LOCUS       62056_20440        5280 bp DNA     circular SYN 17-NOV-2020
DEFINITION  Cloning vector pT-PGK-mCherry, complete sequence.
ACCESSION   MT891341
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5280)
  AUTHORS   Frei T, Cella F, Tedeschi F, Gutierrez J, Stan GB, Khammash M, 
            Siciliano V.
  TITLE     Characterization and mitigation of gene expression burden in 
            mammalian cells
  JOURNAL   Nat Commun 11 (1), 4641 (2020)
  PUBMED    32934213
REFERENCE   2  (bases 1 to 5280)
  AUTHORS   Frei T, Cella F, Tedeschi F, Gutierrez J, Stan G-B., Khammash M, 
            Siciliano V.
  TITLE     Direct Submission
  JOURNAL   Submitted (15-AUG-2020) Synthetic and Systems Biology for 
            Biomedicine, Italian Institute of Technology-IIT, Largo Barsanti e 
            Martteucci 53, Naples, Campania 80125, Italy
REFERENCE   3  (bases 1 to 5280)
  AUTHORS   .
  TITLE     Direct Submission
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Nat 
            Commun"; date: "2020"; volume: "11"; issue: "1"; pages: "4641"
COMMENT     SGRef: number: 2; type: "Journal Article"; journalName: "Submitted 
            (15-AUG-2020) Synthetic and Systems Biology for Biomedicine, Italian
            Institute of Technology-IIT, Largo Barsanti e Martteucci 53, Naples,
            Campania 80125, Italy"
COMMENT     ##Assembly-Data-START##
            Sequencing Technology :: Sanger dideoxy sequencing 
            ##Assembly-Data-END##
FEATURES             Location/Qualifiers
     source          1..5280
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        349..848
                     /label=PGK promoter
                     /note="mouse phosphoglycerate kinase 1 promoter"
     CDS             1273..1980
                     /codon_start=1
                     /label=mCherry
                     /note="monomeric derivative of DsRed fluorescent protein
                     (Shaner et al., 2004)"
                     /translation="VVSKGEEDNMAIIKEFMRFKVHMEGSVNGHEFEIEGEGEGRPYEG
                     TQTAKLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNF
                     EDGGVVTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASSERMYPEDGALK
                     GEIKQRLKLKDGGHYDAEVKTTYKAKKPVQLPGAYNVNIKLDITSHNEDYTIVEQYERA
                     EGRHSTGGMDELYK"
     polyA_signal    complement(2005..2139)
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     promoter        complement(2466..2484)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(2494..2510)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     rep_origin      2652..3107
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow
                     indicates direction of (+) strand synthesis"
     promoter        3133..3237
                     /label=AmpR promoter
     CDS             3238..4095
                     /codon_start=1
                     /label=AmpR
                     /note="beta-lactamase"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     rep_origin      4269..4857
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     protein_bind    5145..5166
                     /label=CAP binding site
                     /note="CAP binding activates transcription in the presence
                     of cAMP."
     promoter        5181..5211
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5219..5235
                     /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     5243..5259
