LOCUS       62056_3400        8274 bp DNA     circular SYN 01-JAN-1980
DEFINITION  synthetic circular DNA.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8274)
  AUTHORS   .
  TITLE     Direct Submission
FEATURES             Location/Qualifiers
     source          1..8274
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        complement(1..105)
                     /label=AmpR promoter
     rep_origin      complement(131..586)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow
                     indicates direction of (+) strand synthesis"
     primer_bind     728..744
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     promoter        1382..1760
                     /label=TRE3G promoter
                     /note="3rd-generation Tet-responsive promoter that can be 
                     activated by binding of Tet-On(R) 3G"
     CDS             1873..2589
                     /codon_start=1
                     /label=EGFP
                     /note="enhanced GFP"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKPPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     polyA_signal    2673..2721
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase
                     polyadenylation signal (Cole and Stacy, 1985)"
     promoter        2952..3163
                     /label=EF-1-alpha core promoter
                     /note="core promoter for human elongation factor 
                     EF-1-alpha"
     LTR             3176..3444
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from human
                     T-cell leukemia virus (HTLV) type 1"
     CDS             3478..4221
                     /codon_start=1
                     /label=Tet-On(R) 3G
                     /note="modified rtTA protein that binds tightly to
                     promoters containing the tet operator in the presence of 
                     doxycycline"
                     /translation="MSRLDKSKVINSALELLNGVGIEGLTTRKLAQKLGVEQPTLYWHV
                     KNKRALLDALPIEMLDRHHTHSCPLEGESWQDFLRNNAKSYRCALLSHRDGAKVHLGTR
                     PTEKQYETLENQLAFLCQQGFSLENALYALSAVGHFTLGCVLEEQEHQVAKEERETPTT
                     DSMPPLLKQAIELFDRQGAEPAFLFGLELIICGLEKQLKCESGGPTDALDDFDLDMLPA
                     DALDDFDLDMLSADALDDFDLDMLPG"
     CDS             4222..4275
                     /codon_start=1
                     /label=T2A
                     /note="2A peptide from Thosea asigna virus capsid protein"
                     /translation="EGRGSLLTCGDVEENPGP"
     CDS             4282..4878
                     /codon_start=1
                     /label=PuroR
                     /note="puromycin N-acetyltransferase"
                     /translation="MTEYKPTVRLATRDDVPRAVRTLAAAFADYPATRHTVDPDRHIER
                     VTELQELFLTRVGLDIGKVWVADDGAAVAVWTTPESVEAGAVFAEIGPRMAELSGSRLA
                     AQQQMEGLLAPHRPKEPAWFLATVGVSPDHQGKGLGSAVVLPGVEAAERAGVPAFLETS
                     APRNLPFYERLGFTVTADVEVPEGPRTWCMTRKPGA"
     polyA_signal    4934..5055
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(6253..6269)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    complement(6277..6293)
                     /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(6301..6331)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    complement(6346..6367)
                     /label=CAP binding site
                     /note="CAP binding activates transcription in the presence
                     of cAMP."
