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:
- LIC-pKLAC-LC1
- 载体抗性:
- Ampicillin
- 载体长度:
- 10540 bp
- 载体类型:
- Expression vector
- 复制子:
- ori
- 载体来源:
- Dortay H, Akula UM, Westphal C, Sittig M, Mueller-Roeber B.
- 启动子:
- ADH1
LIC-pKLAC-LC1 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。
LIC-pKLAC-LC1 载体序列
LOCUS 40924_1769 10540 bp DNA circular SYN 17-DEC-2018
DEFINITION Expression vector LIC-pKLAC-LC1, complete sequence.
ACCESSION .
VERSION .
KEYWORDS .
SOURCE synthetic DNA construct
ORGANISM synthetic DNA construct
REFERENCE 1 (bases 1 to 10540)
AUTHORS Dortay H, Akula UM, Westphal C, Sittig M, Mueller-Roeber B.
TITLE High-Throughput Protein Expression Using a Combination of
Ligation-Independent Cloning (LIC) and Infrared Fluorescent Protein
(IFP) Detection
JOURNAL PLoS ONE 6 (4), E18900 (2011)
PUBMED 21541323
REFERENCE 2 (bases 1 to 10540)
AUTHORS Dortay H, Mueller-Roeber B.
TITLE Direct Submission
JOURNAL Submitted (14-FEB-2011) Molecular Biology, University of Potsdam,
Germany, Karl-Liebknecht-Str. 24-25, Haus 20, Potsdam, Brandenburg
14476, Germany
REFERENCE 3 (bases 1 to 10540)
TITLE Direct Submission
REFERENCE 4 (bases 1 to 10540)
AUTHORS .
TITLE Direct Submission
COMMENT SGRef: number: 1; type: "Journal Article"; journalName: "PLoS ONE";
date: "2011"; volume: "6"; issue: "4"; pages: "E18900"
COMMENT SGRef: number: 2; type: "Journal Article"; journalName: "Submitted
(14-FEB-2011) Molecular Biology, University of Potsdam, Germany,
Karl-Liebknecht-Str. 24-25, Haus 20, Potsdam, Brandenburg 14476,
Germany"
COMMENT SGRef: number: 3; type: "Journal Article"
FEATURES Location/Qualifiers
source 1..10540
/mol_type="other DNA"
/organism="synthetic DNA construct"
misc_feature 14..16
/label=start codon
/note="start codon"
CDS 26..43
/codon_start=1
/label=6xHis
/note="6xHis affinity tag"
/translation="HHHHHH"
CDS 53..1036
/codon_start=1
/label=IFP1.4
/note="bacteriophytochrome-based monomeric infrared
fluorescent protein (Shu et al., 2009)"
/translation="MARDPLPFFPPLYLGGPEITTENCEREPIHIPGSIQPHGALLTAD
GHSGEVLQVSLNAATFLGQEPTVLRGQTLAALLPEQWPALQAALPPGCPDALQYRATLD
WPAAGHLSLTVHRVAELLILEFEPTEAWDSIGPHALRNAMFALESAPNLRALAEVATQT
VRELTGFDRVMLYKFAPDATGEMIAEARREGMQAFLGHRFPASHTPAQARALYTRHLLR
LTADTRAAAVPLDPVLNPQTNAPTPLGGAVLRATSPMHMQYLRNMGVGSSLSVSVVVGG
QLWGLIVCHHQTPYVLPPDLRTTLEELGRKLSGQVQRKEAGMDELYK"
CDS 1037..1057
/codon_start=1
/label=TEV site
/note="tobacco etch virus (TEV) protease recognition and
cleavage site"
/translation="ENLYFQG"
misc_feature 1076..1083
/label=PmeI recognition site
/note="PmeI recognition site"
misc_feature 1084..1750
/label=LIC stuffer fragment
/note="LIC stuffer fragment"
misc_feature 1751..1758
/label=PmeI recognition site
/note="PmeI recognition site"
terminator 1804..2422
/label=LAC4 terminator
/note="transcription terminator for Kluyveromyces lactis
LAC4"
promoter 2423..3125
/label=ADH1 promoter
/note="promoter for alcohol dehydrogenase 1"
CDS 3146..4789
/codon_start=1
/label=amdS
/note="acetamidase from Aspergillus nidulans"
/translation="MPQSWEELAADKRARLAKTIPDEWKVQTLPAEDSVIDFPKKSGIL
SEAELKITEASAADLVSKLAAGELTSVEVTLAFCKRAAIAQQLTNCAHEFFPDAALAQA
RELDEYYAKHKRPVGPLHGLPISLKDQLRVKGYETSMGYISWLNKYDEGDSVLTTMLRK
AGAVFYVKTSVPQTLMVCETVNNIIGRTVNPRNKNWSCGGSSGGEGAIVGIRGGVIGVG
TDIGGSIRVPAAFNFLYGLRPSHGRLPYAKMANSMEGQETVHSVVGPITHSVEDLRLFT
KSVLGQEPWKYDSKVIPMPWRQSESDIIASKIKNGGLNIGYYNFDGNVLPHPPILRGVE
TTVAALAKAGHTVTPWTPYKHDFGHDLISHIYAADGSADVMRDISASGEPAIPNIKDLL
NPNIKAVNMNELWDTHLQKWNYQMEYLEKWREAEEKAGKELDAIIAPITPTAAVRHDQF
RYYGYASVINLLDFTSVVVPVTFADKNIDKKNESFKAVSELDALVQEEYDPEAYHGAPV
AVQVIGRRLSEERTLAIAEEVGKLLGNVVTP"
promoter 5428..6084
/label=LAC4 promoter (5')
/note="5' portion of the Kluyveromyces lactis LAC4
promoter"
promoter complement(6101..6119)
/label=T7 promoter
/note="promoter for bacteriophage T7 RNA polymerase"
primer_bind complement(6133..6149)
/label=M13 rev
/note="common sequencing primer, one of multiple similar
variants"
protein_bind 6157..6173
/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(6181..6211)
/label=lac promoter
/note="promoter for the E. coli lac operon"
protein_bind complement(6226..6247)
/label=CAP binding site
/note="CAP binding activates transcription in the presence
of cAMP."
rep_origin complement(6535..7123)
/direction=LEFT
/label=ori
/note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
replication"
CDS complement(7297..8154)
/codon_start=1
/label=AmpR
/note="beta-lactamase"
/translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
LIKHW"
promoter complement(8155..8259)
/label=AmpR promoter
rep_origin 8286..8741
/label=f1 ori
/note="f1 bacteriophage origin of replication; arrow
indicates direction of (+) strand synthesis"
primer_bind 8882..8898
/label=M13 fwd
/note="common sequencing primer, one of multiple similar
variants"
promoter 8912..10440
/label=LAC4 promoter (3')
/note="3' portion of a modified Kluyveromyces lactis LAC4
promoter"
protein_bind 10446..10462
/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)."