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pCMV-Red Firefly Luc 质粒 (编号: V011552)

Note: pCMV-Red Firefly Luc is a mammalian reporter gene vector with CMV promoter, red-shifted firefly luciferase (emission peak 610-617 nm), for high-sensitivity bioluminescence detection and in vivo imaging.

pCMV-Red Firefly Luc6574 bp300600900120015001800210024002700300033003600390042004500480051005400570060006300CMV enhancerCMV promoterT7 promoterRed Firefly LuciferasebGH poly(A) signalSV40 promoterEM7 promoterPuroRSV40 poly(A) signalAmpRoriM13 fwdlac operatorpause site
复制序列 NovoBuilder中查看图谱

基本信息

载体名称:
pCMV-Red Firefly Luc
载体抗性:
Ampicillin
载体长度:
6574 bp
载体类型:
Luciferase Vectors
复制子:
ori
载体来源:
Thermo Scientific (Pierce)
拷贝数:
High copy number
启动子:
SV40
感受态:
DH5a
培养温度:
37℃

产品信息

质粒编号:V011552
质粒名称:pCMV-Red Firefly Luc
规格:5 μg (冻干粉)

下载资源

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，供学术研究使用。

确保质粒的关键元件正确，但是我们并不能保证实验效果。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。

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。

参考文献

  • Branchini BR, Ablamsky DM, Davis AL, et al. Red-emitting luciferases for bioluminescence reporter and imaging applications. Analytical Biochemistry. 2010;396(2):290-297. doi: 10.1016/j.ab.2009.09.009

  • Caysa H, Jacob R, Müther N, Branchini B, Messerle M, Söling A. A redshifted codon-optimized firefly luciferase is a sensitive reporter for bioluminescence imaging. Photochemical & Photobiological Sciences. 2009;8(1):52-56. doi: 10.1039/b814566k

  • Branchini BR, Southworth TL, Fontaine DM, Kohrt D, Florentine CM, Grossel MJ. A Firefly Luciferase Dual Color Bioluminescence Reporter Assay Using Two Substrates To Simultaneously Monitor Two Gene Expression Events. Scientific Reports. 2018;8(1). doi: 10.1038/s41598-018-24278-2

pCMV-Red Firefly Luc 质粒 (编号: V011552)序列

LOCUS       pCMV-Red-Firefly        6574 bp    DNA     circular SYN 08-APR-2026
DEFINITION  Control vector for constitutive high-level expression of 
            intracellular red firefly luciferase under control of the CMV 
            promoter.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6574)
  AUTHORS   Thermo Scientific (Pierce)
  TITLE     Direct Submission
REFERENCE   2  (bases 1 to 6574)
  TITLE     Direct Submission
REFERENCE   3  (bases 1 to 6574)
  AUTHORS   .
  TITLE     Direct Submission
COMMENT     SGRef: number: 1; type: "Journal Article"
COMMENT     SGRef: number: 2; type: "Journal Article"
FEATURES             Location/Qualifiers
     source          1..6574
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        63..366
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        367..570
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early
                     promoter"
     promoter        615..633
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             646..2289
                     /codon_start=1
                     /label=Red Firefly Luciferase
                     /note="intracellular red firefly luciferase"
                     /translation="MENMENDENIVVGPKPFYPIEEGSAGTQLRKYMERYAKLGAIAFT
                     NAVTGVDYSYAEYLEKSCCLGKALQNYGLVVDGRIALCSENCEEFFIPVIAGLFIGVGV
                     APTNEIYTLRELVHSLGISKPTIVFSSKKGLDKVITVQKTVTTIKTIVILDSKVDYRGY
                     QCLDTFIKRNTPPGFQASSFKTVEVDRKEQVALIMNSSGSTGLPKGVQLTHENTVTRFS
                     HARDPIYGNQVSPGTAVLTVVPFHHGFGMFTTLGYLICGFRVVMLTKFDEETFLKTLQD
                     YKCTYVILVPTLFAILNKSELLNKYDLSNLVEIASGGAPLSKEVGEAVARRFNLPGVRQ
                     GYGLTETTSAIIITPEGDDKPGASGKVVPLFKAKVIDLDTKKSLGPNRRGEVCVKGPML
                     MKGYVNNPEATKELIDEEGWLHTGDIGYYDEEKHFFIVDRLKSLIKYKGYQVPPAELES
                     VLLQHPSIFDAGVAGVPDPVAGELPGAVVVLESGKNMTEKEVMDYVASQVSNAKRLRGG
                     VRFVDEVPKGLTGKIDGRAIREILKKPVAKM"
     polyA_signal    2318..2429
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     promoter        2552..2881
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     promoter        2929..2976
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter"
     CDS             2995..3591
                     /codon_start=1
                     /label=PuroR
                     /note="puromycin N-acetyltransferase"
                     /translation="MTEYKPTVRLATRDDVPRAVRTLAAAFADYPATRHTVDPDRHIER
                     VTELQELFLTRVGLDIGKVWVADDGAAVAVWTTPESVEAGAVFAEIGPRMAELSGSRLA
                     AQQQMEGLLAPHRPKEPAWFLATVGVSPDHQGKGLGSAVVLPGVEAAERAGVPAFLETS
                     APRNLPFYERLGFTVTADVEVPEGPRTWCMTRKPGA"
     polyA_signal    3724..3857
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     CDS             complement(3901..4758)
                     /codon_start=1
                     /label=AmpR
                     /note="beta-lactamase"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     rep_origin      5022..5610
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5880..5896
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    6352..6368
                     /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)."
     misc_feature    6483..6574
                     /label=pause site
                     /note="RNA polymerase II transcriptional pause signal from
                     the human alpha-2 globin gene"