pYD1 载体 (V012745) COA报告测序报告

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。

pYD1 is a 5.0 kb expression vector designed for expression, secretion, and display of proteins on the extracellular surface of Saccharomyces cerevisiae cells. The vector contains the following elements:1. AGA2 gene from Saccharomyces cerevisiae. This gene encodes one of the subunits of the a-agglutinin receptor. Fusion of the gene of interest to AGA2 allows secretion and display of the protein of interest. 2. GAL1 promoter for regulated expression of the AGA2 gene fusion. 3. Xpress epitope and V5 epitope for detection of the displayed protein. 4. Polyhistidine (6xHis) tag for detection and possible purification on metal chelating resin. 5. TRP1 gene for selection in Saccharomyces cerevisiae. 6. CEN6/ARS4 for stable, episomal replication in yeast. 7. Ampicillin resistance gene and the pUC origin for selection and replication in E. coli.

载体名称:: pYD1

载体抗性:: Ampicillin

载体长度:: 5009 bp

载体类型:: Yeast Plasmids

复制子:: ori

载体来源:: Invitrogen

筛选标记:: TRP1

拷贝数:: High copy number

启动子:: GAL1

克隆方法:: Enzyme Cut

5'引物:: pYD1-F: AGTAACGTTTGTCAGTAATTGC

3'引物:: pYD1-R: GTCGATTTTGTTACATCTACAC

感受态:: DH10B

培养温度:: 37℃

pYD1 载体图谱

复制序列 NovoBuilder中查看图谱

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。

pYD1 载体序列

下载GenBank文件(.gb)

LOCUS       Exported                5009 bp DNA     circular SYN 03-SEP-2024
DEFINITION  Exported.
ACCESSION   V012745
VERSION     .
KEYWORDS    pYD1
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5009)
  TITLE     Direct Submission
REFERENCE   2  (bases 1 to 5009)
  TITLE     Direct Submission
REFERENCE   3  (bases 1 to 5009)
  AUTHORS   .
  TITLE     Direct Submission
COMMENT     SGRef: number: 1; type: "Journal Article"
COMMENT     SGRef: number: 2; type: "Journal Article"
FEATURES             Location/Qualifiers
     source          1..5009
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        2..443
                     /label=GAL1 promoter
                     /note="inducible promoter, regulated by Gal4"
     promoter        475..493
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             536..796
                     /gene="AGA2"
                     /label=A-agglutinin-binding subunit
                     /note="A-agglutinin-binding subunit from Saccharomyces 
                     cerevisiae (strain ATCC 204508 / S288c). Accession#: 
                     P32781"
     CDS             860..889
                     /codon_start=1
                     /product="leader peptide from bacteriophage T7 gene 10"
                     /label=leader peptide from bacteriophage T7 gene 10
                     /note="T7 tag (gene 10 leader)"
                     /note="promotes efficient translation in E. coli"
                     /translation="ASMTGGQQMG"
     CDS             893..916
                     /label=Xpress(TM) tag
                     /note="Xpress(TM) epitope tag, including an enterokinase 
                     recognition and cleavage site"
     CDS             995..1036
                     /label=V5 tag
                     /note="epitope tag from simian virus 5"
     CDS             1046..1063
                     /label=6xHis
                     /note="6xHis affinity tag"
     CDS             complement(1437..2108)
                     /label=TRP1
                     /note="phosphoribosylanthranilate isomerase, required for 
                     tryptophan biosynthesis"
     promoter        complement(2109..2210)
                     /label=TRP1 promoter
     misc_feature    complement(2287..2790)
                     /label=CEN/ARS
                     /note="S. cerevisiae CEN6 centromere fused to an
                     autonomously replicating sequence"
     promoter        2827..2931
                     /label=AmpR promoter
     CDS             2932..3789
                     /label=AmpR
                     /note="beta-lactamase"
     rep_origin      3963..4551
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     protein_bind    4839..4860
                     /label=CAP binding site
                     /note="CAP binding activates transcription in the presence
                     of cAMP."
     promoter        4875..4905
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4913..4929
                     /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     4937..4953
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     promoter        4974..4992
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"

pYD1 载体 (V012745) .cls { stroke-width: 0px; fill: url(#c6); } COA报告 测序报告

pYD1 载体图谱

质粒操作方法

pYD1 载体序列

pYD1 载体 (V012745) COA报告测序报告