pUC19-ACMV 载体 (V015684)

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:
pUC19-ACMV
载体抗性:
Ampicillin
载体长度:
6876 bp
载体类型:
Cloning vector
复制子:
ori
载体来源:
Hoyer JS, Fondong VN, Dallas MM, Aimone CD

pUC19-ACMV 载体图谱

pUC19-ACMV6876 bp3006009001200150018002100240027003000330036003900420045004800510054005700600063006600oriCAP binding sitelac promoterlac operatorM13 revACMV DNA-B monomer unit, nick site to nick siteM13 fwdAmpR promoterAmpR

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。

pUC19-ACMV 载体序列

LOCUS       62056_21980        6876 bp DNA     circular SYN 18-NOV-2020
DEFINITION  Cloning vector pUC19 ACMV DNA-B 1.5mer, complete sequence.
ACCESSION   MT856194
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6876)
  AUTHORS   Hoyer JS, Fondong VN, Dallas MM, Aimone CD, Deppong DO, Duffy S, 
            Hanley-Bowdoin L.
  TITLE     Deeply Sequenced Infectious Clones of Key Cassava Begomovirus 
            Isolates from Cameroon
  JOURNAL   Microbiol Resour Announc 9 (46), e00802-20 (2020)
  PUBMED    33184153
REFERENCE   2  (bases 1 to 6876)
  AUTHORS   Hoyer JS, Fondong VN, Dallas MM, Aimone CD, Deppong DO, Duffy S, 
            Hanley-Bowdoin L.
  TITLE     Direct Submission
  JOURNAL   Submitted (06-AUG-2020) Ecology, Evolution, and Natural Resources, 
            Rutgers University, 14 College Farm Rd, New Brunswick, NJ 08901, USA
REFERENCE   3  (bases 1 to 6876)
  AUTHORS   .
  TITLE     Direct Submission
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Microbiol 
            Resour Announc"; date: "2020"; volume: "9"; issue: "46"; pages: 
            "e00802-20"
COMMENT     SGRef: number: 2; type: "Journal Article"; journalName: "Submitted 
            (06-AUG-2020) Ecology, Evolution, and Natural Resources, Rutgers 
            University, 14 College Farm Rd, New Brunswick, NJ 08901, USA"
COMMENT     ##Assembly-Data-START##
            Assembly Method       :: Benchling v. 2019-01-22
            Sequencing Technology :: Illumina; Sanger dideoxy sequencing 
            ##Assembly-Data-END##
FEATURES             Location/Qualifiers
     source          1..6876
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      171..759
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     protein_bind    1047..1068
                     /label=CAP binding site
                     /note="CAP binding activates transcription in the presence
                     of cAMP."
     promoter        1083..1113
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    1121..1137
                     /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)."
     primer_bind     1145..1161
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     misc_feature    1174..2656
                     /note="duplicated part of ACMV DNA-B partial tandem dimer, 
                     HindIII to PstI inclusive, copy 1"
     misc_feature    2558..5282
                     /label=ACMV DNA-B monomer unit, nick site to nick site
                     /note="ACMV DNA-B monomer unit, nick site to nick site"
     misc_feature    2657..3898
                     /note="unique part of ACMV DNA-B partial tandem dimer, PstI
                     to HindIII, noninclusive"
     gene            2992..3762
                     /gene="BV1"
                     /label=BV1
     CDS             2992..3762
                     /codon_start=1
                     /transl_table=11
                     /gene="BV1"
                     /product="NSP"
                     /label=BV1
                     /protein_id="QNR00651.1"
                     /translation="MYSIRKQSRNLQRKYNSNTTNRYPIRRKYVAGHTRPCVRRRLSYE
                     PVERPLVHNVLCEKQHGDVFNLQQNTSYTSFVTYPARGSSGDGRSRDYIKLQSMSVSGV
                     IHAKADGNDDPMELSQVVNGVFVFSLIMDTKPYLPAGVQALPTFEELFGPYSACYVNLR
                     LLNNQQHRYRVLHSVKRFVSSSGDTKVSQFRFNKRLSTRRYTIWASFHDVDLVNAGGNY
                     RNISKNAILVSYAFVSEHAMSCKPFVQIETSYVG"
     gene            complement(3771..4667)
                     /gene="BC1"
                     /label=BC1
     CDS             complement(3771..4667)
                     /codon_start=1
                     /transl_table=11
                     /gene="BC1"
                     /product="MP"
                     /label=BC1
                     /protein_id="QNR00652.1"
                     /translation="MDTSVPVISSDYIHSARTEYKLTNDESPITLQFPSTLERTRVRIM
                     GKCMKVDHVVIEYRNQVPFNAQGSVIVTIRDTRLSDEQQDQAQFTFPIGCNVDLHYFSA
                     SYFSIDDNVPWQLLYKVEDSNVKNGITFAQIKAKLKLSAAKHSTDIKFKQPTIKILSKD
                     YGPDCVDFWSVGKPKPIRRLIQNEPGTDYDTGPKYRPITVQPGETWATKSTIGRSTSMR
                     YTGPQPIDIDASSSKQYASEAEFPLRGLHRLPEASLDPGDSVSQTQSMSKKDIESIIEQ
                     TVNKCLIAHRGSSHKDL"
     misc_feature    3899..5381
                     /note="duplicated part of ACMV DNA-B partial tandem dimer, 
                     HindIII to PstI inclusive, copy 2"
     primer_bind     complement(5421..5437)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     promoter        5911..6015
                     /label=AmpR promoter
     CDS             6016..6873
                     /label=AmpR
                     /note="beta-lactamase"