LOCUS       40924_36793        6293 bp DNA     circular SYN 18-DEC-2018
DEFINITION  Retrofitting vector pRetroES, complete sequence.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6293)
  AUTHORS   Wang Z, Engler P, Longacre A, Storb U.
  TITLE     An efficient method for high-fidelity BAC/PAC retrofitting with a 
            selectable marker for mammalian cell transfection
  JOURNAL   Genome Res. 11 (1), 137-142 (2001)
  PUBMED    11156622
REFERENCE   2  (bases 1 to 6293)
  AUTHORS   Wang Z, Engler P, Longacre A, Storb U.
  TITLE     Direct Submission
  JOURNAL   Submitted (05-JUL-2001) Mol. Genet. Cell Biol., University of 
            Chicago, 920 E. 58th St., Chicago, IL 60637, USA
REFERENCE   3  (bases 1 to 6293)
  TITLE     Direct Submission
REFERENCE   4  (bases 1 to 6293)
  AUTHORS   .
  TITLE     Direct Submission
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Genome 
            Res."; date: "2001"; volume: "11"; issue: "1"; pages: "137-142"
COMMENT     SGRef: number: 2; type: "Journal Article"; journalName: "Submitted 
            (05-JUL-2001) Mol. Genet. Cell Biol., University of Chicago, 920 E. 
            58th St., Chicago, IL 60637, USA"
COMMENT     SGRef: number: 3; type: "Journal Article"
FEATURES             Location/Qualifiers
     source          1..6293
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        470..498
                     /label=tac promoter
                     /note="strong E. coli promoter; hybrid between the trp and
                     lac UV5 promoters"
     protein_bind    506..522
                     /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)."
     CDS             561..1205
                     /codon_start=1
                     /label=GST
                     /note="glutathione S-transferase from Schistosoma
                     japonicum"
                     /translation="MLGYWKIKGLVQPTRLLLEYLEEKYEEHLYERDEGDKWRNKKFEL
                     GLEFPNLPYYIDGDVKLTQSMAIIRYIADKHNMLGGCPKERAEISMLEGAVLDIRYGVS
                     RIAYSKDFETLKVDFLSKLPEMLKMFEDRLCHKTYLNGDHVTHPDFMLYDALDVVLYMD
                     PMCLDAFPKLVCFKKRIEAIPQIDKYLKSSKYIAWPLQGWQATFGGGDHPPK"
     CDS             1212..1229
                     /codon_start=1
                     /label=thrombin site
                     /note="thrombin recognition and cleavage site"
                     /translation="LVPRGS"
     protein_bind    complement(1266..1299)
                     /label=loxP
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     CDS             1332..2360
                     /codon_start=1
                     /label=Cre
                     /note="site-specific recombinase"
                     /translation="MANLLTVHQNLPALPVDATSDEVRKNLMDMFRDRQAFSEHTWKML
                     LSVCRSWAAWCKLNNRKWFPAEPEDVRDYLLYLQARGLAVKTIQQHLGQLNMLHRRSGL
                     PRPSDSNAVSLVMRRIRKENVDAGERAKQALAFERTDFDQVRSLMENSDRCQDIRNLAF
                     LGIAYNTLLRIAEIARIRVKDISRTDGGRMLIHIGRTKTLVSTAGVEKALSLGVTKLVE
                     RWISVSGVADDPNNYLFCRVRKNGVAAPSATSQLSTRALEGIFEATHRLIYGAKDDSGQ
                     RYLAWSGHSARVGAARDMARAGVSIPEIMQAGGWTNVNIVMNYIRNLDSETGAMVRLLE
                     DGD"
     terminator      2463..2510
