pADGal4 2.1 载体 (V009580)

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:
pADGal4 2.1
载体抗性:
Ampicillin
载体长度:
7653 bp
载体类型:
Cloning vector
复制子:
ori
宿主:
Yeast
载体来源:
Lu Q.
启动子:
LEU2

pADGal4 2.1 载体载体图谱

pADGal4 2.17653 bp3006009001200150018002100240027003000330036003900420045004800510054005700600063006600690072007500ADH1 promoterSV40 NLSGAL4 activation domainT7 promoterADH1 terminatorLEU2LEU2 promoterM13 revlac operatorlac promoterCAP binding sitef1 oriAmpR promoterSV40 enhanceroriAmpRAmpR promoter2u ori

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。

pADGal4 2.1 载体载体序列

LOCUS       40924_4050        7653 bp DNA     circular SYN 17-DEC-2018
DEFINITION  Cloning vector pADGal4 2.1, complete sequence.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7653)
  AUTHORS   Lu Q.
  TITLE     Direct Submission
  JOURNAL   Submitted (06-NOV-1997) Technical Services, Stratagene, 11011 North 
            Torrey Pines Road, La Jolla, CA 92037, USA
REFERENCE   2  (bases 1 to 7653)
  AUTHORS   Lu Q.
  TITLE     Direct Submission
  JOURNAL   Submitted (25-JAN-1999) Technical Services, Stratagene, 11011 North 
            Torrey Pines Road, La Jolla, CA 92037, USA
REFERENCE   3  (bases 1 to 7653)
  TITLE     Direct Submission
REFERENCE   4  (bases 1 to 7653)
  AUTHORS   .
  TITLE     Direct Submission
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Submitted 
            (06-NOV-1997) Technical Services, Stratagene, 11011 North Torrey 
            Pines Road, La Jolla, CA 92037, USA"
COMMENT     SGRef: number: 2; type: "Journal Article"; journalName: "Submitted 
            (25-JAN-1999) Technical Services, Stratagene, 11011 North Torrey 
            Pines Road, La Jolla, CA 92037, USA"
COMMENT     SGRef: number: 3; type: "Journal Article"
COMMENT     On Jan 25, 1999 this sequence version replaced gi:2695664.
FEATURES             Location/Qualifiers
     source          1..7653
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        5..406
                     /label=ADH1 promoter
                     /note="promoter for alcohol dehydrogenase 1"
     CDS             452..472
                     /codon_start=1
                     /label=SV40 NLS
                     /note="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /translation="PKKKRKV"
     CDS             488..829
                     /codon_start=1
                     /label=GAL4 activation domain
                     /note="activation domain of the GAL4 transcriptional
                     activator"
                     /translation="ANFNQSGNIADSSLSFTFTNSSNGPNLITTQTNSQALSQPIASSN
                     VHDNFMNNEITASKIDDGNNSKPLSPGWTDQTAYNAFGITTGMFNTTTMDDVYNYLFDD
                     EDTPPNPKKE"
     promoter        complement(905..923)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     terminator      1310..1497
                     /label=ADH1 terminator
                     /note="transcription terminator for the S. cerevisiae
                     alcohol dehydrogenase 1 (ADH1) gene"
     CDS             complement(1618..2709)
                     /codon_start=1
                     /label=LEU2
                     /note="3-isopropylmalate dehydrogenase, required for
                     leucine biosynthesis"
                     /translation="MSAPKKIVVLPGDHVGQEITAEAIKVLKAISDVRSNVKFDFENHL
                     IGGAAIDATGVPLPDEALEASKKVDAVLLGAVGGPKWGTGSVRPEQGLLKIRKELQLYA
                     NLRPCNFASDSLLDLSPIKPQFAKGTDFVVVRELVGGIYFGKRKEDDGDGVAWDSEQYT
                     VPEVQRITRMAAFMALQHEPPLPIWSLDKANVLASSRLWRKTVEETIKNEFPTLKVQHQ
                     LIDSAAMILVKNPTHLNGIIITSNMFGDIISDEASVIPGSLGLLPSASLASLPDKNTAF
                     GLYEPCHGSAPDLPKNKVDPIATILSAAMMLKLSLNLPEEGKAIEDAVKKVLDAGIRTG
                     DLGGSNSTTEVGDAVAEEVKKILA"
     promoter        complement(2710..3114)
                     /label=LEU2 promoter
     primer_bind     complement(3156..3172)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    complement(3180..3196)
                     /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)."
     promoter        complement(3204..3234)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    complement(3249..3270)
                     /label=CAP binding site
                     /note="CAP binding activates transcription in the presence
                     of cAMP."
     rep_origin      complement(3351..3806)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow
                     indicates direction of (+) strand synthesis"
     promoter        3833..3937
                     /label=AmpR promoter
     enhancer        3939..4130
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     rep_origin      complement(4483..5071)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     CDS             complement(5245..6102)
                     /codon_start=1
                     /label=AmpR
                     /note="beta-lactamase"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     promoter        complement(6103..6207)
                     /label=AmpR promoter
     rep_origin      6489..7653
                     /label=2u ori
                     /note="yeast 2u plasmid origin of replication"