LOCUS       sequence-234036-        6181 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 6181)
  AUTHORS   Galietta LJ, Haggie PM, Verkman AS
  TITLE     Green fluorescent protein-based halide indicators with improved 
            chloride and iodide affinities.
  JOURNAL   FEBS Lett. 2001 Jun 22. 499(3):220-4.
  PUBMED    11423120
REFERENCE   2  (bases 1 to 6181)
  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: "FEBS Lett. 
            2001 Jun 22. 499(3):220-4."
COMMENT     Sequence Label: pcDNA3.1 EYFP H148Q/I152L
FEATURES             Location/Qualifiers
     source          1..6181
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        1..380
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        381..584
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     535..555
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     629..648
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        629..647
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     regulatory      726..735
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(732..734,735..737,738..1451)
                     /codon_start=1
                     /product="enhanced YFP"
                     /label=EYFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTFGYGLQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSQNVYLMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSYQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(777..798)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(1038..1057)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1385..1406
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     complement(1538..1555)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    1544..1768
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      1814..2242
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1901..1920)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2111..2132
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(2251..2271)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        2256..2585
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      2436..2571
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     2498..2517
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             2652..3446
                     /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(2706..2725)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     3316..3335
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    3620..3741
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3657..3676)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3711..3730
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(3790..3806)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3790..3806)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3803..3825)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    3814..3830
                     /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(3838..3844,3845..3862,3863..3868))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3883..3904
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4021..4038)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4192..4780)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4272..4291)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4951..5742,5743..5811))
                     /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     5574..5593
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5812..5916)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(5991..6010)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
ORIGIN
        1 gacattgatt attgactagt tattaatagt aatcaattac ggggtcatta gttcatagcc
       61 catatatgga gttccgcgtt acataactta cggtaaatgg cccgcctggc tgaccgccca
      121 acgacccccg cccattgacg tcaataatga cgtatgttcc catagtaacg ccaataggga
      181 ctttccattg acgtcaatgg gtggagtatt tacggtaaac tgcccacttg gcagtacatc
      241 aagtgtatca tatgccaagt acgcccccta ttgacgtcaa tgacggtaaa tggcccgcct
      301 ggcattatgc ccagtacatg accttatggg actttcctac ttggcagtac atctacgtat
      361 tagtcatcgc tattaccatg gtgatgcggt tttggcagta catcaatggg cgtggatagc
