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:
- pTrc-tpiA
- 载体抗性:
- Ampicillin
- 载体长度:
- 4921 bp
- 载体类型:
- Cloning vector
- 复制子:
- ori
- 载体来源:
- Kim B, Binkley R, Kim HU, Lee SY.
pTrc-tpiA 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。
pTrc-tpiA 载体序列
LOCUS V002546 4921 bp DNA circular SYN 18-DEC-2018 DEFINITION Exported. ACCESSION V002546 VERSION V002546 KEYWORDS . SOURCE synthetic DNA construct ORGANISM synthetic DNA construct . REFERENCE 1 (bases 1 to 4921) AUTHORS Kim B, Binkley R, Kim HU, Lee SY. TITLE Metabolic engineering of Escherichia coli for the enhanced production of l-tyrosine JOURNAL Biotechnol. Bioeng. (2018) In press PUBMED 30019750 REFERENCE 2 (bases 1 to 4921) AUTHORS Yang D, Kim WJ, Yoo SM, Choi JH, Ha SH, Lee MH, Lee SY. TITLE Direct Submission JOURNAL Submitted (15-JUN-2018) Dept. Chemical and Biomolecular Engineering, Korea Advanced Institute of Science and Technology, Daehak-ro 291, Daejeon 34141, South Korea REFERENCE 3 (bases 1 to 4921) TITLE Direct Submission REFERENCE 4 (bases 1 to 4921) AUTHORS . TITLE Direct Submission COMMENT SGRef: number: 1; type: "Journal Article"; journalName: "Biotechnol. Bioeng. (2018) In press" SGRef: number: 2; type: "Journal Article"; journalName: "Submitted (15-JUN-2018) Dept. Chemical and Biomolecular Engineering, Korea Advanced Institute of Science and Technology, Daehak-ro 291, Daejeon 34141, South Korea" SGRef: number: 3; type: "Journal Article" FEATURES Location/Qualifiers source 1..4921 /mol_type="other DNA" /organism="synthetic DNA construct" promoter 193..222 /label="trc promoter" /note="strong E. coli promoter; hybrid between the trp and lac UV5 promoters" protein_bind 230..246 /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)." CDS 267..1031 /gene="tpiA" /label="Triosephosphate isomerase" /note="Triosephosphate isomerase from Escherichia coli (strain ATCC 8739 / DSM 1576 / NBRC 3972 / NCIMB 8545 / WDCM 00012 / Crooks). Accession#: B1IVG0" terminator 1274..1360 /label="rrnB T1 terminator" /note="transcription terminator T1 from the E. coli rrnB gene" terminator 1452..1479 /label="rrnB T2 terminator" /note="transcription terminator T2 from the E. coli rrnB gene" promoter 1499..1590 /label="AmpR promoter" CDS 1591..2448 /label="AmpR" /note="beta-lactamase" rep_origin 2622..3210 /label="ori" /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" misc_feature complement(3396..3536) /label="bom" /note="basis of mobility region from pBR322" promoter 3722..3799 /label="lacIq promoter" /note="In the lacIq allele, a single base change in the promoter boosts expression of the lacI gene about 10-fold." CDS 3800..4879 /label="lacI" /note="lac repressor"