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     promoter        5280
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
ORIGIN
        1 attaaccctc actaaaggga acaaaagctg gagctcggat ctgctgggct tgctgaaggt
       61 agggggtcaa gaaccagagg tgtaaagtac ttgagtaatt ttacttgatt actgtactta
      121 agtattattt ttggggattt ttactttact tgagtacaat taaaaatcaa tacttttact
      181 tttacttaat tacatttttt tagaaaaaaa agtacttttt actccttaca attttattta
      241 cagtcaaaaa gtacttattt tttggagatc acttgggccc tctaggtcga cctcgagctg
      301 cagcccgggg gatccagcgg ccgcgctagc ggcgcgccga attctaccgg gtaggggagg
      361 cgcttttccc aaggcagtct ggagcatgcg ctttagcagc cccgctgggc acttggcgct
      421 acacaagtgg cctctggcct cgcacacatt ccacatccac cggtaggcgc caaccggctc
      481 cgttctttgg tggccccttc gcgccacctt ctactcctcc cctagtcagg aagttccccc
      541 ccgccccgca gctcgcgtcg tgcaggacgt gacaaatgga agtagcacgt ctcactagtc
      601 tcgtgcagat ggacagcacc gctgagcaat ggaagcgggt aggcctttgg ggcagcggcc
      661 aatagcagct ttgctccttc gctttctggg ctcagaggct gggaaggggt gggtccgggg
      721 gcgggctcag gggcgggctc aggggcgggg cgggcgcccg aaggtcctcc ggaggcccgg
      781 cattctgcac gcttcaaaag cgcacgtctg ccgcgctgtt ctcctcttcc tcatctccgg
      841 gcctttcgac caattcccaa tactatcgtc gagtgagccg ccaccatgcc agagccagcg
      901 aagtctgctc ccgccccgaa aaagggctcc aagaaggcgg tgactaaggc gcagaagaaa
      961 ggcggcaaga agcgcaagcg cagccgcaag gagagctatt ccatctatgt gtacaaggtt
     1021 ctgaagcagg tccaccctga caccggcatt tcgtccaagg ccatgggcat catgaattcg
     1081 tttgtgaacg acattttcga gcgcatcgca ggtgaggctt cccgcctggc gcattacaac
     1141 aagcgctcga ccatcacctc cagggagatc cagacggccg tgcgcctgct gctgcctggg
     1201 gagttggcca agcacgccgt gtccgagggt actaaggcca tcaccaagta caccagcgct
     1261 aaggatccac cggtcgtgag caagggcgag gaggataaca tggccatcat caaggagttc
     1321 atgcgcttca aggtgcacat ggagggctcc gtgaacggcc acgagttcga gatcgagggc
     1381 gagggcgagg gccgccccta cgagggcacc cagaccgcca agctgaaggt gaccaagggt
     1441 ggccccctgc ccttcgcctg ggacatcctg tcccctcagt tcatgtacgg ctccaaggcc
     1501 tacgtgaagc accccgccga catccccgac tacttgaagc tgtccttccc cgagggcttc
     1561 aagtgggagc gcgtgatgaa cttcgaggac ggcggcgtgg tgaccgtgac ccaggactcc
     1621 tccctgcagg acggcgagtt catctacaag gtgaagctgc gcggcaccaa cttcccctcc
     1681 gacggccccg taatgcagaa gaagaccatg ggctgggagg cctcctccga gcggatgtac
     1741 cccgaggacg gcgccctgaa gggcgagatc aagcagaggc tgaagctgaa ggacggcggc
     1801 cactacgacg ctgaggtcaa gaccacctac aaggccaaga agcccgtgca gctgcccggc
     1861 gcctacaacg tcaacatcaa gttggacatc acctcccaca acgaggacta caccatcgtg
     1921 gaacagtacg aacgcgccga gggccgccac tccaccggcg gcatggacga gctgtacaag
     1981 tagtctagtg gatccaccga tgatgatcca gacatgataa gatacattga tgagtttgga
     2041 caaaccacaa ctagaatgca gtgaaaaaaa tgctttattt gtgaaatttg tgatgctatt
     2101 gctttatttg taaccattat aagctgcaat aaacaagtta acaacaacaa ttgcattcat
     2161 tttatgtttc aggttcaggg ggaggtgtgg gaggtttttt aaagcaagta aaacctctac
     2221 aaatgtggta tggctgatta tgatcctcta gatcagatct ttcttgcttt tacttttact
     2281 tccttaatac tcaagtacaa ttttaatgga gtactttttt acttttactc aagtaagatt
     2341 ctagccagat acttttactt ttaattgagt aaaattttcc ctaagtactt gtactttcac
     2401 ttgagtaaaa tttttgagta ctttttacac ctctgtcaag aaccatatgc cggtacccaa
     2461 ttcgccctat agtgagtcgt attacgcgcg ctcactggcc gtcgttttac aacgtcgtga
     2521 ctgggaaaac cctggcgtta cccaacttaa tcgccttgca gcacatcccc ctttcgccag
