质粒编号
质粒名称
规格
价格
V017496
pGERM
5 μg (冻干粉)
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:
- pGERM
- 载体抗性:
- Ampicillin
- 载体长度:
- 4807 bp
- 载体类型:
- Bacteroides Plasmid
- 载体来源:
- Shoemaker,N.B., Wang,G.R. and Salyers,A.A.
- 拷贝数:
- High copy number
- 感受态:
- stbl3
- 培养温度:
- 37℃
pGERM 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。
pGERM 载体序列
LOCUS pGERM 4807 bp DNA circular SYN 26-JUL-2024
DEFINITION PGERM gene disruption vector, complete sequence.
ACCESSION EF155418
VERSION .
KEYWORDS .
SOURCE synthetic DNA construct
ORGANISM synthetic DNA construct
REFERENCE 1 (bases 1 to 4807)
AUTHORS Shoemaker NB, Wang GR, Salyers AA.
TITLE Multiple gene products and sequences required for excision of the
mobilizable integrated Bacteroides element NBU1
JOURNAL J. Bacteriol. 182 (4), 928-936 (2000)
PUBMED 10648516
REFERENCE 2 (bases 1 to 4807)
AUTHORS Chiang HC, Manchester JK, Gordon JI.
TITLE Regulation of nitrogen metabolism by a novel hybrid two-component
system protein in a prominent human gut symbiont, Bacteroides
thetaiotaomicron
JOURNAL Unpublished
REFERENCE 3 (bases 1 to 4807)
AUTHORS Chiang HC, Manchester JK, Gordon JI.
TITLE Direct Submission
JOURNAL Submitted (03-DEC-2006) Center for Genome Sciences, Washington
University School of Medicine, 4444 Forest Park Ave. Campus Box
8510, Saint Louis, MO 63108, USA
REFERENCE 4 (bases 1 to 4807)
AUTHORS .
TITLE Direct Submission
COMMENT SGRef: number: 1; type: "Journal Article"; journalName: "J.
Bacteriol."; date: "2000"; volume: "182"; issue: "4"; pages:
"928-936"
COMMENT SGRef: number: 2; type: "Journal Article"; journalName:
"Unpublished"
COMMENT SGRef: number: 3; type: "Journal Article"; journalName: "Submitted
(03-DEC-2006) Center for Genome Sciences, Washington University
School of Medicine, 4444 Forest Park Ave. Campus Box 8510, Saint
Louis, MO 63108, USA"
FEATURES Location/Qualifiers
source 1..4807
/mol_type="other DNA"
/label=pGERM is pUC19 with the RK2 oriT and ermG
/note="pGERM is pUC19 with the RK2 oriT and ermG"
/db_xref="taxon:433620"
/organism="pGERM gene disruption vector"
CDS complement(44..367)
/codon_start=1
/gene="lacZ fragment"
/product="LacZ-alpha fragment of beta-galactosidase"
/label=lacZ-alpha
/translation="MTMITPSLHACRSTLEDPRVPSSNSLAVVLQRRDWENPGVTQLNR
LAAHPPFASWRNSEEARTDRPSQQLRSLNGEWRLMRYFLLTHLCGISHRIWCTLSTICS
DAA"
primer_bind 277..293
/label=M13 fwd
/note="common sequencing primer, one of multiple similar
variants"
misc_feature 294..350
/label=MCS
/note="pUC18/19 multiple cloning site"
primer_bind complement(363..379)
/label=M13 rev
/note="common sequencing primer, one of multiple similar
variants"
protein_bind 387..403
/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(411..441)
/label=lac promoter
/note="promoter for the E. coli lac operon"
protein_bind 456..477
/label=CAP binding site
/bound_moiety="E. coli catabolite activator protein"
/note="CAP binding activates transcription in the presence
of cAMP."
CDS complement(709..1080)
/codon_start=1
/gene="traJ"
/product="oriT-recognizing protein"
/label=traJ
/translation="MADETKPTRKGSPPIKVYCLPDERRAIEEKAAAAGMSLSAYLLAV
GQGYKITGVVDYEHVRELARINGDLGRLGGLLKLWLTDDPRTARFGDATILALLAKIEE
KQDELGKVMMGVVRPRAEP"
oriT 1113..1224
/label=origin of transfer from 56 kb circular plasmid RK2
/note="origin of transfer from 56 kb circular plasmid RK2"
oriT 1113..1222
/note="incP origin of transfer"
rep_origin complement(1543..2131)
/direction=LEFT
/label=ori
/note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
replication"
gene complement(2302..3162)
/gene="AmpR"
/label=AmpR
CDS complement(2302..3162)
/codon_start=1
/transl_table=11
/gene="AmpR"
/product="beta lactamase"
/label=AmpR
/protein_id="ABO69572.1"
/translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
LIKHW"
gene 3650..4384
/gene="ermG"
/label=ermG
CDS 3650..4384
/codon_start=1
/transl_table=11
/gene="ermG"
/product="rRNA methyltransferase"
/label=ermG
/protein_id="ABO69573.1"
/translation="MNKVNIKDSQNFITSKYHIEKIMNCISLDEKDNIFEIGAGKGHFT
AGLVKRCNFVTAIEIDSKLCEVTRNKLLNYPNYQIVNDDILKFTFPSHNPYKIFGSIPY
NISTNIIRKIVFESSATISYLIVEYGFAKMLLDTNRSLALLLMAEVDISILAKIPRYYF
HPKPKVDSTLIVLKRKPAKMAFKERKKYETFVMKWVNKEYEKLFTKNQFNKALKHARIY
DINNISFEQFVSLFNSYKIFNG"