     rep_origin      complement(6655..7243)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     CDS             complement(7417..8274)
                     /codon_start=1
                     /label=AmpR
                     /note="beta-lactamase"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
ORIGIN
        1 actcttcctt tttcaatatt attgaagcat ttatcagggt tattgtctca tgagcggata
       61 catatttgaa tgtatttaga aaaataaaca aataggggtt ccgcgcacat ttccccgaaa
      121 agtgccacct aaattgtaag cgttaatatt ttgttaaaat tcgcgttaaa tttttgttaa
      181 atcagctcat tttttaacca ataggccgaa atcggcaaaa tcccttataa atcaaaagaa
      241 tagaccgaga tagggttgag tgttgttcca gtttggaaca agagtccact attaaagaac
      301 gtggactcca acgtcaaagg gcgaaaaacc gtctatcagg gcgatggccc actacgtgaa
      361 ccatcaccct aatcaagttt tttggggtcg aggtgccgta aagcactaaa tcggaaccct
      421 aaagggagcc cccgatttag agcttgacgg ggaaagccgg cgaacgtggc gagaaaggaa
      481 gggaagaaag cgaaaggagc gggcgctagg gcgctggcaa gtgtagcggt cacgctgcgc
      541 gtaaccacca cacccgccgc gcttaatgcg ccgctacagg gcgcgtccca ttcgccattc
      601 aggctgcgca actgttggga agggcgatcg gtgcgggcct cttcgctatt acgccagctg
      661 gcgaaagggg gatgtgctgc aaggcgatta agttgggtaa cgccagggtt ttcccagtca
      721 cgacgttgta aaacgacggc cagtgagcgc gcctcgttca ttcacgtttt tgaacccgtg
      781 gaggacgggc agactcgcgg tgcaaatgtg ttttacagcg tgatggagca gatgaagatg
      841 ctcgacacgc tgcagaacac gcagctagat taaccctaga aagataatca tattgtgacg
      901 tacgttaaag ataatcatgc gtaaaattga cgcatgtgtt ttatcggtct gtatatcgag
      961 gtttatttat taatttgaat agatattaag ttttattata tttacactta catactaata
     1021 ataaattcaa caaacaattt atttatgttt atttatttat taaaaaaaaa caaaaactca
     1081 aaatttcttc tataaagtaa caaaactttt atgagggaca gccccccccc aaagccccca
     1141 gggatgtaat tacgtccctc ccccgctagg gggcagcagc gagccgcccg gggctccgct
     1201 ccggtccggc gctccccccg catccccgag ccggcagcgt gcggggacag cccgggcacg
     1261 gggaaggtgg cacgggatcg ctttcctctg aacgcttctc gctgctcttt gagcctgcag
     1321 acacctgggg ggatacgggg aaaaggcctc caaggcctac tagtcccttt cgtcttcact
     1381 cgagtttact ccctatcagt gatagagaac gtatgaagag tttactccct atcagtgata
     1441 gagaacgtat gcagacttta ctccctatca gtgatagaga acgtataagg agtttactcc
     1501 ctatcagtga tagagaacgt atgaccagtt tactccctat cagtgataga gaacgtatct
     1561 acagtttact ccctatcagt gatagagaac gtatatccag tttactccct atcagtgata
     1621 gagaacgtat aagctatagg cgtgtacggt gggcgcctat aaaagcagag ctcgtttagt
     1681 gaaccgtcag atcgcctgga gcaattccac aacacttttg tcttatacca actttccgta
     1741 ccacttccta ccctcgtaaa ggaagctttc tagagctagc gtcgacacgc gtgcggccgc
     1801 tgtacaatgg gtaccgccac taacttctcc ctgttgaaac aagcagggga tgtcgaagag