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     polyA_signal    complement(2587..3048)
                     /label=PGK poly(A) signal
                     /note="mouse phosphoglycerate kinase 1 polyadenylation
                     signal"
     CDS             complement(3067..3867)
                     /codon_start=1
                     /label=NeoR/KanR
                     /note="aminoglycoside phosphotransferase from Tn5"
                     /translation="MGSAIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQ
                     GRPVLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQD
                     LLSSHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDE
                     EHQGLAPAELFARLKARMPDGDDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQ
                     DIALATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     promoter        complement(3881..4377)
                     /label=PGK promoter
                     /note="mouse phosphoglycerate kinase 1 promoter"
     rep_origin      4509..4897
                     /label=R6K gamma ori
                     /note="gamma replication origin from E. coli plasmid R6K;
                     requires the R6K initiator protein pi for replication"
     CDS             complement(5035..5892)
                     /codon_start=1
                     /label=AmpR
                     /note="beta-lactamase"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERSPMMSTFKVLLCGAVLSRVDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     promoter        complement(5893..5997)
                     /label=AmpR promoter
     primer_bind     6218..6234
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
ORIGIN
        1 ggccgcgtgg cgaatggcgc tttgcctggt ttccggcacc agaagcggtg ccggaaagct
       61 ggctggagtg cgatcttcct gaggccgata ctgtcgtcgt cccctcaaac tggcagatgc
      121 acggttacga tgcgcccatc tacaccaacg taacctatcc cattacggtc aatccgccgt
      181 ttgttcccac ggagaatccg acgggttgtt actcgctcac atttaatgtt gatgaaagct
      241 ggctacagga aggccagacg cgaattattt ttgatggcgt tggaattagc ttatcgactg
      301 cacggtgcac caatgcttct ggcgtcaggc agccatcgga agctgtggta tggctgtgca
      361 ggtcgtaaat cactgcataa ttcgtgtcgc tcaaggcgca ctcccgttct ggataatgtt
      421 ttttgcgccg acatcataac ggttctggca aatattctga aatgagctgt tgacaattaa
      481 tcatcggctc gtataatgtg tggaattgtg agcggataac aatttcacac aggaaacagt
      541 tctagagtcc cctatgtaga atgctaggtt attggaaaat taagggcctt gtgcaaccca
      601 ctcgacttct tttggaatat cttgaagaaa aatatgaaga gcatttgtat gagcgcgatg
      661 aaggtgataa atggcgaaac aaaaagtttg aattgggttt ggagtttccc aatcttcctt
      721 attatattga tggtgatgtt aaattaacac agtctatggc catcatacgt tatatagctg
      781 acaagcacaa catgttgggt ggttgtccaa aagagcgtgc agagatttca atgcttgaag
      841 gagcggtttt ggatattaga tacggtgttt cgagaattgc atatagtaaa gactttgaaa
      901 ctctcaaagt tgattttctt agcaagctac ctgaaatgct gaaaatgttc gaagatcgtt
      961 tatgtcataa aacatattta aatggtgatc atgtaaccca tcctgacttc atgttgtatg
     1021 acgctcttga tgttgtttta tacatggacc caatgtgcct ggatgcgttc ccaaaattag
     1081 tttgttttaa aaaacgtatt gaagctatcc cacaaattga taagtacttg aaatccagca
     1141 agtatatagc atggcctttg cagggctggc aagccacgtt tggtggtggc gaccatcctc
