基本信息
- 载体名称:
- PiggyBac Dual promoter-1600014C10Rik(mouse)-luciferase
- 载体抗性:
- Ampicillin
- Refseq ID:
- NM_001085385.1
- 载体长度:
- 9359 bp
- 载体类型:
- Protein expression, Transposition
- 复制子:
- ori
- 宿主:
- Mammalian cells
- 筛选标记:
- Puro;CopGFP
- 拷贝数:
- High
- 启动子:
- CMV
- 5'引物:
- CMV-F
- 载体标签:
- luciferase
- 感受态:
- Stbl3
- 培养温度:
- 37℃
- 表达方法:
- Transient
- 描述:
- Mus musculus RIKEN cDNA 1600014C10 gene (1600014C10Rik), transcript variant 1, mRNA
产品信息
质粒编号:V044325
质粒名称:PiggyBac Dual promoter-1600014C10Rik(mouse)-luciferase
规格:100 μg (转染级)
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，可供科学家使用。
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。
小鼠1600014C10Rik ORF 克隆 (NM_001085385) (编号: V044325)序列
LOCUS V044325 9359 bp DNA circular SYN 01-JAN-1980
DEFINITION synthetic circular DNA.
ACCESSION V044325
VERSION V044325
KEYWORDS .
SOURCE .
ORGANISM .
.
FEATURES Location/Qualifiers
promoter complement(1..105)
/gene="bla"
/label="AmpR promoter"
rep_origin complement(131..586)
/direction=LEFT
/note="f1 bacteriophage origin of replication; arrow
indicates direction of (+) strand synthesis"
/label="f1 ori"
primer_bind 728..744
/note="common sequencing primer, one of multiple similar
variants"
/label="M13 fwd"
promoter 1442..1645
/note="human cytomegalovirus (CMV) immediate early
promoter"
/label="CMV promoter"
CDS 1734..2156
/label="1600014C10Rik(NM_001085385)"
/note="1600014C10Rik(NM_001085385)"
/gene="1600014C10Rik"
CDS 2193..3845
/EC_number=""
/codon_start=1
/gene="luc+"
/note="enhanced luc+ version of the luciferase gene"
/product="firefly luciferase"
/protein_id=""
/transl_table=1
/translation="MEDAKNIKKGPAPFYPLEDGTAGEQLHKAMKRYALVPGTIAFTDA
HIEVDITYAEYFEMSVRLAEAMKRYGLNTNHRIVVCSENSLQFFMPVLGALFIGVAVAP
ANDIYNERELLNSMGISQPTVVFVSKKGLQKILNVQKKLPIIQKIIIMDSKTDYQGFQS
MYTFVTSHLPPGFNEYDFVPESFDRDKTIALIMNSSGSTGLPKGVALPHRTACVRFSHA
RDPIFGNQIIPDTAILSVVPFHHGFGMFTTLGYLICGFRVVLMYRFEEELFLRSLQDYK
IQSALLVPTLFSFFAKSTLIDKYDLSNLHEIASGGAPLSKEVGEAVAKRFHLPGIRQGY
GLTETTSAILITPEGDDKPGAVGKVVPFFEAKVVDLDTGKTLGVNQRGELCVRGPMIMS
GYVNNPEATNALIDKDGWLHSGDIAYWDEDEHFFIVDRLKSLIKYKGYQVAPAELESIL
LQHPNIFDAGVAGLPDDDAGELPAAVVVLEHGKTMTEKEIVDYVASQVTTAKKLRGGVV
FVDEVPKGLTGKLDARKIREILIKAKKGGKIAV*"
/label="luciferase"
promoter 3886..4097
/note="core promoter for human elongation factor
EF-1α"
/label="EF-1α core promoter"
LTR 4110..4378
/note="truncated 5' long terminal repeat (LTR) from human
T-cell leukemia virus (HTLV) type 1"
/label="5' LTR (truncated)"
regulatory 4407..4416
/note="vertebrate consensus sequence for strong initiation
of translation (Kozak, 1987)"
/regulatory_class="other"
/label="Kozak sequence"
CDS 4437..5099
/codon_start=1
/product="green fluorescent protein 2 from Pontellina
plumata, also known as ppluGFP2 (Shagin et al., 2004)"
/transl_table=1
/translation="PAMEIECRITGTLNGVEFELVGGGEGTPKQGRMTNKMKSTKGALT
FSPYLLSHVMGYGFYHFGTYPSGYENPFLHAINNGGYTNTRIEKYEDGGVLHVSFSYRY
EAGRVIGDFKVVGTGFPEDSVIFTDKIIRSNATVEHLHPMGDNVLVGSFARTFSLRDGG
YYSFVVDSHMHFKSAIHPSILQNGGPMFAFRRVEELHSNTELGIVEYQHAFKTPIAFA"
/label="CopGFP"
CDS 5169..5222
/codon_start=1
/note="Eukaryotic ribosomes fail to insert a peptide bond
between the Gly and Pro residues, yielding separate
polypeptides."
/product="2A peptide from Thosea asigna virus capsid
protein"
/protein_id=""
/transl_table=1
/translation="EGRGSLLTCGDVEENPGP"
/label="T2A"
CDS 5223..5822
/codon_start=1
/gene="pac from Streptomyces alboniger"
/note="confers resistance to puromycin"
/product="puromycin N-acetyltransferase"
/transl_table=1
/translation="MTEYKPTVRLATRDDVPRAVRTLAAAFADYPATRHTVDPDRHIER
VTELQELFLTRVGLDIGKVWVADDGAAVAVWTTPESVEAGAVFAEIGPRMAELSGSRLA
AQQQMEGLLAPHRPKEPAWFLATVGVSPDHQGKGLGSAVVLPGVEAAERAGVPAFLETS
APRNLPFYERLGFTVTADVEVPEGPRTWCMTRKPGA*"
/label="PuroR"
polyA_signal 5930..6051
/note="SV40 polyadenylation signal"
/label="SV40 poly(A) signal"
primer_bind complement(7338..7354)
/note="common sequencing primer, one of multiple similar
variants"
/label="M13 rev"
protein_bind 7362..7378
/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-β-D-thiogalactopyranoside (IPTG)."
/label="lac operator"
promoter complement(7386..7416)
/note="promoter for the E. coli lac operon"
/label="lac promoter"
protein_bind 7431..7452
/bound_moiety="E. coli catabolite activator protein"
/note="CAP binding activates transcription in the presence
of cAMP."
/label="CAP binding site"
rep_origin complement(7740..8328)
/direction=LEFT
/note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
replication"
/label="ori"
CDS complement(8499..9359)
/codon_start=1
/gene="bla"
/note="confers resistance to ampicillin, carbenicillin, and
related antibiotics"
/product="β-lactamase"
/transl_table=1
/translation="MSIQHFRVALIPFFAAFCLPVFA,HPETLVKVKDAEDQLGARVGY
IELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEY
SPVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDR
WEPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRS
ALPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGA
SLIKHW*"
/label="AmpR"