LOCUS       sequence-200607-        6523 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 6523)
  AUTHORS   Alford SC, Ding Y, Simmen T, Campbell RE
  TITLE     Dimerization-dependent green and yellow fluorescent proteins.
  JOURNAL   ACS Synth Biol. 2012 Dec 21;1(12):569-75. doi: 10.1021/sb300050j. 
            Epub 2012 Aug 14.
  PUBMED    23656278
REFERENCE   2  (bases 1 to 6523)
  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"; doi: "10.1021/sb300050j";
            journalName: "ACS Synth Biol"; date: "2012-12-21- 21"; volume: "1"; 
            issue: "12"; pages: "569-75"
COMMENT     Sequence Label: pTOM20-ddGFP-B
FEATURES             Location/Qualifiers
     source          1..6523
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        167..546
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        547..750
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     701..721
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     795..814
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        795..813
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     regulatory      838..847
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     primer_bind     complement(1460..1478)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     1728..1747
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     primer_bind     1922..1945
                     /label=DsRed1-C
                     /note="DsRed1, forward primer"
     primer_bind     complement(2046..2063)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    2052..2276
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      2322..2750
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2409..2428)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2619..2640
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(2759..2779)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        2764..3093
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      2944..3079
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     3006..3025
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             3160..3954
                     /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
                     GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(3214..3233)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     3824..3843
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    4128..4249
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4165..4184)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4219..4238
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(4298..4314)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4298..4314)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4311..4333)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4322..4338
                     /label=lac operator
                     /bound_moiety="lac repressor encoded by 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)."
     promoter        complement(join(4346..4352,4353..4370,4371..4376))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4391..4412
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4529..4546)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4700..5288)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4780..4799)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5459..6250,6251..6319))
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     6082..6101
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6320..6424)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(6499..6518)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
ORIGIN
        1 ttaagccagt atctgctccc tgcttgtgtg ttggaggtcg ctgagtagtg cgcgagcaaa
       61 atttaagcta caacaaggca aggcttgacc gacaattgca tgaagaatct gcttagggtt
      121 aggcgttttg cgctgcttcg cgatgtacgg gccagatata cgcgttgaca ttgattattg
      181 actagttatt aatagtaatc aattacgggg tcattagttc atagcccata tatggagttc
