pAAV-RC 载体 (V009666)

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:
pAAV-RC
载体抗性:
Ampicillin
载体长度:
7327 bp
载体类型:
Cloning vector
复制子:
ori
载体来源:
Grafsky AJ III.
启动子:
lac

pAAV-RC 载体载体图谱

pAAV-RC7327 bp300600900120015001800210024002700300033003600390042004500480051005400570060006300660069007200SK primerFRTProtein Rep78Capsid protein VP1M13 fwdf1 oriAmpR promoterAmpRoriCAP binding sitelac promoterlac operatorM13 rev

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。

pAAV-RC 载体载体序列

LOCUS       V009666                 7327 bp    DNA     circular SYN 17-DEC-2018
DEFINITION  Exported.
ACCESSION   V009666
VERSION     V009666
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
            .
REFERENCE   1  (bases 1 to 7327)
  AUTHORS   Grafsky AJ III.
  TITLE     pAAV-RC for AAV Helper-Free System
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 7327)
  AUTHORS   Grafsky AJ III.
  TITLE     Direct Submission
  JOURNAL   Submitted (16-APR-2001) Technical Services, Stratagene, 11011 N.
            Torrey Pines Rd., La Jolla, CA 92037, USA
REFERENCE   3  (bases 1 to 7327)
  TITLE     Direct Submission
REFERENCE   4  (bases 1 to 7327)
  AUTHORS   .
  TITLE     Direct Submission
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName:
            "Unpublished"
            SGRef: number: 2; type: "Journal Article"; journalName: "Submitted
            (16-APR-2001) Technical Services, Stratagene, 11011 N. Torrey Pines
            Rd., La Jolla, CA 92037, USA"
            SGRef: number: 3; type: "Journal Article"
FEATURES             Location/Qualifiers
     source          1..7327
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     36..52
                     /label="SK primer"
                     /note="common sequencing primer, one of multiple similar
                     variants"
     protein_bind    66..113
                     /label="FRT"
                     /note="FLP-mediated recombination occurs in the 8-bp core
                     sequence TCTAGAAA (Turan and Bode, 2011)."
     CDS             131..1993
                     /gene="Rep78"
                     /label="Protein Rep78"
                     /note="Protein Rep78 from Adeno-associated virus 2 (isolate
                     Srivastava/1982). Accession#: Q89268"
     CDS             2013..4217
                     /gene="VP1"
                     /label="Capsid protein VP1"
                     /note="Capsid protein VP1 from Adeno-associated virus 2
                     (isolate Srivastava/1982). Accession#: P03135"
     primer_bind     complement(4548..4564)
                     /label="M13 fwd"
                     /note="common sequencing primer, one of multiple similar
                     variants"
     rep_origin      complement(4705..5160)
                     /direction=LEFT
                     /label="f1 ori"
                     /note="f1 bacteriophage origin of replication; arrow
                     indicates direction of (+) strand synthesis"
     promoter        5187..5291
                     /label="AmpR promoter"
     CDS             5292..6149
                     /label="AmpR"
                     /note="beta-lactamase"
     rep_origin      6323..6911
                     /label="ori"
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
                     replication"
     protein_bind    7199..7220
                     /label="CAP binding site"
                     /note="CAP binding activates transcription in the presence
                     of cAMP."
     promoter        7235..7265
                     /label="lac promoter"
                     /note="promoter for the E. coli lac operon"
     protein_bind    7273..7289
                     /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)."
     primer_bind     7297..7313
                     /label="M13 rev"
                     /note="common sequencing primer, one of multiple similar
                     variants"