LOCUS       sequence-367860-        6737 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  synthetic circular DNA
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6737)
  AUTHORS   Wu SM, Fujiwara Y, Cibulsky SM, Clapham DE, Lien CL, Schultheiss TM,
            Orkin SH
  TITLE     Developmental origin of a bipotential myocardial and smooth muscle 
            cell precursor in the mammalian heart.
  JOURNAL   Cell. 2006 Dec 15;127(6):1137-50. Epub 2006 Nov 22.
  PUBMED    17123591
REFERENCE   2  (bases 1 to 6737)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Dec 15, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Cell."; 
            date: "2006-12-15"; volume: "127(6)"; pages: "1137-50. Epub 2006 Nov
            22"
COMMENT     Sequence Label: pNkx-GFP
FEATURES             Location/Qualifiers
     source          1..6737
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     regulatory      2791..2800
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(2797..2799,2800..2802,2803..3516)
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             join(2797..2799,2800..2802,2803..3513)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(2842..2863)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(3103..3122)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     3450..3471
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     polyA_signal    3639..3760
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3676..3695)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3730..3749
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(3767..4222)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3904..3925)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     4116..4135
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        4249..4353
                     /gene="bla"
                     /label=AmpR promoter
     promoter        4355..4712
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        4355..4546
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(4378..4398)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      4563..4698
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             4747..5541
                     /codon_start=1
                     /product="aminoglycoside phosphotransferase"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     CDS             4747..5541
                     /codon_start=1
                     /gene="aph(3')-II (or nptII)"
                     /product="aminoglycoside phosphotransferase from Tn5"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(4801..4820)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     5411..5430
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(5729..5748)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    5773..5820
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      6149..6737
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6638..6657
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 aacgccagca acgcggcctt tttacggttc ctggcctttt gctggccttt tgctcacatg
       61 ttctttcctg cgttatcccc tgattctgtg gataaccgta ttaccgccat gcattagtta
      121 ttactagcgc taccggactc agatctcgag ctccaccgcg gtggcggccg ctctagaact
      181 agtccagtct gggtcctaat gcgggtggcg tctctcttga caggcagcgt ttggggacaa
      241 cagcggggaa gggagataag atgacatacc agagcagatt tggtgtgcgc gctgatactc
      301 ctggcccgac aggaaactcg gagctattta aaaaggccct atcgattact ttatcttccc
      361 cggaggaaaa cttcttgccg agagacaaaa gatgtccccc tacctaaaga tacaaggcca
      421 cacagcagag ggtttgtaca ggcgacgcgc gagtagatcc ctgggagcgc agagacgcct
      481 ttctaccggc agagactgaa gtttgactgg agcgaggcgg ggcggagcca cctcccggtc
      541 cccgtgcact ccggaattgt gaacgctccc gcaaagtccc cgcgagtgtt gtgttcacac
      601 taattggtgt gcctctgagt tccatccgta ctcccccccc cccaagttta aatgctcctt