     2581 ctggcgtaat agcgaagagg cccgcaccga tcgcccttcc caacagttgc gcagcctgaa
     2641 tggcgaatgg gacgcgccct gtagcggcgc attaagcgcg gcgggtgtgg tggttacgcg
     2701 cagcgtgacc gctacacttg ccagcgccct agcgcccgct cctttcgctt tcttcccttc
     2761 ctttctcgcc acgttcgccg gctttccccg tcaagctcta aatcgggggc tccctttagg
     2821 gttccgattt agtgctttac ggcacctcga ccccaaaaaa cttgattagg gtgatggttc
     2881 acgtagtggg ccatcgccct gatagacggt ttttcgccct ttgacgttgg agtccacgtt
     2941 ctttaatagt ggactcttgt tccaaactgg aacaacactc aaccctatct cggtctattc
     3001 ttttgattta taagggattt tgccgatttc ggcctattgg ttaaaaaatg agctgattta
     3061 acaaaaattt aacgcgaatt ttaacaaaat attaacgctt acaatttagg tggcactttt
     3121 cggggaaatg tgcgcggaac ccctatttgt ttatttttct aaatacattc aaatatgtat
     3181 ccgctcatga gacaataacc ctgataaatg cttcaataat attgaaaaag gaagagtatg
     3241 agtattcaac atttccgtgt cgcccttatt cccttttttg cggcattttg ccttcctgtt
     3301 tttgctcacc cagaaacgct ggtgaaagta aaagatgctg aagatcagtt gggtgcacga
     3361 gtgggttaca tcgaactgga tctcaacagc ggtaagatcc ttgagagttt tcgccccgaa
     3421 gaacgttttc caatgatgag cacttttaaa gttctgctat gtggcgcggt attatcccgt
     3481 attgacgccg ggcaagagca actcggtcgc cgcatacact attctcagaa tgacttggtt
     3541 gagtactcac cagtcacaga aaagcatctt acggatggca tgacagtaag agaattatgc
     3601 agtgctgcca taaccatgag tgataacact gcggccaact tacttctgac aacgatcgga
     3661 ggaccgaagg agctaaccgc ttttttgcac aacatggggg atcatgtaac tcgccttgat
     3721 cgttgggaac cggagctgaa tgaagccata ccaaacgacg agcgtgacac cacgatgcct
     3781 gtagcaatgg caacaacgtt gcgcaaacta ttaactggcg aactacttac tctagcttcc
     3841 cggcaacaat taatagactg gatggaggcg gataaagttg caggaccact tctgcgctcg
     3901 gcccttccgg ctggctggtt tattgctgat aaatctggag ccggtgagcg tgggtctcgc
     3961 ggtatcattg cagcactggg gccagatggt aagccctccc gtatcgtagt tatctacacg
     4021 acggggagtc aggcaactat ggatgaacga aatagacaga tcgctgagat aggtgcctca
     4081 ctgattaagc attggtaact gtcagaccaa gtttactcat atatacttta gattgattta
     4141 aaacttcatt tttaatttaa aaggatctag gtgaagatcc tttttgataa tctcatgacc
     4201 aaaatccctt aacgtgagtt ttcgttccac tgagcgtcag accccgtaga aaagatcaaa
     4261 ggatcttctt gagatccttt ttttctgcgc gtaatctgct gcttgcaaac aaaaaaacca
     4321 ccgctaccag cggtggtttg tttgccggat caagagctac caactctttt tccgaaggta
     4381 actggcttca gcagagcgca gataccaaat actgtccttc tagtgtagcc gtagttaggc
     4441 caccacttca agaactctgt agcaccgcct acatacctcg ctctgctaat cctgttacca
     4501 gtggctgctg ccagtggcga taagtcgtgt cttaccgggt tggactcaag acgatagtta
     4561 ccggataagg cgcagcggtc gggctgaacg gggggttcgt gcacacagcc cagcttggag
     4621 cgaacgacct acaccgaact gagataccta cagcgtgagc tatgagaaag cgccacgctt
     4681 cccgaaggga gaaaggcgga caggtatccg gtaagcggca gggtcggaac aggagagcgc
     4741 acgagggagc ttccaggggg aaacgcctgg tatctttata gtcctgtcgg gtttcgccac
     4801 ctctgacttg agcgtcgatt tttgtgatgc tcgtcagggg ggcggagcct atggaaaaac
     4861 gccagcaacg cggccttttt acggttcctg gccttttgct ggccttttgc tcacatgttc
     4921 tttcctgcgt tatcccctga ttctgtggat aaccgtatta ccgcctttga gtgagctgat
     4981 accgctcgcc gcagccgaac gaccgagcgc agcgagtcag tgagcgagga agcggaagag
     5041 cgcccaatac gcaaaccgcc tctccccgcg cgttggccga ttcattaatg cagctggcac
     5101 gacaggtttc ccgactggaa agcgggcagt gagcgcaacg caattaatgt gagttagctc
     5161 actcattagg caccccaggc tttacacttt atgcttccgg ctcgtatgtt gtgtggaatt
     5221 gtgagcggat aacaatttca cacaggaaac agctatgacc atgattacgc caagcgcgca
//
