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:
- pUB-Cas9-@BAR
- 载体抗性:
- Kanamycin
- 载体长度:
- 14835 bp
- 载体类型:
- Binary vector
- 复制子:
- ori
- 宿主:
- Plants
- 载体来源:
- Hahn F, Mantegazza O, Greiner A, Hegemann P, Eisenhut M, Weber AP.
- 启动子:
- CaMV 35S (enhanced)
pUB-Cas9-@BAR 载体图谱
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。
pUB-Cas9-@BAR 载体序列
LOCUS 40924_44784 14835 bp DNA circular SYN 18-DEC-2018
DEFINITION Binary vector pUB-Cas9-@BAR, complete sequence.
ACCESSION .
VERSION .
KEYWORDS .
SOURCE synthetic DNA construct
ORGANISM synthetic DNA construct
REFERENCE 1 (bases 1 to 14835)
AUTHORS Hahn F, Mantegazza O, Greiner A, Hegemann P, Eisenhut M, Weber AP.
TITLE An Efficient Visual Screen for CRISPR/Cas9 Activity in Arabidopsis
thaliana
JOURNAL Front Plant Sci 8, 39 (2017)
PUBMED 28174584
REFERENCE 2 (bases 1 to 14835)
AUTHORS Hahn F, Mantegazza O, Eisenhut M, Greiner A, Hegemann P, Weber APM.
TITLE Direct Submission
JOURNAL Submitted (03-NOV-2016) Plant Biochemistry, Heinrich Heine
University Dusseldorf, Universitatsstr. 1, Dusseldorf 40225, Germany
REFERENCE 3 (bases 1 to 14835)
TITLE Direct Submission
REFERENCE 4 (bases 1 to 14835)
AUTHORS .
TITLE Direct Submission
COMMENT SGRef: number: 1; type: "Journal Article"; journalName: "Front Plant
Sci 8, 39 (2017)"
COMMENT SGRef: number: 2; type: "Journal Article"; journalName: "Submitted
(03-NOV-2016) Plant Biochemistry, Heinrich Heine University
Dusseldorf, Universitatsstr. 1, Dusseldorf 40225, Germany"
COMMENT SGRef: number: 3; type: "Journal Article"
FEATURES Location/Qualifiers
source 1..14835
/mol_type="other DNA"
/organism="synthetic DNA construct"
polyA_signal 40..214
/label=CaMV poly(A) signal
/note="cauliflower mosaic virus polyadenylation signal"
misc_feature complement(292..316)
/label=LB T-DNA repeat
/note="left border repeat from nopaline C58 T-DNA"
CDS 741..1532
/label=KanR
/note="aminoglycoside phosphotransferase"
rep_origin 1622..2210
/label=ori
/note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
replication"
misc_feature complement(2396..2536)
/label=bom
/note="basis of mobility region from pBR322"
rep_origin complement(2880..3074)
/direction=LEFT
/label=pVS1 oriV
/note="origin of replication for the Pseudomonas plasmid
pVS1 (Heeb et al., 2000)"
CDS complement(3143..4207)
/label=pVS1 RepA
/note="replication protein from the Pseudomonas plasmid
pVS1 (Heeb et al., 2000)"
CDS complement(4644..5270)
/label=pVS1 StaA
/note="stability protein from the Pseudomonas plasmid pVS1
(Heeb et al., 2000)"
misc_feature complement(6570..6594)
/label=RB T-DNA repeat
/note="right border repeat from nopaline C58 T-DNA"
primer_bind 6797..6813
/label=M13 fwd
/note="common sequencing primer, one of multiple similar
variants"
regulatory 6846..6855
/note="vertebrate consensus sequence for strong initiation
of translation (Kozak, 1987)"
/regulatory_class="other"
regulatory 6850..6859
/note="vertebrate consensus sequence for strong initiation
of translation (Kozak, 1987)"
/regulatory_class="other"
promoter 6903..7326
/label=AtU6-26
/note="Arabidopsis U6-26 gene promoter"
misc_feature 7336..7356
/label=bar1 protospacer sequence
/note="bar1 protospacer sequence"
misc_RNA 7357..7432
/label=gRNA scaffold
/note="guide RNA scaffold for the Streptococcus pyogenes
CRISPR/Cas9 system"
regulatory 7557..8196
/label=Arabidopsis thaliana Ubiquitin10 promoter
/note="Arabidopsis thaliana Ubiquitin10 promoter"
/regulatory_class="promoter"
protein_bind 8236..8260
/label=attB1
/note="recombination site for the Gateway(R) BP reaction"
CDS 8281..8301
/label=SV40 NLS
/note="nuclear localization signal of SV40 (simian virus
40) large T antigen"
CDS 8299..12402
/label=Cas9
/note="Cas9 (Csn1) endonuclease from the Streptococcus
pyogenes Type II CRISPR/Cas system"
protein_bind complement(12407..12431)
/label=attB2
/note="recombination site for the Gateway(R) BP reaction"
terminator 12497..12744
/label=NOS terminator
/note="nopaline synthase terminator and poly(A) signal"
primer_bind complement(12760..12776)
/label=M13 rev
/note="common sequencing primer, one of multiple similar
variants"
protein_bind 12784..12800
/label=lac operator
/bound_moiety="lac repressor encoded by lacI"
/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(12808..12838)
/label=lac promoter
/note="promoter for the E. coli lac operon"
protein_bind complement(12853..12874)
/label=CAP binding site
/note="CAP binding activates transcription in the presence
of cAMP."
promoter 13065..13742
/label=CaMV 35S promoter (enhanced)
/note="cauliflower mosaic virus 35S promoter with a
duplicated enhancer region"
CDS 13810..14832
/label=HygR
/note="aminoglycoside phosphotransferase from E. coli"