质粒编号
质粒名称
规格
价格
V010598
pCX-OKS-2A
5 μg (冻干粉)
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:
- pCX-OKS-2A
- 载体抗性:
- Ampicillin
- 载体长度:
- 8495 bp
- 载体类型:
- Mammalian Expression
- 复制子:
- ori
- 拷贝数:
- High Copy
- 启动子:
- CAG
- 克隆方法:
- Restriction Enzyme
- 5'引物:
- GCGAGCCGCAGCCATTGCCTTTTA
- 3'引物:
- TTAGCCAGAAGTCAGATGCTC
pCX-OKS-2A 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。
pCX-OKS-2A 载体序列
LOCUS 40924_13930 8495 bp DNA circular SYN 13-MAY-2021
DEFINITION Non-integrating (episomal) expression of mouse Oct3/4, Klf4 and
Sox2.
ACCESSION .
VERSION .
KEYWORDS .
SOURCE synthetic DNA construct
ORGANISM synthetic DNA construct
REFERENCE 1 (bases 1 to 8495)
AUTHORS Okita K, Nakagawa M, Hyenjong H, Ichisaka T, Yamanaka S
TITLE Generation of Mouse Induced Pluripotent Stem Cells Without Viral
Vectors.
JOURNAL Science. 2008 Oct 9. ():.
PUBMED 18845712
REFERENCE 2 (bases 1 to 8495)
TITLE Direct Submission
REFERENCE 3 (bases 1 to 8495)
AUTHORS .
TITLE Direct Submission
COMMENT SGRef: number: 1; type: "Journal Article"; journalName: "Science.
2008 Oct 9. ():."
COMMENT SGRef: number: 2; type: "Journal Article"
FEATURES Location/Qualifiers
source 1..8495
/mol_type="other DNA"
/organism="synthetic DNA construct"
enhancer 4..383
/label=CMV enhancer
/note="human cytomegalovirus immediate early enhancer"
promoter 385..661
/label=chicken beta-actin promoter
intron 662..1678
/label=chimeric intron
/note="chimera between introns from chicken beta-actin and
rabbit beta-globin"
primer_bind 1686..1705
/label=pCAG-F
/note="Rabbit beta-globin intron, for pCAG plasmids,
forward primer"
CDS 1762..2817
/codon_start=1
/label=mPou5f1
/note="Mus musculus Oct-4 gene. Encodes a transcription
factor containing a POU homeodomain that plays a key role
in embryonic development and stem cell pluripotency."
/translation="MAGHLASDFAFSPPPGGGDGSAGLEPGWVDSRTWLSFQGPPGGPG
IGPGSEVLGISPCPPAYEFCGGMAYCGPQVGLGLVPQVGVETLQPEGQAGARVESNSEG
TSSEPCADRPNAVKLEKVEPTPEESQDMKALQKELEQFAKLLKQKRITLGYTQADVGLT
LGVLFGKVFSQTTICRFEALQLSLKNMCKLRPLLEKWVEEADNNENLQEICKSETLVQA
RKRKRTSIENRVRWSLETMFLKCPKPSLQQITHIANQLGLEKDVVRVWFCNRRQKGKRS
SIEYSQREEYEATGTPFPGGAVSFPLPPGPHFGTPGYGSPHFTTLYSVPFPEGEAFPSV
PVTALGSPMHSN"
CDS 2842..2907
/codon_start=1
/product="2A peptide from foot-and-mouth disease virus
polyprotein"
/label=F2A
/note="Eukaryotic ribosomes fail to insert a peptide bond
between the Gly and Pro residues, yielding separate
polypeptides."
/translation="VKQTLNFDLLKLAGDVESNPGP"
CDS 2911..4332
/codon_start=1
/label=mKlf4
/note="Mouse Kruppel-like factor (Klf4) gene. Belongs to
the relatively large family of SP1-like transcription
factors and is involved in the regulation of proliferation,
differentiation, apoptosis and somatic cell reprogramming."