     1861 aatcccgggc caatggtgag caagggcgag gagctgttca ccggggtggt gcccatcctg
     1921 gtcgagctgg acggcgacgt aaacggccac aagttcagcg tgtccggcga gggcgagggc
     1981 gatgccacct acggcaagct gaccctgaag ttcatctgca ccaccggcaa gctgcccgtg
     2041 ccctggccca ccctcgtgac caccctgacc tacggcgtgc agtgcttcag ccgctacccc
     2101 gaccacatga agcagcacga cttcttcaag tccgccatgc ccgaaggcta cgtccaggag
     2161 cgcaccatct tcttcaagga cgacggcaac tacaagaccc gcgccgaggt gaagttcgag
     2221 ggcgacaccc tggtgaaccg catcgagctg aagggcatcg acttcaagga ggacggcaac
     2281 atcctggggc acaagctgga gtacaactac aacagccaca acgtctatat catggccgac
     2341 aagcagaaga acggcatcaa ggtgaacttc aagatccgcc acaacatcga ggacggcagc
     2401 gtgcagctcg ccgaccacta ccagcagaac acccccatcg gcgacggccc cgtgctgctg
     2461 cccgacaacc actacctgag cacccagtcc gccctgagca aacctcccaa cgagaagcgc
     2521 gatcacatgg tcctgctgga gttcgtgacc gccgccggga tcactctcgg catggacgag
     2581 ctgtacaagt aaaccggtta gtaatgagtt taaacggggg aggctaactg aaacacggaa
     2641 ggagacaata ccggaaggaa cccgcgctat gacggcaata aaaagacaga ataaaacgca
     2701 cgggtgttgg gtcgtttgtt cataaacgcg gggttcggtc ccagggctgg cactctgtcg
     2761 ataccccacc gagaccccat tggggccaat acgcccgcgt ttcttccttt tccccacccc
     2821 accccccaag ttcgggtgaa ggcccagggc tcgcagccaa cgtcggggcg gcaggccctg
     2881 ccatagcaga tctgcgaaat cggatccagc tgaatctaag tggatctgcg atcgctccgg
     2941 tgcccgtcag tgggcagagc gcacatcgcc cacagtcccc gagaagttgg ggggaggggt
     3001 cggcaattga acgggtgcct agagaaggtg gcgcggggta aactgggaaa gtgatgtcgt
     3061 gtactggctc cgcctttttc ccgagggtgg gggagaaccg tatataagtg cagtagtcgc
     3121 cgtgaacgtt ctttttcgca acgggtttgc cgccagaaca cagctgaagc atcgaggggc
     3181 tcgcatctct ccttcacgcg cccgccgccc tacctgaggc cgccatccac gccggttgag
     3241 tcgcgttctg ccgcctcccg cctgtggtgc ctcctgaact gcgtccgccg tctaggtaag
     3301 tttaaagctc aggtcgagac cgggcctttg tccggcgctc ccttggagcc tacctagact
     3361 cagccggctc tccacgcttt gcctgaccct gcttgctcaa ctctacgtct ttgtttcgtt
     3421 ttctgttctg cgccgttaca gatccaagct gtgaccggcg cctactctag agccgccatg
     3481 tctagactgg acaagagcaa agtcataaac tctgctctgg aattactcaa tggagtcggt
     3541 atcgaaggcc tgacgacaag gaaactcgct caaaagctgg gagttgagca gcctaccctg
     3601 tactggcacg tgaagaacaa gcgggccctg ctcgatgccc tgccaatcga gatgctggac
     3661 aggcatcata cccactcctg ccccctggaa ggcgagtcat ggcaagactt tctgcggaac
     3721 aacgccaagt cataccgctg tgctctcctc tcacatcgcg acggggctaa agtgcatctc
     3781 ggcacccgcc caacagagaa acagtacgaa accctggaaa atcagctcgc gttcctgtgt
     3841 cagcaaggct tctccctgga gaacgcactg tacgctctgt ccgccgtggg ccactttaca
     3901 ctgggctgcg tattggagga acaggagcat caagtagcaa aagaggaaag agagacacct
     3961 accaccgatt ctatgccccc acttctgaaa caagcaattg agctgttcga ccggcaggga
