..5457
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      134..1298
                     /label=2u ori
                     /note="yeast 2u plasmid origin of replication"
     primer_bind     1356..1372
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     promoter        1379..1397
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     misc_feature    complement(1406..1513)
                     /label=MCS
                     /note="pBluescript multiple cloning site"
     CDS             1523..1549
                     /label=HA
                     /note="HA (human influenza hemagglutinin) epitope tag"
     regulatory      1553..1799
                     /note="TPS1 terminator; interior"
                     /regulatory_class="terminator"
     promoter        1821..2036
                     /label=URA3 promoter
     CDS             2037..2837
                     /label=URA3
                     /note="orotidine-5'-phosphate decarboxylase, required for
                     uracil biosynthesis"
     terminator      2841..3088
                     /label=CYC1 terminator
                     /note="transcription terminator for CYC1"
     regulatory      3089..3301
                     /note="TPS1 terminator; lateral"
                     /regulatory_class="terminator"
     promoter        complement(3315..3333)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(3354..3370)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    complement(3378..3394)
                     /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        complement(3402..3432)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    complement(3447..3468)
                     /label=CAP binding site
                     /note="CAP binding activates transcription in the presence
                     of cAMP."
     rep_origin      complement(3756..4344)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     CDS             complement(4517..5305)
                     /label=NeoR/KanR
                     /note="aminoglycoside phosphotransferase"
     promoter        complement(5330..5434)
                     /label=AmpR promoter