/translation="MAVSDALLPSFSTFASGPAGREKTLRPAGAPTNRWREELSHMKRL
PPLPGRPYDLAATVATDLESGGAGAACSSNNPALLARRETEEFNDLLDLDFILSNSLTH
QESVAATVTTSASASSSSSPASSGPASAPSTCSFSYPIRAGGDPGVAASNTGGGLLYSR
ESAPPPTAPFNLADINDVSPSGGFVAELLRPELDPVYIPPQQPQPPGGGLMGKFVLKAS
LTTPGSEYSSPSVISVSKGSPDGSHPVVVAPYSGGPPRMCPKIKQEAVPSCTVSRSLEA
HLSAGPQLSNGHRPNTHDFPLGRQLPTRTTPTLSPEELLNSRDCHPGLPLPPGFHPHPG
PNYPPFLPDQMQSQVPSLHYQELMPPGSCLPEEPKPKRGRRSWPRKRTATHTCDYAGCG
KTYTKSSHLKAHLRTHTGEKPYHCDWDGCGWKFARSDELTRHYRKHTGHRPFQCQKCDR
AFSRSDHLALHMKRHF"
CDS 4357..4422
/codon_start=1
/product="2A peptide from foot-and-mouth disease virus
polyprotein"
/label=F2A
/note="Eukaryotic ribosomes fail to insert a peptide bond
between the Gly and Pro residues, yielding separate
polypeptides."
/translation="VKQTLNFDLLKLAGDVESNPGP"
CDS 4447..5403
/codon_start=1
/label=mSox2
/note="Mus musculus transcription factor SOX-2 gene.
Belongs to the SRY-related HMG-box (SOX) family of
transcription factors, which is involved in the regulation
of embryonic development and in the determination of cell
fate"
/translation="MYNMMETELKPPGPQQASGGGGGGGNATAAATGGNQKNSPDRVKR
PMNAFMVWSRGQRRKMAQENPKMHNSEISKRLGAEWKLLSETEKRPFIDEAKRLRALHM
KEHPDYKYRPRRKTKTLMKKDKYTLPGGLLAPGGNSMASGVGVGAGLGAGVNQRMDSYA
HMNGWSNGSYSMMQEQLGYPQHPGLNAHGAAQMQPMHRYDVSALQYNSMTSSQTYMNGS
PTYSMSYSQQGTPGMALGSMGSVVKSEASSSPPVVTSSSHSRAPCQAGDLRDMISMYLP
GAEVPEPAAPSRLHMAQHYQSGPVPGTAINGTLPLSHM"
primer_bind complement(5516..5535)
/label=Bglob-pA-R
/note="Rabbit beta-globin polyA region, reverse primer"
polyA_signal 5581..5636
/label=beta-globin poly(A) signal
/note="rabbit beta-globin polyadenylation signal (Gil and
Proudfoot, 1987)"
primer_bind complement(5635..5654)
/label=rbglobpA-R
/note="Rabbit beta-globin polyA, reverse primer. Also
called rb-glob-pA-term-R"
primer_bind complement(5997..6013)
/label=M13 rev
/note="common sequencing primer, one of multiple similar
variants"
protein_bind complement(6021..6037)
/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(6045..6075)
/label=lac promoter
/note="promoter for the E. coli lac operon"
protein_bind complement(6090..6111)
/label=CAP binding site
/note="CAP binding activates transcription in the presence
of cAMP."
promoter 6169..6365
/label=SV40 promoter
/note="SV40 early promoter"
polyA_signal 6371..6505
/label=SV40 poly(A) signal
/note="SV40 polyadenylation signal"
primer_bind complement(6573..6590)
/label=L4440
/note="L4440 vector, forward primer"
rep_origin complement(6744..7332)
/direction=LEFT
/label=ori
/note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
replication"
CDS complement(7506..8363)
/codon_start=1
/label=AmpR
/note="beta-lactamase"
/translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
LIKHW"
promoter complement(8364..8468)
/label=AmpR promoter
promoter 8495
/label=CAG
/note="CMV early enhancer fused to modified chicken
beta-actin promoter"