     4021 gccgaacctg ccttcctttt cggcctggaa ctaatcatat gtggcctgga gaaacagcta
     4081 aagtgcgaaa gcggcgggcc gaccgacgcc cttgacgatt ttgacttaga catgctccca
     4141 gccgatgccc ttgacgactt tgaccttgat atgctgtctg ctgacgctct tgacgatttt
     4201 gaccttgaca tgctccccgg ggagggcaga ggaagtcttc taacatgcgg tgacgtggag
     4261 gagaatcccg gccctcttaa gatgaccgag tacaagccca cggtgcgcct cgccacccgc
     4321 gacgacgtcc ccagggccgt acgcaccctc gccgccgcgt tcgccgacta ccccgccacg
     4381 cgccacaccg tcgatccgga ccgccacatc gagcgggtca ccgagctgca agaactcttc
     4441 ctcacgcgcg tcgggctcga catcggcaag gtgtgggtcg cggacgacgg cgccgcggtg
     4501 gcggtctgga ccacgccgga gagcgtcgaa gcgggggcgg tgttcgccga gatcggcccg
     4561 cgcatggccg agttgagcgg ttcccggctg gccgcgcagc aacagatgga aggcctcctg
     4621 gcgccgcacc ggcccaagga gcccgcgtgg ttcctggcca ccgtcggcgt ctcgcccgac
     4681 caccagggca agggtctggg cagcgccgtc gtgctccccg gagtggaggc ggccgagcgc
     4741 gccggggtgc ccgccttcct ggagacctcc gcgccccgca acctcccctt ctacgagcgg
     4801 ctcggcttca ccgtcaccgc cgacgtcgag gtgcccgaag gaccgcgcac ctggtgcatg
     4861 acccgcaagc ccggtgcctg agaattctac ctacgctatg tggatacgct gctttaatgc
     4921 ctttgtatca gttaacttgt ttattgcagc ttataatggt tacaaataaa gcaatagcat
     4981 cacaaatttc acaaataaag catttttttc actgcattct agttgtggtt tgtccaaact
     5041 catcaatgta tcttaagctc atctggtctc ccttccgggg gacaagacat ccctgtttaa
     5101 tatttaaaca gcagtgttcc caaactgggt tcttatatcc cttgctctgg tcaaccaggt
     5161 tgcagggttt cctgtcctca caggaacgaa gtccctaaag aaacagtggc agccaggttt
     5221 agccccggaa ttgactggat tcctttttta gggcccattg gtatggcttt ttccccgtat
     5281 ccccccaggt gtctgcaggc tcaaagagca gcgagaagcg ttcagaggaa agcgatcccg
     5341 tgccaccttc cccgtgcccg ggctgtcccc gcacgctgcc ggctcgggga tgcgggggga
     5401 gcgccggacc ggagcggagc cccgggcggc tcgctgctgc cccctagcgg gggaggatct
     5461 ataacaagaa aatatatata taataagtta tcacgtaagt agaacatgaa ataacaatat
     5521 aattatcgta tgagttaaat cttaaaagtc acgtaaaaga taatcatgcg tcattttgac
     5581 tcacgcggtc gttatagttc aaaatcagtg acacttaccg cattgacaag cacgcctcac
     5641 gggagctcca agcggcgact gagatgtcct aaatgcacag cgacggattc gcgctattta
     5701 gaaagagaga gcaatatttc aagaatgcat gcgtcaattt tacgcagact atctttctag
     5761 ggttaatcta gctgcatcag gatcatatcg tcgggtcttt tttccggctc agtcatcgcc
     5821 caagctggcg ctatctgggc atcggggagg aagaagcccg tgccttttcc cgcgaggttg
     5881 aagcggcatg gaaagagttt gccgaggatg actgctgctg cattgacgtt gagcgaaaac
     5941 gcacgtttac catgatgatt cgggaaggtg tggccatgca cgcctttaac ggtgaactgt
     6001 tcgttcaggc cacctgggat accagttcgt cgcggctttt ccggacacag ttccggatgg
     6061 tcagcccgaa gcgcatcagc aacccgaaca ataccggcga cagccggaac tgccgtgccg
