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:
- pCL-Eco
- 载体抗性:
- Ampicillin
- 载体长度:
- 12341 bp
- 载体类型:
- Mammalian Expression, Retroviral
- 复制子:
- ori
- 拷贝数:
- High Copy
- 启动子:
- CMVd1
- 克隆方法:
- Restriction Enzyme
- 5'引物:
- LXSN primer
- 感受态:
- DH10B
- 培养温度:
- 37℃
pCL-Eco 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。
pCL-Eco 载体序列
LOCUS Exported 12341 bp ds-DNA circular SYN 20-DEC-2021
DEFINITION synthetic circular DNA
ACCESSION .
VERSION .
KEYWORDS pCL-Eco
SOURCE synthetic DNA construct
ORGANISM synthetic DNA construct
REFERENCE 1 (bases 1 to 12341)
AUTHORS Naviaux RK, Costanzi E, Haas M, Verma IM
TITLE The pCL vector system: rapid production of helper-free, high-titer,
recombinant retroviruses.
JOURNAL J Virol. 1996 Aug . 70(8):5701-5.
PUBMED 8764092
REFERENCE 2 (bases 1 to 12341)
AUTHORS .
TITLE Direct Submission
COMMENT SGRef: number: 1; type: "Journal Article"; journalName: "J Virol.
1996 Aug . 70(8):5701-5."
FEATURES Location/Qualifiers
source 1..12341
/organism="synthetic DNA construct"
/mol_type="other DNA"
primer_bind 133..152
/label=pBR322ori-F
/note="pBR322 origin, forward primer"
primer_bind 386..403
/label=L4440
/note="L4440 vector, forward primer"
rep_origin 468..602
/label=SV40 ori
/note="SV40 origin of replication"
primer_bind complement(521..540)
/label=SV40pro-F
/note="SV40 promoter/origin, forward primer"
polyA_signal 1060..1194
/label=SV40 poly(A) signal
/note="SV40 polyadenylation signal"
primer_bind complement(1084..1103)
/label=EBV-rev
/note="SV40 polyA terminator, reverse primer"
primer_bind 1138..1157
/label=SV40pA-R
/note="SV40 polyA, reverse primer"
CDS complement(1466..1486)
/codon_start=1
/product="nuclear localization signal of SV40 (simian virus
40) large T antigen"
/label=SV40 NLS
/translation="PKKKRKV"
intron 1616..1681
/label=small t intron
/note="SV40 (simian virus 40) small t antigen intron"
CDS complement(4331..9547)
/codon_start=1
/gene="gag-pol"
/product="Moloney murine leukemia virus Gag-Pol
polyprotein"
/label=gag-pol
/note="contains a read-through stop codon"
/translation="MGQTVTTPLSLTLGHWKDVERIAHNQSVDVKKRRWVTFCSAEWPT
FNVGWPRDGTFNRDLITQVKIKVFSPGPHGHPDQVPYIVTWEALAFDPPPWVKPFVHPK
PPPPLPPSAPSLPLEPPRSTPPRSSLYPALTPSLGAKPKPQVLSDSGGPLIDLLTEDPP
PYRDPRPPPSDRDGNGGEATPAGEAPDPSPMASRLRGRREPPVADSTTSQAFPLRAGGN
GQLQYWPFSSSDLYNWKNNNPSFSEDPGKLTALIESVLITHQPTWDDCQQLLGTLLTGE
EKQRVLLEARKAVRGDDGRPTQLPNEVDAAFPLERPDWDYTTQAGRNHLVHYRQLLLAG
LQNAGRSPTNLAKVKGITQGPNESPSAFLERLKEAYRRYTPYDPEDPGQETNVSMSFIW
QSAPDIGRKLERLEDLKNKTLGDLVREAEKIFNKRETPEEREERIRRETEEKEERRRTE
DEQKEKERDRRRHREMSKLLATVVSGQKQDRQGGERRRSQLDRDQCAYCKEKGHWAKDC
PKKPRGPRGPRPQTSLLTLDD*GGQGQEPPPEPRITLKVGGQPVTFLVDTGAQHSVLTQ
NPGPLSDKSAWVQGATGGKRYRWTTDRKVHLATGKVTHSFLHVPDCPYPLLGRDLLTKL