     1201 caaaatcgga tctggttccg cgtggatctc gtcgtgcatc tgttggatcg catatgccca
     1261 tggctataac ttcgtatagc atacattata cgaagttatc tggaattccc cgggctcgag
     1321 aacatatggc catggccaat ttactgaccg tacaccaaaa tttgcctgca ttaccggtcg
     1381 atgcaacgag tgatgaggtt cgcaagaacc tgatggacat gttcagggat cgccaggcgt
     1441 tttctgagca tacctggaaa atgcttctgt ccgtttgccg gtcgtgggcg gcatggtgca
     1501 agttgaataa ccggaaatgg tttcccgcag aacctgaaga tgttcgcgat tatcttctat
     1561 atcttcaggc gcgcggtctg gcagtaaaaa ctatccagca acatttgggc cagctaaaca
     1621 tgcttcatcg tcggtccggg ctgccacgac caagtgacag caatgctgtt tcactggtta
     1681 tgcggcggat ccgaaaagaa aacgttgatg ccggtgaacg tgcaaaacag gctctagcgt
     1741 tcgaacgcac tgatttcgac caggttcgtt cactcatgga aaatagcgat cgctgccagg
     1801 atatacgtaa tctggcattt ctggggattg cttataacac cctgttacgt atagccgaaa
     1861 ttgccaggat cagggttaaa gatatctcac gtactgacgg tgggagaatg ttaatccata
     1921 ttggcagaac gaaaacgctg gttagcaccg caggtgtaga gaaggcactt agcctggggg
     1981 taactaaact ggtcgagcga tggatttccg tctctggtgt agctgatgat ccgaataact
     2041 acctgttttg ccgggtcaga aaaaatggtg ttgccgcgcc atctgccacc agccagctat
     2101 caactcgcgc cctggaaggg atttttgaag caactcatcg attgatttac ggcgctaagg
     2161 atgactctgg tcagagatac ctggcctggt ctggacacag tgcccgtgtc ggagccgcgc
     2221 gagatatggc ccgcgctgga gtttcaatac cggagatcat gcaagctggt ggctggacca
     2281 atgtaaatat tgtcatgaac tatatccgta acctggatag tgaaacaggg gcaatggtgc
     2341 gcctgctgga agatggcgat tagcccggga atttagcagg catgctgggg attctagaag
     2401 atccggctgc taacaaagcc cgaaaggaag ctgagttggc tgctgccacc gctgagcaat
     2461 aactagcata accccttggg gcctctaaac gggtcttgag gggttttttg ctgaaaggag
     2521 gaactatatc cgggctgcag gaattaattc gatatcaagc ttcgtggatc cagatctaga
     2581 gtcagcttct gatggaatta gaacttggca aaacaatact gagaatgaag tgtatgtgga
     2641 acagaggctg ctgatctcgt tcttcaggct atgaaactga cacatttgga aaccacagta
     2701 cttagaacca caaagtggga atcaagagaa aaacaatgat cccacgagag atctatagat
     2761 ctatagatca tgagtgggag gaatgagctg gcccttaatt tggttttgct tgtttaaatt
     2821 atgatatcca actatgaaac attatcataa agcaatagta aagagccttc agtaaagagc
     2881 aggcatttat ctaatcccac cccaccccca cccccgtagc tccaatcctt ccattcaaaa
     2941 tgtaggtact ctgttctcac ccttcttaac aaagtatgac aggaaaaact tccattttag
     3001 tggacatctt tattgtttaa tagatcatca atttctgcag acttacagga cggatcgatc
     3061 ccctcagaag aactcgtcaa gaaggcgata gaaggcgatg cgctgcgaat cgggagcggc
     3121 gataccgtaa agcacgagga agcggtcagc ccattcgccg ccaagctctt cagcaatatc
     3181 acgggtagcc aacgctatgt cctgatagcg gtccgccaca cccagccggc cacagtcgat
     3241 gaatccagaa aagcggccat tttccaccat gatattcggc aagcaggcat cgccatgggt
     3301 cacgacgaga tcatcgccgt cgggcatgcg cgccttgagc ctggcgaaca gttcggctgg
     3361 cgcgagcccc tgatgctctt cgtccagatc atcctgatcg acaagaccgg cttccatccg
     3421 agtacgtgct cgctcgatgc gatgtttcgc ttggtggtcg aatgggcagg tagccggatc
     3481 aagcgtatgc agccgccgca ttgcatcagc catgatggat actttctcgg caggagcaag
     3541 gtgagatgac aggagatcct gccccggcac ttcgcccaat agcagccagt cccttcccgc
     3601 ttcagtgaca acgtcgagca cagctgcgca aggaacgccc gtcgtggcca gccacgatag
     3661 ccgcgctgcc tcgtcctgca gttcattcag ggcaccggac aggtcggtct tgacaaaaag
