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:
- w-repeats
- 载体抗性:
- Ampicillin
- 载体长度:
- 14656 bp
- 载体类型:
- Cloning vector
- 复制子:
- ori
- 载体来源:
- Akmammedov A, Geigges M, Paro R.
- 启动子:
- hsp70
w-repeats 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。
w-repeats 载体序列
LOCUS 40924_49372 14656 bp DNA circular SYN 18-DEC-2018 DEFINITION Cloning vector w-repeats, complete sequence. ACCESSION . VERSION . KEYWORDS . SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 14656) AUTHORS Akmammedov A, Geigges M, Paro R. TITLE Single vector non-leaky gene expression system for Drosophila melanogaster JOURNAL Sci Rep 7 (1), 6899 (2017) PUBMED 28761084 REFERENCE 2 (bases 1 to 14656) AUTHORS Akmammedov A, Geigges M, Paro R. TITLE Direct Submission JOURNAL Submitted (04-MAY-2017) Biosystem Science and Engineering, ETH Zurich, Mattenstrasse 26, Basel 4058, Switzerland REFERENCE 3 (bases 1 to 14656) TITLE Direct Submission REFERENCE 4 (bases 1 to 14656) AUTHORS . TITLE Direct Submission COMMENT SGRef: number: 1; type: "Journal Article"; journalName: "Sci Rep"; date: "2017"; volume: "7"; issue: "1"; pages: "6899" COMMENT SGRef: number: 2; type: "Journal Article"; journalName: "Submitted (04-MAY-2017) Biosystem Science and Engineering, ETH Zurich, Mattenstrasse 26, Basel 4058, Switzerland" COMMENT SGRef: number: 3; type: "Journal Article" FEATURES Location/Qualifiers source 1..14656 /mol_type="other DNA" /organism="synthetic DNA construct" rep_origin complement(3..458) /direction=LEFT /label=f1 ori /note="f1 bacteriophage origin of replication; arrow indicates direction of (+) strand synthesis" primer_bind 600..616 /label=M13 fwd /note="common sequencing primer, one of multiple similar variants" promoter 623..641 /label=T7 promoter /note="promoter for bacteriophage T7 RNA polymerase" gene 666..4802 /label=mini-white /note="This modified version of the white gene lacks part of the first intron." protein_bind 4842..5227 /label=gypsy insulator /note="chromatin insulator from Drosophila" polyA_signal complement(5235..5661) /label=hsp70 poly(A) /note="polyadenlyation signal from the Drosophila melanogaster hsp70Ab gene" CDS complement(5680..8724) /codon_start=1 /label=lacZ /note="beta-galactosidase" /translation="VVLQRRDWENPGVTQLNRLAAHPPFASWRNSEEARTDRPSQQLRS LNGEWRFAWFPAPEAVPESWLECDLPEADTVVVPSNWQMHGYDAPIYTNVTYPITVNPP FVPTENPTGCYSLTFNVDESWLQEGQTRIIFDGVNSAFHLWCNGRWVGYGQDSRLPSEF DLSAFLRAGENRLAVMVLRWSDGSYLEDQDMWRMSGIFRDVSLLHKPTTQISDFHVATR FNDDFSRAVLEAEVQMCGELRDYLRVTVSLWQGETQVASGTAPFGGEIIDERGGYADRV TLRLNVENPKLWSAEIPNLYRAVVELHTADGTLIEAEACDVGFREVRIENGLLLLNGKP LLIRGVNRHEHHPLHGQVMDEQTMVQDILLMKQNNFNAVRCSHYPNHPLWYTLCDRYGL YVVDEANIETHGMVPMNRLTDDPRWLPAMSERVTRMVQRDRNHPSVIIWSLGNESGHGA NHDALYRWIKSVDPSRPVQYEGGGADTTATDIICPMYARVDEDQPFPAVPKWSIKKWLS LPGETRPLILCEYAHAMGNSLGGFAKYWQAFRQYPRLQGGFVWDWVDQSLIKYDENGNP WSAYGGDFGDTPNDRQFCMNGLVFADRTPHPALTEAKHQQQFFQFRLSGQTIEVTSEYL FRHSDNELLHWMVALDGKPLASGEVPLDVAPQGKQLIELPELPQPESAGQLWLTVRVVQ PNATAWSEAGHISAWQQWRLAENLSVTLPAASHAIPHLTTSEMDFCIELGNKRWQFNRQ SGFLSQMWIGDKKQLLTPLRDQFTRAPLDNDIGVSEATRIDPNAWVERWKAAGHYQAEA ALLQCTADTLADAVLITTAHAWQHQGKTLFISRKTYRIDGSGQMAITVDVEVASDTPHP ARIGLNCQLAQVAERVNWLGLGPQENYPDRLTAACFDRWDLPLSDMYTPYVFPSENGLR CGTRELNYGPHQWRGDFQFNISRYSQQQLMETSHRHLLHAEEGTWLNIDGFHMGIGGDD SWSPSVSAELQLSAGRYHYQLVWCQK" promoter complement(8846..9294) /label=hsp70 promoter /note="Drosophila melanogaster hsp70Bb promoter" misc_feature complement(9307..11747) /label=w-repeats /note="w-repeats" protein_bind 11755..12140 /label=gypsy insulator /note="chromatin insulator from Drosophila" protein_bind 12180..12249 /label=attB /note="attB site for the phi-C31 integrase (Groth et al., 2000)" promoter complement(12468..12486) /label=T3 promoter /note="promoter for bacteriophage T3 RNA polymerase" primer_bind complement(12507..12523) /label=M13 rev /note="common sequencing primer, one of multiple similar variants" protein_bind 12531..12547 /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(12555..12585) /label=lac promoter /note="promoter for the E. coli lac operon" protein_bind complement(12600..12621) /label=CAP binding site /note="CAP binding activates transcription in the presence of cAMP." rep_origin complement(12909..13497) /direction=LEFT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" CDS complement(13671..14528) /codon_start=1 /label=AmpR /note="beta-lactamase" /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS LIKHW" promoter complement(14529..14633) /label=AmpR promoter