KAQIHFEGSGAQVMGPMGQPLQVLTLNIEDEYRLHETSKEPDVSLGSTWLSDFPQAWAE
TGGMGLAVRQAPLIIPLKATSTPVSIKQYPMSQEARLGIKPHIQRLLDQGILVPCQSPW
NTPLLPVKKPGTNDYRPVQDLREVNKRVEDIHPTVPNPYNLLSGLPPSHQWYTVLDLKD
AFFCLRLHPTSQPLFAFEWRDPEMGISGQLTWTRLPQGFKNSPTLFDEALHRDLADFRI
QHPDLILLQYVDDLLLAATSELDCQQGTRALLQTLGNLGYRASAKKAQICQKQVKYLGY
LLKEGQRWLTEARKETVMGQPTPKTPRQLREFLGTAGFCRLWIPGFAEMAAPLYPLTKT
GTLFNWGPDQQKAYQEIKQALLTAPALGLPDLTKPFELFVDEKQGYAKGVLTQKLGPWR
RPVAYLSKKLDPVAAGWPPCLRMVAAIAVLTKDAGKLTMGQPLVILAPHAVEALVKQPP
DRWLSNARMTHYQALLLDTDRVQFGPVVALNPATLLPLPEEGLQHNCLDILAEAHGTRP
DLTDQPLPDADHTWYTDGSSLLQEGQRKAGAAVTTETEVIWAKALPAGTSAQRAELIAL
TQALKMAEGKKLNVYTDSRYAFATAHIHGEIYRRRGLLTSEGKEIKNKDEILALLKALF
LPKRLSIIHCPGHQKGHSAEARGNRMADQAARKAAITETPDTSTLLIENSSPYTSEHFH
YTVTDIKDLTKLGAIYDKTKKYWVYQGKPVMPDQFTFELLDFLHQLTHLSFSKMKALLE
RSHSPYYMLNRDRTLKNITETCKACAQVNASKSAVKQGTRVRGHRPGTHWEIDFTEIKP
GLYGYKYLLVFIDTFSGWIEAFPTKKETAKVVTKKLLEEIFPRFGMPQVLGTDNGPAFV
SKVSQTVADLLGIDWKLHCAYRPQSSGQVERMNRTIKETLTKLTLATGSRDWVLLLPLA
LYRARNTPGPHGLTPYEILYGAPPPLVNFPDPDMTRVTNSPSLQAHLQALYLVQHEVWR
PLAAAYQEQLDRPVVPHPYRVGDTVWVRRHQTKNLEPRWKGPYTVLLTTPTALKVDGIA
AWIHAAHVKAADPGGGPSSRLTWRVQRSQNPLKIRLTREAP"
misc_feature 4394..4768
/label=pol region
/note="Moloney murine leukemia virus (MMLV) pol region
containing the splice acceptor site"
primer_bind complement(4440..4456)
/label=pBMN 5'
/note="MMLV sequence, for inserts in pBMN retroviral
vector"
primer_bind complement(9144..9160)
/label=pBABE 5'
/note="Psi packaging signal, forward primer for pBABE
vectors"
primer_bind complement(9176..9198)
/label=pLXSN 5'
/note="Murine stem cell virus, forward primer. Also called
MSCV"
promoter complement(9856..10059)
/label=CMV promoter
/note="human cytomegalovirus (CMV) immediate early
promoter"
primer_bind complement(9885..9905)
/label=CMV-F
/note="Human CMV immediate early promoter, forward primer"
enhancer 10060..10439
/label=CMV enhancer
/note="human cytomegalovirus immediate early enhancer"
primer_bind complement(10584..10603)
/label=pRS-marker
/note="pRS vectors, use to sequence yeast selectable
marker"
primer_bind complement(10763..10781)
/label=pBRforEco
/note="pBR322 vectors, upsteam of EcoRI site, forward
primer"
promoter 10849..10953
/gene="bla"
/label=AmpR promoter
CDS 10954..11814
/codon_start=1
/gene="bla"
/product="beta-lactamase"
/label=AmpR
/note="confers resistance to ampicillin, carbenicillin, and
related antibiotics"
/translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
EPELNEAIPNDERDTTMPAAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
LIKHW"
primer_bind complement(11172..11191)
/label=Amp-R
/note="Ampicillin resistance gene, reverse primer"
rep_origin join(11985..12341,1..232)
/direction=RIGHT
/label=ori
/note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
replication"