pGAL1PNiST-1 载体 (V006115)

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:
pGAL1PNiST-1
载体抗性:
Ampicillin
载体长度:
7176 bp
载体类型:
Yeast expression vector
复制子:
ori
载体来源:
De Schamphelaire W, Olbrechts A, Meert J, Verhelst K, Roggeman Fonseca M, Vanhoucke M, Beyaert R.

pGAL1PNiST-1 载体图谱

pGAL1PNiST-17176 bp30060090012001500180021002400270030003300360039004200450048005100540057006000630066006900SP6 promoterM13 revlac operatorlac promoterCAP binding siteoriAmpRAmpR promoterf1 oriM13 fwdT7 promoterOrotidine 5'-phosphate decarboxylaseCaGAL1 promoter3' UTR of hIFNbetaInterferon betaCYC1 terminator

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。

pGAL1PNiST-1 载体序列

LOCUS       V006115                 7176 bp    DNA     circular SYN 18-DEC-2018
DEFINITION  Exported.
ACCESSION   V006115
VERSION     V006115
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
            .
REFERENCE   1  (bases 1 to 7176)
  AUTHORS   De Schamphelaire W, Olbrechts A, Meert J, Verhelst K, Roggeman
            Fonseca M, Vanhoucke M, Beyaert R.
  TITLE     BCCM/LMBP Plasmid collection
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 7176)
  AUTHORS   De Schamphelaire W.
  TITLE     Direct Submission
  JOURNAL   Submitted (02-FEB-2017) BCCM/LMBP, Universiteit Gent,
            Technologiepark 927, 9052, BELGIUM
REFERENCE   3  (bases 1 to 7176)
  TITLE     Direct Submission
REFERENCE   4  (bases 1 to 7176)
  AUTHORS   .
  TITLE     Direct Submission
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName:
            "Unpublished"
            SGRef: number: 2; type: "Journal Article"; journalName: "Submitted
            (02-FEB-2017) BCCM/LMBP, Universiteit Gent, Technologiepark 927,
            9052, BELGIUM"
            SGRef: number: 3; type: "Journal Article"
FEATURES             Location/Qualifiers
     source          1..7176
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        complement(10..28)
                     /label="SP6 promoter"
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     complement(46..62)
                     /label="M13 rev"
                     /note="common sequencing primer, one of multiple similar
                     variants"
     protein_bind    complement(70..86)
                     /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(94..124)
                     /label="lac promoter"
                     /note="promoter for the E. coli lac operon"
     protein_bind    complement(139..160)
                     /label="CAP binding site"
                     /note="CAP binding activates transcription in the presence
                     of cAMP."
     rep_origin      complement(448..1036)
                     /direction=LEFT
                     /label="ori"
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
                     replication"
     CDS             complement(1210..2067)
                     /label="AmpR"
                     /note="beta-lactamase"
     promoter        complement(2068..2172)
                     /label="AmpR promoter"
     rep_origin      complement(2506..2961)
                     /direction=LEFT
                     /label="f1 ori"
                     /note="f1 bacteriophage origin of replication; arrow
                     indicates direction of (+) strand synthesis"
     primer_bind     3102..3118
                     /label="M13 fwd"
                     /note="common sequencing primer, one of multiple similar
                     variants"
     promoter        3125..3143
                     /label="T7 promoter"
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             3692..4501
                     /gene="URA3"
                     /label="Orotidine 5'-phosphate decarboxylase"
                     /note="Orotidine 5'-phosphate decarboxylase from Candida
                     albicans (strain SC5314 / ATCC MYA-2876). Accession#:
                     P13649"
     regulatory      4799..6011
                     /label="CaGAL1 promoter"
                     /note="CaGAL1 promoter"
                     /regulatory_class="promoter"
     misc_feature    complement(6069..6268)
                     /label="3' UTR of hIFNbeta"
                     /note="3' UTR of hIFNbeta"
     CDS             complement(6272..6832)
                     /gene="IFNB1"
                     /label="Interferon beta"
                     /note="Interferon beta from Homo sapiens. Accession#:
                     P01574"
     terminator      6926..7173
                     /label="CYC1 terminator"
                     /note="transcription terminator for CYC1"