     3721 aaccgggcgc ccctgcgctg acagccggaa cacggcggca tcagagcagc cgattgtctg
     3781 ttgtgcccag tcatagccga atagcctctc cacccaagcg gccggagaac ctgcgtgcaa
     3841 tccatcttgt tcaatggccg atcccatatt ggctgcaggt cgaaaggccc ggagatgagg
     3901 aagaggagaa cagcgcggca gacgtgcgct tttgaagcgt gcgagaatgc cgggcctcgg
     3961 gaggaccttc gcgcccgccc cgcccctgag cccgcccctg agcccgcccc cggacccacc
     4021 ccttcccagc ctctgagccc agaaagcgaa ggagcaaagc tgctattggc cgctgcccca
     4081 aaggcctacc cgcttccatt gctcagcggt gctgtccatc tgcacgagac tagtgagacg
     4141 tgctacttcc atttgtcacg tcctgcacga cgcgagctgc ggggcggggg ggaacttcct
     4201 gactagggga ggagtagaag gtggcgcgaa ggggccacca aagaacggag ccggttggcg
     4261 ctaccggtgg atgtggaatg tgtgcgaggc cagaggccac ttgtgtagcg ccaagtgcca
     4321 gcggggctgc taaagcgcat gctccagact gccttgggaa aagcgcctcc cctacccggt
     4381 agaattgacc tgcaggggcc ctcgaatcct gcaggaattt taacaaaata ttaacgctta
     4441 caatttaggt ggcacttttc ggggaaatgt gcgcggaacc agatccccac gatgcgtccg
     4501 gcgtagagga tctgaagatc agcagttcaa cctgttgata gtacgtacta agctctcatg
     4561 tttcacgtac taagctctca tgtttaacgt actaagctct catgtttaac gaactaaacc
     4621 ctcatggcta acgtactaag ctctcatggc taacgtacta agctctcatg tttcacgtac
     4681 taagctctca tgtttgaaca ataaaattaa tataaatcag caacttaaat agcctctaag
     4741 gttttaagtt ttataagaaa aaaaagaata tataaggctt ttaaagcttt taaggtttaa
     4801 cggttgtgga caacaagcca gggatgtaac gcactgagaa gcccttagag cctctcaaag
     4861 caattttgag tgacacagga acacttaacg gctgacatgg gaattttaac aaaatattaa
     4921 cgtttacaat ttaaatattt gcttatacaa tcttcctgtt tttggggctt ttctgattat
     4981 caaccggggt aaatcaatct aaagtatata tgagtaaact tggtctgaca gttaccaatg
     5041 cttaatcagt gaggcaccta tctcagcgat ctgtctattt cgttcatcca tagttgcctg
     5101 actccccgtc gtgtagataa ctacgatacg ggagggctta ccatctggcc ccagtgctgc
     5161 aatgataccg cgagacccac gctcaccggc tccagattta tcagcaataa accagccagc
     5221 cggaagggcc gagcgcagaa gtggtcctgc aactttatcc gcctccatcc agtctattaa
     5281 ttgttgccgg gaagctagag taagtagttc gccagttaat agtttgcgca acgttgttgc
     5341 cattgctaca ggcatcgtgg tgtcacgctc gtcgtttggt atggcttcat tcagctccgg
     5401 ttcccaacga tcaaggcgag ttacatgatc ccccatgttg tgcaaaaaag cggttagctc
     5461 cttcggtcct ccgatcgttg tcagaagtaa gttggccgca gtgttatcac tcatggttat
     5521 ggcagcactg cataattctc ttactgtcat gccatccgta agatgctttt ctgtgactgg
     5581 tgagtactca accaagtcat tctgagaata gtgtatgcgg cgaccgagtt gctcttgccc
     5641 ggcgtcaaca cgggataata ccgcgccaca tagcagaact ttaaaagtgc tcatcattgg
     5701 agaacgttct tcggggcgaa aactctcaag gatcttaccg ctgttgagat ccagttcgat
     5761 gtaacccact cgtgcaccca actgatcttc agcatctttt actttcacca gcgtttctgg
     5821 gtgagcaaaa acaggaaggc aaaatgccgc aaaaaaggga ataagggcga cacggaaatg
     5881 ttgaatactc atactcttcc tttttcaata ttattgaagc atttatcagg gttattgtct
     5941 catgagcgga tacatatttg aatgtattta gaaaaataaa caaatagggg ttccgcgcac
     6001 atttccccga aaagtgccac ctgacgtagt taacaaaaaa aagcccgccg aagcgggctt
     6061 tattaccaag cgaagcgcca ttcgccattc aggctgcgca actgttggga agggcgatcg
     6121 gtgcgggcct cttcgctatt acgccagctg gcgaaagggg gatgtgctgc aaggcgatta
     6181 agttgggtaa cgccagggtt ttcccagtca cgacgttgta aaacgacggc cagtccgtaa
     6241 tacgactcac ttaaggcctt gactagaggg taccagagct ccaccgcggt ggc
//
