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:
- pBAC-TS
- 载体抗性:
- Tetracycline
- 载体长度:
- 10475 bp
- 载体类型:
- Cloning vector
- 复制子:
- ori2
- 载体来源:
- Lee JW, Gyorgy A, Cameron DE, Pyenson N, Choi KR, Way JC, Silver PA, Del Vecchio D, Collins JJ.
pBAC-TS 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。
pBAC-TS 载体序列
LOCUS 40924_5554 10475 bp DNA circular SYN 17-DEC-2018 DEFINITION Cloning vector pBAC-TS, complete sequence. ACCESSION . VERSION . KEYWORDS . SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 10475) AUTHORS Lee JW, Gyorgy A, Cameron DE, Pyenson N, Choi KR, Way JC, Silver PA, Del Vecchio D, Collins JJ. TITLE Creating Single-Copy Genetic Circuits JOURNAL Mol. Cell 63 (2), 329-336 (2016) PUBMED 27425413 REFERENCE 2 (bases 1 to 10475) AUTHORS Lee JW, Gyorgy A, Cameron DE, Pyenson N, Choi KR, Way JC, Silver PA, Del Vecchio D, Collins JJ. TITLE Direct Submission JOURNAL Submitted (18-MAY-2016) Biological Engineering, MIT, 45 Carleton St, MIT Building E25-Rm302, Cambridge, MA 02142, USA REFERENCE 3 (bases 1 to 10475) TITLE Direct Submission REFERENCE 4 (bases 1 to 10475) AUTHORS . TITLE Direct Submission COMMENT SGRef: number: 1; type: "Journal Article"; journalName: "Mol. Cell"; date: "2016"; volume: "63"; issue: "2"; pages: "329-336" COMMENT SGRef: number: 2; type: "Journal Article"; journalName: "Submitted (18-MAY-2016) Biological Engineering, MIT, 45 Carleton St, MIT Building E25-Rm302, Cambridge, MA 02142, USA" COMMENT SGRef: number: 3; type: "Journal Article" COMMENT ##Assembly-Data-START## Sequencing Technology :: Sanger dideoxy sequencing ##Assembly-Data-END## FEATURES Location/Qualifiers source 1..10475 /mol_type="other DNA" /organism="synthetic DNA construct" CDS complement(105..1073) /label=sopB /note="partitioning protein for the bacterial F plasmid" CDS complement(1076..2248) /label=sopA /note="partitioning protein for the bacterial F plasmid" misc_feature complement(2574..2824) /label=incC /note="incompatibility region of the bacterial F plasmid" CDS complement(2830..3582) /label=repE /note="replication initiation protein for the bacterial F plasmid" rep_origin complement(3673..3892) /direction=LEFT /label=ori2 /note="secondary origin of replication for the bacterial F plasmid; also known as oriS" rep_origin complement(3968..4582) /direction=LEFT /label=oriV /note="origin of replication for the bacterial F plasmid" promoter 5440..5542 /label=cat promoter /note="promoter of the E. coli cat gene encoding chloramphenicol acetyltransferase" CDS 5543..6199 /label=CmR /note="chloramphenicol acetyltransferase" terminator complement(6337..6431) /label=lambda t0 terminator /note="transcription terminator from phage lambda" CDS complement(6494..7207) /label=yeGFP /note="yeast-enhanced green fluorescent protein" regulatory complement(7259..7332) /label=PLtetO /note="PLtetO" /regulatory_class="promoter" promoter complement(7259..7332) /label=PLtetO-1 promoter /note="modified phage lambda PL promoter with tet operator sites (Lutz and Bujard, 1997)" prim_transcript 7278 /label=transcription start /note="transcription start" protein_bind 7289..7307 /gene="tetO" /label=tet operator /bound_moiety="tetracycline repressor TetR" /note="bacterial operator O2 for the tetR and tetA genes" protein_bind 7314..7332 /gene="tetO" /label=tet operator /bound_moiety="tetracycline repressor TetR" /note="bacterial operator O2 for the tetR and tetA genes" misc_feature complement(7375..7458) /label=mfLon-specific tag pdt2 /note="mfLon-specific tag pdt2" CDS complement(7459..8538) /label=lacI /note="lac repressor" promoter complement(8575..8648) /label=PLtetO-1 promoter /note="modified phage lambda PL promoter with tet operator sites (Lutz and Bujard, 1997)" promoter 8680..8709 /label=trc promoter /note="strong E. coli promoter; hybrid between the trp and lac UV5 promoters" prim_transcript 8715 /label=transcription start /note="transcription start" protein_bind 8717..8733 /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)." CDS 8780..9400 /label=TetR /note="tetracycline repressor TetR" regulatory 9446..9520 /label=Ptrc-2 /note="Ptrc-2" /regulatory_class="promoter" promoter 9459..9488 /label=trc promoter /note="strong E. coli promoter; hybrid between the trp and lac UV5 promoters" protein_bind 9496..9512 /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)." prim_transcript 9496 /label=transcription start /note="transcription start" CDS 9563..10270 /label=mCherry /note="monomeric derivative of DsRed fluorescent protein (Shaner et al., 2004)" terminator 10333..10419 /label=rrnB T1 terminator /note="transcription terminator T1 from the E. coli rrnB gene"