      241 cgcgttacat aacttacggt aaatggcccg cctggctgac cgcccaacga cccccgccca
      301 ttgacgtcaa taatgacgta tgttcccata gtaacgccaa tagggacttt ccattgacgt
      361 caatgggtgg agtatttacg gtaaactgcc cacttggcag tacatcaagt gtatcatatg
      421 ccaagtacgc cccctattga cgtcaatgac ggtaaatggc ccgcctggca ttatgcccag
      481 tacatgacct tatgggactt tcctacttgg cagtacatct acgtattagt catcgctatt
      541 accatggtga tgcggttttg gcagtacatc aatgggcgtg gatagcggtt tgactcacgg
      601 ggatttccaa gtctccaccc cattgacgtc aatgggagtt tgttttggca ccaaaatcaa
      661 cgggactttc caaaatgtcg taacaactcc gccccattga cgcaaatggg cggtaggcgt
      721 gtacggtggg aggtctatat aagcagagct ctctggctaa ctagagaacc cactgcttac
      781 tggcttatcg aaattaatac gactcactat agggagaccc aagctggcta gcgtgccgcc
      841 accatggcct cgagaatggt gggtcggaac agcgccatcg ccgccggtgt atgcggggcc
      901 cttttcattg ggtactgcat ctacttcgac cgcaaaagac gaagtgaccc caacttcaag
      961 aacaggcttc gagaacgaag aaagaaacag aagcttgcca aggagagagc tgggctttcc
     1021 aagttacctg accttaaaga tgctgaagct gttcagaagt tcttccttga agaaatacag
     1081 cttggtgaag agttactagc tcaaggtgaa tatgagaagg gcgtagacca tctgacaaat
     1141 gcaattgctg tgtgtggaca gccacagcag ttactgcagg tcttacagca aactcttcca
     1201 ccaccagtgt tccagatgct tctgactaag ctcccaacaa ttagtcagag aattgtaagt
     1261 gctcagagct tggctgaaga tgatgtggaa ggcggtagcg gggatccacc ggtcggtacc
     1321 gcctcctccg aggacaacaa catggccacc atcaaagagt tcatgcgctt caaggtgcgc
     1381 atggagggct ccatgaacgg ccacgagttc gagatcgagg gcgagggcga gggccgcccc
     1441 tacgagggca cccagaccgc caagctgaag gtgaccaagg gcggccccct gccctttgcc
     1501 tgggacatcc tgtcccccca gatcatgtac ggctccaaag cgtacgtgag gcaccccgcc
     1561 gacatccccg attacaagaa gctgcccttc cccgagggct tcaagtggga gcgcgtgatg
     1621 aacttcgaag acggcggtct ggtgaccgtt acccaggact cctccctgca ggacggcacg
     1681 ctgatctgca aggtgaagat tcgcggcacc aacttccccc ccgacggccc cgtaatgcag
     1741 aagaagacca tgggctggga ggcctccacc gagatgctgt accccaagga cggcgtgctg
     1801 aagggccata gctatcaggc cctgaagctg aaggacggcg gccactacct ggtggagttc
     1861 gagaccatct acatggccaa gaagcccgtg caactgcccg gcgattactg tgtggacacc
     1921 aagctggaca tcacctccca caacgaggac tacaccatcg tggaacagta cgggcgctcc
     1981 gagggccgcc accgtctggg catggacgag cggtacaagt aaaagcttgg taaaccgctg
     2041 atcagcctcg actgtgcctt ctagttgcca gccatctgtt gtttgcccct cccccgtgcc
     2101 ttccttgacc ctggaaggtg ccactcccac tgtcctttcc taataaaatg aggaaattgc
     2161 atcgcattgt ctgagtaggt gtcattctat tctggggggt ggggtggggc aggacagcaa
     2221 gggggaggat tgggaagaca atagcaggca tgctggggat gcggtgggct ctatggcttc
     2281 tgaggcggaa agaaccagct ggggctctag ggggtatccc cacgcgccct gtagcggcgc
     2341 attaagcgcg gcgggtgtgg tggttacgcg cagcgtgacc gctacacttg ccagcgccct
     2401 agcgcccgct cctttcgctt tcttcccttc ctttctcgcc acgttcgccg gctttccccg
     2461 tcaagctcta aatcgggggc tccctttagg gttccgattt agtgctttac ggcacctcga
     2521 ccccaaaaaa cttgattagg gtgatggttc acgtagtggg ccatcgccct gatagacggt
     2581 ttttcgccct ttgacgttgg agtccacgtt ctttaatagt ggactcttgt tccaaactgg
     2641 aacaacactc aaccctatct cggtctattc ttttgattta taagggattt tgccgatttc
     2701 ggcctattgg ttaaaaaatg agctgattta acaaaaattt aacgcgaatt aattctgtgg
     2761 aatgtgtgtc agttagggtg tggaaagtcc ccaggctccc cagcaggcag aagtatgcaa
     2821 agcatgcatc tcaattagtc agcaaccagg tgtggaaagt ccccaggctc cccagcaggc
     2881 agaagtatgc aaagcatgca tctcaattag tcagcaacca tagtcccgcc cctaactccg
     2941 cccatcccgc ccctaactcc gcccagttcc gcccattctc cgccccatgg ctgactaatt
     3001 ttttttattt atgcagaggc cgaggccgcc tctgcctctg agctattcca gaagtagtga
     3061 ggaggctttt ttggaggcct aggcttttgc aaaaagctcc cgggagcttg tatatccatt
     3121 ttcggatctg atcaagagac aggatgagga tcgtttcgca tgattgaaca agatggattg
     3181 cacgcaggtt ctccggccgc ttgggtggag aggctattcg gctatgactg ggcacaacag
     3241 acaatcggct gctctgatgc cgccgtgttc cggctgtcag cgcaggggcg cccggttctt
     3301 tttgtcaaga ccgacctgtc cggtgccctg aatgaactgc aggacgaggc agcgcggcta
     3361 tcgtggctgg ccacgacggg cgttccttgc gcagctgtgc tcgacgttgt cactgaagcg