      661 ttaagggctt gagtgtctgc agccgtcatg tgcaccttga aagcttggcg tgtgacattg
      721 tgttcctcta gcgtctccat cccgcttttc accaattcgt gccctggccg aatcccgtgg
      781 gagcagaggt ggcccctcga agtcccgggc cacccaggga ggctccgggt acacccagat
      841 tcccagcagt ccgtagtctg ggctttcccc gtgtgacact ttgccgccta ccggttcaca
      901 atcctctcgg cccgcgtgag agaaggcggc atgttccgtg ggacacagtt tattttatgc
      961 acattaaggg ctgtaaagca cggtctgaaa gaaaaagatg gtgcggcggg gaggagcggg
     1021 agaaggggca gcgatgatcc tgggactgtg cagcgaccac ttacaacctg attcatgccc
     1081 accggggacc ccagcggtcc tggagggagg cggaggctcc gaggcttccg gttgggtttc
     1141 agccaccagg ttcgggcagt gactcttgga ctctgaccac gtcccgcggc acactcggag
     1201 acccggctcg tccgcgaagc taggagcgaa tggcccgcag ctgcgcgtcg ctcaatctga
     1261 gctctcaagc ccaaagcgtc tgcagagccg cttcccagag ctccaagcca ccgctagcag
     1321 agctctgaag gcaccttgaa gcctcgaccc acttcaatgc cacctgattt ttcaaattat
     1381 caccttctga gacgcgatcc ttcggaaccg gcagagcccc cacctttgag cgccgccgtt
     1441 tatgctgtag cgctgtagcg gtttgcagca gtcagggcag gtggaggacc ttgttactgg
     1501 cactgcggag cccggagctg aggggaaaga aaacggagat aaggaaaagg gctgcagagg
     1561 tcggaaagca gctcccatcg gatccatcaa atcaatcaca gccccaagtg accaaagaga
     1621 aattttgcgt tttggaaata tccccaccac caaaagcacg aggaaagtgt gagcagcaag
     1681 aaactaacaa taaattttag gtgcaaaaat ataaagaagc acacagaatt tgaaaaaaaa
     1741 aacccgcaca ttctgctatt tgagttttcc ataaacatac aaataaaacc tgcggaagat
     1801 gtacttcttc aagtctgtga cagaccctgg actgactaac ggggacgcgc tcatttaaaa
     1861 tgtcagcggt tcctatattt caccgaactt tcaacgagct tttccacatg tccagactca
     1921 tgggacagta tgcttatgac cctgttaatt gtagttcaga cttactatgt ctcaataaag
     1981 tgactcagca cttgaccagc atatgaggct ccccgagtcc agtccccaga aacaagtaga
     2041 atatacatat atatataatt ctatatatct atctatatat tattctatat atatattcta
     2101 cttgtatata tatgtattac ataatatata tacaagtaga atatatataa tataatttat
     2161 attatatata gaatatatat agaaacaagt agattatata tatatatata tagtatatat
     2221 atatatatat atatatatat atagtattgt attacataca tgtgggtata catcatacac
     2281 aggactagtg gatcccccag cttggggctt tcaatccatc ctcaccagcc catttagtga
     2341 tacaggacac agtgcaaaga atatttattc aaatcctcga cccaccccca accctgcgtt
     2401 tagactcagc ataacagaat caggtatttt ctttgagtgt gtctggctgg gattttcagg
     2461 ctaacgagga gcaatctcct tgcaggagtg ggagactatc ccaaatgtgt cgcggcccct
     2521 ttaaaaaaag ctgagcagga tttttttttc ccatccctat gtctatatta ggtgacgcag
     2581 aactgcccgt cgctcctgtc atccaggccc ctggcccaat ggcaggctga atccccccta
     2641 ctccagcctg ctcccgcctc tctgccctgg atgtggcgcg cggcgggcgg cgggcaccat
     2701 gcggggaggc tgtccccagg ggtgggcagc accactctct gctacccacc tggcccaagc
     2761 tgggtaccgc gggcccggga tccaccggtc gccaccatgg tgagcaaggg cgaggagctg
     2821 ttcaccgggg tggtgcccat cctggtcgag ctggacggcg acgtaaacgg ccacaagttc
     2881 agcgtgtccg gcgagggcga gggcgatgcc acctacggca agctgaccct gaagttcatc
     2941 tgcaccaccg gcaagctgcc cgtgccctgg cccaccctcg tgaccaccct gacctacggc
     3001 gtgcagtgct tcagccgcta ccccgaccac atgaagcagc acgacttctt caagtccgcc
     3061 atgcccgaag gctacgtcca ggagcgcacc atcttcttca aggacgacgg caactacaag
     3121 acccgcgccg aggtgaagtt cgagggcgac accctggtga accgcatcga gctgaagggc
     3181 atcgacttca aggaggacgg caacatcctg gggcacaagc tggagtacaa ctacaacagc
     3241 cacaacgtct atatcatggc cgacaagcag aagaacggca tcaaggtgaa cttcaagatc
     3301 cgccacaaca tcgaggacgg cagcgtgcag ctcgccgacc actaccagca gaacaccccc
     3361 atcggcgacg gccccgtgct gctgcccgac aaccactacc tgagcaccca gtccgccctg
     3421 agcaaagacc ccaacgagaa gcgcgatcac atggtcctgc tggagttcgt gaccgccgcc
     3481 gggatcactc tcggcatgga cgagctgtac aagtaaagcg gccgcgactc tagatcataa
     3541 tcagccatac cacatttgta gaggttttac ttgctttaaa aaacctccca cacctccccc
     3601 tgaacctgaa acataaaatg aatgcaattg ttgttgttaa cttgtttatt gcagcttata
     3661 atggttacaa ataaagcaat agcatcacaa atttcacaaa taaagcattt ttttcactgc
