pACYCDuet-1

pACYCDuet-1

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pACYCDuet-1,Plasmid pACYCDuet-1,pACYCDuet-1 vector

pACYCDuet-1 plasmid information
Promoter: T7/Lac
Replicon: p15A ori
Terminator: T7 Terminator
Plasmid classification: large intestine series plasmid; other large intestine plasmid; large intestine double frame plasmid
Plasmid size: 4008bp
Plasmid Tags: N-6 x His, C-S
Prokaryotic resistance: chloramphenicol Chl (50 g/ml)
Clone strain: DH5 alpha and other Escherichia coli
Culture conditions: 37, aerobic, LB
Expression host: BL21 (DE3) and other Escherichia coli
Culture conditions: 37, aerobic, LB
Induction: IPTG or lactose and its analogues
Primers for 5'sequencing: ACYCDuetUP1 (GGATCTCGACGCTCTCCCT) DuetUP2 (TTGTACACGGCCGCATAATC)

pACYCDuet-1 plasmid  Description
pACYCDuet-1 is designed for the coexpression of two target genes. The vector contains two multiple cloning sites (MCS), each of which is preceded by a T7 promoter/lac operator and ribosome binding site (rbs). The vector also carries the P15A replicon, lacI gene and chloramphenicol resistance gene. This vector can be used in combination with pETDuet™-1 in an appropriate host strain for the coexpression of up to 4 target genes. Genes inserted into MCS1 can be sequenced using the ACYCDuetUP1 Primer and DuetDOWN1 Primer . Genes inserted into MCS2 can be sequenced using the DuetUP2 Primer and T7 Terminator Primer .
 

pACYCDuet-1 plasmid Sequence
LOCUS       Exported                4008 bp ds-DNA     circular SYN 27-8-2015
DEFINITION  synthetic circular DNA
KEYWORDS    Untitled
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4008)
  TITLE     Direct Submission
  JOURNAL   Exported 2015-8-27   
FEATURES             Location/Qualifiers
     source          1..4008
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     protein_bind    3..27
                     /bound_moiety="lac repressor encoded by lacI"
                     /note="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)."
     CDS             83..100
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /note="6xHis"
                     /translation="HHHHHH"
     promoter        214..232
                     /note="T7 promoter"
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    233..257
                     /bound_moiety="lac repressor encoded by lacI"
                     /note="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)."
     CDS             366..410
                     /codon_start=1
                     /product="affinity and epitope tag derived from pancreatic
                     ribonuclease A"
                     /note="S-Tag"
                     /translation="KETAAAKFERQHMDS"
     terminator      462..509
                     /note="T7 terminator"
                     /note="transcription terminator for bacteriophage T7 RNA
                     polymerase"
     CDS             complement(729..1388)
                     /codon_start=1
                     /gene="cat"
                     /product="chloramphenicol acetyltransferase"
                     /note="CmR"
                     /note="confers resistance to chloramphenicol"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA"
     promoter        complement(1389..1491)
                     /note="cat promoter"
                     /note="promoter of the E. coli cat gene encoding
                     chloramphenicol acetyltransferase"
     rep_origin      complement(2017..2561)
                     /direction=LEFT
                     /note="p15A ori"
                     /note="Plasmids containing the medium-copy-number p15A
                     origin of replication can be propagated in E. coli cells
                     that contain a second plasmid with the ColE1 origin."
     CDS             complement(2786..3868)
                     /codon_start=1
                     /gene="lacI"
                     /product="lac repressor"
                     /note="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)."
                     /translation="MKPVTLYDVAEYAGVSYQTVSRVVNQASHVSAKTREKVEAAMAEL
                     NYIPNRVAQQLAGKQSLLIGVATSSLALHAPSQIVAAIKSRADQLGASVVVSMVERSGV
                     EACKAAVHNLLAQRVSGLIINYPLDDQDAIAVEAACTNVPALFLDVSDQTPINSIIFSH
                     EDGTRLGVEHLVALGHQQIALLAGPLSSVSARLRLAGWHKYLTRNQIQPIAEREGDWSA
                     MSGFQQTMQMLNEGIVPTAMLVANDQMALGAMRAITESGLRVGADISVVGYDDTEDSSC
                     YIPPLTTIKQDFRLLGQTSVDRLLQLSQGQAVKGNQLLPVSLVKRKTTLAPNTQTASPR
                     ALADSLMQLARQVSRLESGQ"
     promoter        complement(3869..3946)
                     /gene="lacI"
                     /note="lacI promoter"
     promoter        3992..2
                     /note="T7 promoter"
                     /note="promoter for bacteriophage T7 RNA polymerase"
ORIGIN
        1 ggggaattgt gagcggataa caattcccct gtagaaataa ttttgtttaa ctttaataag
       61 gagatatacc atgggcagca gccatcacca tcatcaccac agccaggatc cgaattcgag
      121 ctcggcgcgc ctgcaggtcg acaagcttgc ggccgcataa tgcttaagtc gaacagaaag
      181 taatcgtatt gtacacggcc gcataatcga aattaatacg actcactata ggggaattgt
      241 gagcggataa caattcccca tcttagtata ttagttaagt ataagaagga gatatacata
      301 tggcagatct caattggata tcggccggcc acgcgatcgc tgacgtcggt accctcgagt
      361 ctggtaaaga aaccgctgct gcgaaatttg aacgccagca catggactcg tctactagcg
      421 cagcttaatt aacctaggct gctgccaccg ctgagcaata actagcataa ccccttgggg
      481 cctctaaacg ggtcttgagg ggttttttgc tgaaacctca ggcatttgag aagcacacgg
      541 tcacactgct tccggtagtc aataaaccgg taaaccagca atagacataa gcggctattt
      601 aacgaccctg ccctgaaccg acgaccgggt cgaatttgct ttcgaatttc tgccattcat
      661 ccgcttatta tcacttattc aggcgtagca ccaggcgttt aagggcacca ataactgcct
      721 taaaaaaatt acgccccgcc ctgccactca tcgcagtact gttgtaattc attaagcatt
      78  <