     3421 ggaagggact ggctgctatt gggcgaagtg ccggggcagg atctcctgtc atctcacctt
     3481 gctcctgccg agaaagtatc catcatggct gatgcaatgc ggcggctgca tacgcttgat
     3541 ccggctacct gcccattcga ccaccaagcg aaacatcgca tcgagcgagc acgtactcgg
     3601 atggaagccg gtcttgtcga tcaggatgat ctggacgaag agcatcaggg gctcgcgcca
     3661 gccgaactgt tcgccaggct caaggcgcgc atgcccgacg gcgaggatct cgtcgtgacc
     3721 catggcgatg cctgcttgcc gaatatcatg gtggaaaatg gccgcttttc tggattcatc
     3781 gactgtggcc ggctgggtgt ggcggaccgc tatcaggaca tagcgttggc tacccgtgat
     3841 attgctgaag agcttggcgg cgaatgggct gaccgcttcc tcgtgcttta cggtatcgcc
     3901 gctcccgatt cgcagcgcat cgccttctat cgccttcttg acgagttctt ctgagcggga
     3961 ctctggggtt cgaaatgacc gaccaagcga cgcccaacct gccatcacga gatttcgatt
     4021 ccaccgccgc cttctatgaa aggttgggct tcggaatcgt tttccgggac gccggctgga
     4081 tgatcctcca gcgcggggat ctcatgctgg agttcttcgc ccaccccaac ttgtttattg
     4141 cagcttataa tggttacaaa taaagcaata gcatcacaaa tttcacaaat aaagcatttt
     4201 tttcactgca ttctagttgt ggtttgtcca aactcatcaa tgtatcttat catgtctgta
     4261 taccgtcgac ctctagctag agcttggcgt aatcatggtc atagctgttt cctgtgtgaa
     4321 attgttatcc gctcacaatt ccacacaaca tacgagccgg aagcataaag tgtaaagcct
     4381 ggggtgccta atgagtgagc taactcacat taattgcgtt gcgctcactg cccgctttcc
     4441 agtcgggaaa cctgtcgtgc cagctgcatt aatgaatcgg ccaacgcgcg gggagaggcg
     4501 gtttgcgtat tgggcgctct tccgcttcct cgctcactga ctcgctgcgc tcggtcgttc
     4561 ggctgcggcg agcggtatca gctcactcaa aggcggtaat acggttatcc acagaatcag
     4621 gggataacgc aggaaagaac atgtgagcaa aaggccagca aaaggccagg aaccgtaaaa
     4681 aggccgcgtt gctggcgttt ttccataggc tccgcccccc tgacgagcat cacaaaaatc
     4741 gacgctcaag tcagaggtgg cgaaacccga caggactata aagataccag gcgtttcccc
     4801 ctggaagctc cctcgtgcgc tctcctgttc cgaccctgcc gcttaccgga tacctgtccg
     4861 cctttctccc ttcgggaagc gtggcgcttt ctcatagctc acgctatagg tatctcagtt
     4921 cggtgtaggt cgttcgctcc aagctgggct gtgtgcacga accccccgtt cagcccgacc
     4981 gctgcgcctt atccggtaac tatcgtcttg agtccaaccc ggtaagacac gacttatcgc
     5041 cactggcagc agccactggt aacaggatta gcagagcgag gtatgtaggc ggtgctacag
     5101 agttcttgaa gtggtggcct aactacggct acactagaag aacagtattt ggtatctgcg
     5161 ctctgctgaa gccagttacc ttcggaaaaa gagttggtag ctcttgatcc ggcaaacaaa
     5221 ccaccgctgg tagcggtggt ttttttgttt gcaagcagca gattacgcgc agaaaaaaag
     5281 gatctcaaga agatcctttg atcttttcta cggggtctga cgctcagtgg aacgaaaact
     5341 cacgttaagg gattttggtc atgagattat caaaaaggat cttcacctag atccttttaa
     5401 attaaaaatg aagttttaaa tcaatctaaa gtatatatga gtaaacttgg tctgacagtt
     5461 accaatgctt aatcagtgag gcacctatct cagcgatctg tctatttcgt tcatccatag
     5521 ttgcctgact ccccgtcgtg tagataacta cgatacggga gggcttacca tctggcccca
     5581 gtgctgcaat gataccgcga gacccacgct caccggctcc agatttatca gcaataaacc
     5641 agccagccgg aagggccgag cgcagaagtg gtcctgcaac tttatccgcc tccatccagt
     5701 ctattaattg ttgccgggaa gctagagtaa gtagttcgcc agttaatagt ttgcgcaacg
     5761 ttgttgccat tgctacaggc atcgtggtgt cacgctcgtc gtttggtatg gcttcattca
     5821 gctccggttc ccaacgatca aggcgagtta catgatcccc catgttgtgc aaaaaagcgg
     5881 ttagctcctt cggtcctccg atcgttgtca gaagtaagtt ggccgcagtg ttatcactca
     5941 tggttatggc agcactgcat aattctctta ctgtcatgcc atccgtaaga tgcttttctg
     6001 tgactggtga gtactcaacc aagtcattct gagaatagtg tatgcggcga ccgagttgct
     6061 cttgcccggc gtcaatacgg gataataccg cgccacatag cagaacttta aaagtgctca
     6121 tcattggaaa acgttcttcg gggcgaaaac tctcaaggat cttaccgctg ttgagatcca
     6181 gttcgatgta acccactcgt gcacccaact gatcttcagc atcttttact ttcaccagcg
     6241 tttctgggtg agcaaaaaca ggaaggcaaa atgccgcaaa aaagggaata agggcgacac
     6301 ggaaatgttg aatactcata ctcttccttt ttcaatatta ttgaagcatt tatcagggtt
     6361 attgtctcat gagcggatac atatttgaat gtatttagaa aaataaacaa ataggggttc
     6421 cgcgcacatt tccccgaaaa gtgccacctg acgtcgacgg atcgggagat ctcccgatcc
     6481 cctatggtgc actctcagta caatctgctc tgatgccgca tag
//