     3721 attctagttg tggtttgtcc aaactcatca atgtatctta aggcgtaaat tgtaagcgtt
     3781 aatattttgt taaaattcgc gttaaatttt tgttaaatca gctcattttt taaccaatag
     3841 gccgaaatcg gcaaaatccc ttataaatca aaagaataga ccgagatagg gttgagtgtt
     3901 gttccagttt ggaacaagag tccactatta aagaacgtgg actccaacgt caaagggcga
     3961 aaaaccgtct atcagggcga tggcccacta cgtgaaccat caccctaatc aagttttttg
     4021 gggtcgaggt gccgtaaagc actaaatcgg aaccctaaag ggagcccccg atttagagct
     4081 tgacggggaa agccggcgaa cgtggcgaga aaggaaggga agaaagcgaa aggagcgggc
     4141 gctagggcgc tggcaagtgt agcggtcacg ctgcgcgtaa ccaccacacc cgccgcgctt
     4201 aatgcgccgc tacagggcgc gtcaggtggc acttttcggg gaaatgtgcg cggaacccct
     4261 atttgtttat ttttctaaat acattcaaat atgtatccgc tcatgagaca ataaccctga
     4321 taaatgcttc aataatattg aaaaaggaag agtcctgagg cggaaagaac cagctgtgga
     4381 atgtgtgtca gttagggtgt ggaaagtccc caggctcccc agcaggcaga agtatgcaaa
     4441 gcatgcatct caattagtca gcaaccaggt gtggaaagtc cccaggctcc ccagcaggca
     4501 gaagtatgca aagcatgcat ctcaattagt cagcaaccat agtcccgccc ctaactccgc
     4561 ccatcccgcc cctaactccg cccagttccg cccattctcc gccccatggc tgactaattt
     4621 tttttattta tgcagaggcc gaggccgcct cggcctctga gctattccag aagtagtgag
     4681 gaggcttttt tggaggccta ggcttttgca aagatcgatc aagagacagg atgaggatcg
     4741 tttcgcatga ttgaacaaga tggattgcac gcaggttctc cggccgcttg ggtggagagg
     4801 ctattcggct atgactgggc acaacagaca atcggctgct ctgatgccgc cgtgttccgg
     4861 ctgtcagcgc aggggcgccc ggttcttttt gtcaagaccg acctgtccgg tgccctgaat
     4921 gaactgcaag acgaggcagc gcggctatcg tggctggcca cgacgggcgt tccttgcgca
     4981 gctgtgctcg acgttgtcac tgaagcggga agggactggc tgctattggg cgaagtgccg
     5041 gggcaggatc tcctgtcatc tcaccttgct cctgccgaga aagtatccat catggctgat
     5101 gcaatgcggc ggctgcatac gcttgatccg gctacctgcc cattcgacca ccaagcgaaa
     5161 catcgcatcg agcgagcacg tactcggatg gaagccggtc ttgtcgatca ggatgatctg
     5221 gacgaagagc atcaggggct cgcgccagcc gaactgttcg ccaggctcaa ggcgagcatg
     5281 cccgacggcg aggatctcgt cgtgacccat ggcgatgcct gcttgccgaa tatcatggtg
     5341 gaaaatggcc gcttttctgg attcatcgac tgtggccggc tgggtgtggc ggaccgctat
     5401 caggacatag cgttggctac ccgtgatatt gctgaagagc ttggcggcga atgggctgac
     5461 cgcttcctcg tgctttacgg tatcgccgct cccgattcgc agcgcatcgc cttctatcgc
     5521 cttcttgacg agttcttctg agcgggactc tggggttcga aatgaccgac caagcgacgc
     5581 ccaacctgcc atcacgagat ttcgattcca ccgccgcctt ctatgaaagg ttgggcttcg
     5641 gaatcgtttt ccgggacgcc ggctggatga tcctccagcg cggggatctc atgctggagt
     5701 tcttcgccca ccctaggggg aggctaactg aaacacggaa ggagacaata ccggaaggaa
     5761 cccgcgctat gacggcaata aaaagacaga ataaaacgca cggtgttggg tcgtttgttc
     5821 ataaacgcgg ggttcggtcc cagggctggc actctgtcga taccccaccg agaccccatt
     5881 ggggccaata cgcccgcgtt tcttcctttt ccccacccca ccccccaagt tcgggtgaag
     5941 gcccagggct cgcagccaac gtcggggcgg caggccctgc catagcctca ggttactcat
     6001 atatacttta gattgattta aaacttcatt tttaatttaa aaggatctag gtgaagatcc
     6061 tttttgataa tctcatgacc aaaatccctt aacgtgagtt ttcgttccac tgagcgtcag
     6121 accccgtaga aaagatcaaa ggatcttctt gagatccttt ttttctgcgc gtaatctgct
     6181 gcttgcaaac aaaaaaacca ccgctaccag cggtggtttg tttgccggat caagagctac
     6241 caactctttt tccgaaggta actggcttca gcagagcgca gataccaaat actgttcttc
     6301 tagtgtagcc gtagttaggc caccacttca agaactctgt agcaccgcct acatacctcg
     6361 ctctgctaat cctgttacca gtggctgctg ccagtggcga taagtcgtgt cttaccgggt
     6421 tggactcaag acgatagtta ccggataagg cgcagcggtc gggctgaacg gggggttcgt
     6481 gcacacagcc cagcttggag cgaacgacct acaccgaact gagataccta cagcgtgagc
     6541 tatgagaaag cgccacgctt cccgaaggga gaaaggcgga caggtatccg gtaagcggca
     6601 gggtcggaac aggagagcgc acgagggagc ttccaggggg aaacgcctgg tatctttata
     6661 gtcctgtcgg gtttcgccac ctctgacttg agcgtcgatt tttgtgatgc tcgtcagggg
     6721 ggcggagcct atggaaa
//