     6121 gtgtgcagat taatgacagc ggtgcggcgc tgggatatta cgtcagcgag gacgggtatc
     6181 ctggctggat gccgcagaaa tggacatgga taccccgtga gttacccggc gggcgcgctt
     6241 ggcgtaatca tggtcatagc tgtttcctgt gtgaaattgt tatccgctca caattccaca
     6301 caacatacga gccggaagca taaagtgtaa agcctggggt gcctaatgag tgagctaact
     6361 cacattaatt gcgttgcgct cactgcccgc tttccagtcg ggaaacctgt cgtgccagct
     6421 gcattaatga atcggccaac gcgcggggag aggcggtttg cgtattgggc gctcttccgc
     6481 ttcctcgctc actgactcgc tgcgctcggt cgttcggctg cggcgagcgg tatcagctca
     6541 ctcaaaggcg gtaatacggt tatccacaga atcaggggat aacgcaggaa agaacatgtg
     6601 agcaaaaggc cagcaaaagg ccaggaaccg taaaaaggcc gcgttgctgg cgtttttcca
     6661 taggctccgc ccccctgacg agcatcacaa aaatcgacgc tcaagtcaga ggtggcgaaa
     6721 cccgacagga ctataaagat accaggcgtt tccccctgga agctccctcg tgcgctctcc
     6781 tgttccgacc ctgccgctta ccggatacct gtccgccttt ctcccttcgg gaagcgtggc
     6841 gctttctcat agctcacgct gtaggtatct cagttcggtg taggtcgttc gctccaagct
     6901 gggctgtgtg cacgaacccc ccgttcagcc cgaccgctgc gccttatccg gtaactatcg
     6961 tcttgagtcc aacccggtaa gacacgactt atcgccactg gcagcagcca ctggtaacag
     7021 gattagcaga gcgaggtatg taggcggtgc tacagagttc ttgaagtggt ggcctaacta
     7081 cggctacact agaaggacag tatttggtat ctgcgctctg ctgaagccag ttaccttcgg
     7141 aaaaagagtt ggtagctctt gatccggcaa acaaaccacc gctggtagcg gtggtttttt
     7201 tgtttgcaag cagcagatta cgcgcagaaa aaaaggatct caagaagatc ctttgatctt
     7261 ttctacgggg tctgacgctc agtggaacga aaactcacgt taagggattt tggtcatgag
     7321 attatcaaaa aggatcttca cctagatcct tttaaattaa aaatgaagtt ttaaatcaat
     7381 ctaaagtata tatgagtaaa cttggtctga cagttaccaa tgcttaatca gtgaggcacc
     7441 tatctcagcg atctgtctat ttcgttcatc catagttgcc tgactccccg tcgtgtagat
     7501 aactacgata cgggagggct taccatctgg ccccagtgct gcaatgatac cgcgagaccc
     7561 acgctcaccg gctccagatt tatcagcaat aaaccagcca gccggaaggg ccgagcgcag
     7621 aagtggtcct gcaactttat ccgcctccat ccagtctatt aattgttgcc gggaagctag
     7681 agtaagtagt tcgccagtta atagtttgcg caacgttgtt gccattgcta caggcatcgt
     7741 ggtgtcacgc tcgtcgtttg gtatggcttc attcagctcc ggttcccaac gatcaaggcg
     7801 agttacatga tcccccatgt tgtgcaaaaa agcggttagc tccttcggtc ctccgatcgt
     7861 tgtcagaagt aagttggccg cagtgttatc actcatggtt atggcagcac tgcataattc
     7921 tcttactgtc atgccatccg taagatgctt ttctgtgact ggtgagtact caaccaagtc
     7981 attctgagaa tagtgtatgc ggcgaccgag ttgctcttgc ccggcgtcaa tacgggataa
     8041 taccgcgcca catagcagaa ctttaaaagt gctcatcatt ggaaaacgtt cttcggggcg
     8101 aaaactctca aggatcttac cgctgttgag atccagttcg atgtaaccca ctcgtgcacc
     8161 caactgatct tcagcatctt ttactttcac cagcgtttct gggtgagcaa aaacaggaag
     8221 gcaaaatgcc gcaaaaaagg gaataagggc gacacggaaa tgttgaatac tcat
//
