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:
- pTip-CH1.2
- 载体抗性:
- Ampicillin
- 载体长度:
- 8337 bp
- 载体类型:
- Expression vector
- 复制子:
- ori
- 载体来源:
- Nakashima N, Tamura T.
pTip-CH1.2 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。
pTip-CH1.2 载体序列
LOCUS V002645 8337 bp DNA circular SYN 18-DEC-2018 DEFINITION Exported. ACCESSION V002645 VERSION V002645 KEYWORDS . SOURCE synthetic DNA construct ORGANISM synthetic DNA construct . REFERENCE 1 (bases 1 to 8337) AUTHORS Nakashima N, Tamura T. TITLE A novel system for expressing recombinant proteins over a wide temperature range from 4 to 35 degrees C JOURNAL Biotechnol. Bioeng. 86 (2), 136-148 (2004) PUBMED 15052633 REFERENCE 2 (bases 1 to 8337) AUTHORS Nakashima N, Tamura T. TITLE Direct Submission JOURNAL Submitted (18-OCT-2002) Nobutaka Nakashima, National Institute of Advanced Industrial Science and Technology; Tsukisamu-Higashi 2-17-2-1, Sapporo, Toyohira-ku 062-8517, Japan (E-mail:n-nakashima@aist.go.jp, Tel:81-11-857-8937) REFERENCE 3 (bases 1 to 8337) TITLE Direct Submission REFERENCE 4 (bases 1 to 8337) AUTHORS . TITLE Direct Submission COMMENT SGRef: number: 1; type: "Journal Article"; journalName: "Biotechnol. Bioeng."; date: "2004"; volume: "86"; issue: "2"; pages: "136-148" SGRef: number: 2; type: "Journal Article"; journalName: "Submitted (18-OCT-2002) Nobutaka Nakashima, National Institute of Advanced Industrial Science and Technology"; volume: " Tsukisamu-Higashi 2-17-2-1, Sapporo, Toyohira-ku 062-8517, Japan (E-mail:n-nakashima@aist.go.jp, Tel"; pages: "81-11-857-8937" SGRef: number: 3; type: "Journal Article" FEATURES Location/Qualifiers source 1..8337 /mol_type="other DNA" /organism="synthetic DNA construct" CDS 170..976 /gene="tsnR" /label="23S rRNA (adenosine(1067)-2'-O)-methyltransferase" /note="23S rRNA (adenosine(1067)-2'-O)-methyltransferase from Streptomyces azureus. Accession#: P18644" CDS 1541..2716 /codon_start=1 /gene="Chlr" /product="chloramphenicol resistance protein" /label="Chlr" /protein_id="BAD11263.1" /translation="VPFAIYVLGLAVFAQGTSEFMLSGLIPDMARDLGVSVPAAGLLTS AFAVGMIIGAPLMAIASMRWPRRRALLTFLITFMLVHVIGALTSSFEVLLVTRIVGALA NAGFLAVALGAAMAMVPADMKGRATSVLLGGVTIACVAGVPGGAFLGEMWGWRAAFWAV VVISAPAVVAIMFATPAEPLAESTPNAKRELSSLRSRKLQLMLVLGALINGATFCSFTY MAPTLTDISGFDSRWIPLLLGLFGLGSFIGVSVGGRLADTRPFQLLAVGSAALLTGWIV FALTASHPAVTLVMLFVQGALSFAVGSTLISQVLYAADAAPTLGGSFATAAFNVGAALG PALGGLAIGMGLSYRAPLWTSAALVTLAIVIGAATLSLWRRPASVHESVPA" gene 1541..2716 /gene="Chlr" /label="Chlr" regulatory 2905..3080 /gene="ThcA" /regulatory_class="promoter" gene 2905..3080 /gene="ThcA" /label="ThcA" CDS 3081..3839 /gene="tipA" /label="HTH-type transcriptional activator TipA" /note="HTH-type transcriptional activator TipA from Streptomyces lividans. Accession#: P0A4T9" promoter 4020..4124 /label="AmpR promoter" CDS 4125..4982 /label="AmpR" /note="beta-lactamase" rep_origin 5156..5744 /direction=RIGHT /label="ori" /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" regulatory complement(5995..6175) /gene="ThcA" /regulatory_class="terminator" gene complement(5995..6175) /gene="ThcA" /label="ThcA" CDS complement(6189..6206) /label="6xHis" /note="6xHis affinity tag" gene complement(6255..6497) /gene="TipA" /label="TipA" regulatory complement(6255..6273) /gene="TipA" /regulatory_class="ribosome_binding_site" regulatory complement(6274..6497) /gene="TipA" /regulatory_class="promoter" CDS 6936..7823 /codon_start=1 /gene="RepA" /label="RepA" /protein_id="BAD11266.1" /translation="MKSATGGVLSDRQWADAVLLPHRPFATNNLQRGQYRMSRDDALAM RYVEHSPHALLGSIVIDCDHVDAAMRAFEQPSDHPAPNWVAQSPSGRAHIGWWLGPNHV CRTDSARLTPLRYAHRIETGLKISVGGDFAYGGQLTKNPIHPDWETIYGPATPYTLRQL ATIHTPRQMPRRPDRAVGLGRNVTMFDATRRWAYPQWWQHRNGTGRDWDHLVLQHCHAV NTEFTTPLPFTEVRATAQSISKWIWRNFTEEQYRARQAHLGQKGGKATTLAKQEAVRNN ARKYDEHTMREAII" gene 6936..7823 /gene="RepA" /label="RepA" CDS 7823..8107 /codon_start=1 /gene="RepB" /label="RepB" /protein_id="BAD11267.1" /translation="MGGAKNPVRRKMTAAAAAEKFGASTRTIQRLFAEPRDDYLGRAKA RRDKAVELRKQGLKYREIAEAMELSTGIVGRLLHDARRHGEISAEDLSA" gene 7823..8107 /gene="RepB" /label="RepB"