      421 ggtttgactc acggggattt ccaagtctcc accccattga cgtcaatggg agtttgtttt
      481 ggcaccaaaa tcaacgggac tttccaaaat gtcgtaacaa ctccgcccca ttgacgcaaa
      541 tgggcggtag gcgtgtacgg tgggaggtct atataagcag agctctctgg ctaactagag
      601 aacccactgc ttactggctt atcgaaatta atacgactca ctatagggag acccaagctg
      661 gctagcgttt aaacttaagc ttgcatgcct gcaggtcgac tctagaggat ccccgggtac
      721 cggtcgccac catggtgagc aagggcgagg agctgttcac cggggtggtg cccatcctgg
      781 tcgagctgga cggcgacgta aacggccaca agttcagcgt gtccggcgag ggcgagggcg
      841 atgccaccta cggcaagctg accctgaagt tcatctgcac caccggcaag ctgcccgtgc
      901 cctggcccac cctcgtgacc accttcggct acggcctgca gtgcttcgcc cgctaccccg
      961 accacatgaa gcagcacgac ttcttcaagt ccgccatgcc cgaaggctac gtccaggagc
     1021 gcaccatctt cttcaaggac gacggcaact acaagacccg cgccgaggtg aagttcgagg
     1081 gcgacaccct ggtgaaccgc atcgagctga agggcatcga cttcaaggag gacggcaaca
     1141 tcctggggca caagctggag tacaactaca acagccagaa cgtctatctg atggccgaca
     1201 agcagaagaa cggcatcaag gtgaacttca agatccgcca caacatcgag gacggcagcg
     1261 tgcagctcgc cgaccactac cagcagaaca cccccatcgg cgacggcccc gtgctgctgc
     1321 ccgacaacca ctacctgagc taccagtccg ccctgagcaa agaccccaac gagaagcgcg
     1381 atcacatggt cctgctggag ttcgtgaccg ccgccgggat cactctcggc atggacgagc
     1441 tgtacaagta aagcggccgc gactctagaa ttctgcagat atccagcaca gtggcggccg
     1501 ctcgagtcta gagggcccgt ttaaacccgc tgatcagcct cgactgtgcc ttctagttgc
     1561 cagccatctg ttgtttgccc ctcccccgtg ccttccttga ccctggaagg tgccactccc
     1621 actgtccttt cctaataaaa tgaggaaatt gcatcgcatt gtctgagtag gtgtcattct
     1681 attctggggg gtggggtggg gcaggacagc aagggggagg attgggaaga caatagcagg
     1741 catgctgggg atgcggtggg ctctatggct tctgaggcgg aaagaaccag ctggggctct
     1801 agggggtatc cccacgcgcc ctgtagcggc gcattaagcg cggcgggtgt ggtggttacg
     1861 cgcagcgtga ccgctacact tgccagcgcc ctagcgcccg ctcctttcgc tttcttccct
     1921 tcctttctcg ccacgttcgc cggctttccc cgtcaagctc taaatcgggg gctcccttta
     1981 gggttccgat ttagtgcttt acggcacctc gaccccaaaa aacttgatta gggtgatggt
     2041 tcacgtagtg ggccatcgcc ctgatagacg gtttttcgcc ctttgacgtt ggagtccacg
     2101 ttctttaata gtggactctt gttccaaact ggaacaacac tcaaccctat ctcggtctat
     2161 tcttttgatt tataagggat tttgccgatt tcggcctatt ggttaaaaaa tgagctgatt
     2221 taacaaaaat ttaacgcgaa ttaattctgt ggaatgtgtg tcagttaggg tgtggaaagt
     2281 ccccaggctc cccagcaggc agaagtatgc aaagcatgca tctcaattag tcagcaacca
     2341 ggtgtggaaa gtccccaggc tccccagcag gcagaagtat gcaaagcatg catctcaatt
     2401 agtcagcaac catagtcccg cccctaactc cgcccatccc gcccctaact ccgcccagtt
     2461 ccgcccattc tccgccccat ggctgactaa ttttttttat ttatgcagag gccgaggccg
     2521 cctctgcctc tgagctattc cagaagtagt gaggaggctt ttttggaggc ctaggctttt
     2581 gcaaaaagct cccgggagct tgtatatcca ttttcggatc tgatcaagag acaggatgag
     2641 gatcgtttcg catgattgaa caagatggat tgcacgcagg ttctccggcc gcttgggtgg
     2701 agaggctatt cggctatgac tgggcacaac agacaatcgg ctgctctgat gccgccgtgt
     2761 tccggctgtc agcgcagggg cgcccggttc tttttgtcaa gaccgacctg tccggtgccc
     2821 tgaatgaact gcaggacgag gcagcgcggc tatcgtggct ggccacgacg ggcgttcctt
     2881 gcgcagctgt gctcgacgtt gtcactgaag cgggaaggga ctggctgcta ttgggcgaag
     2941 tgccggggca ggatctcctg tcatctcacc ttgctcctgc cgagaaagta tccatcatgg
     3001 ctgatgcaat gcggcggctg catacgcttg atccggctac ctgcccattc gaccaccaag
     3061 cgaaacatcg catcgagcga gcacgtactc ggatggaagc cggtcttgtc gatcaggatg
     3121 atctggacga agagcatcag gggctcgcgc cagccgaact gttcgccagg ctcaaggcgc
     3181 gcatgcccga cggcgaggat ctcgtcgtga cccatggcga tgcctgcttg ccgaatatca
     3241 tggtggaaaa tggccgcttt tctggattca tcgactgtgg ccggctgggt gtggcggacc
     3301 gctatcagga catagcgttg gctacccgtg atattgctga agagcttggc ggcgaatggg
     3361 ctgaccgctt cctcgtgctt tacggtatcg ccgctcccga ttcgcagcgc atcgccttct
     3421 atcgccttct tgacgagttc ttctgagcgg gactctgggg ttcgaaatga ccgaccaagc
     3481 gacgcccaac ctgccatcac gagatttcga ttccaccgcc gccttctatg aaaggttggg
     3541 cttcggaatc gttttccggg acgccggctg gatgatcctc cagcgcgggg atctcatgct
     3601 ggagttcttc gcccacccca acttgtttat tgcagcttat aatggttaca aataaagcaa
     3661 tagcatcaca aatttcacaa ataaagcatt tttttcactg cattctagtt gtggtttgtc
     3721 caaactcatc aatgtatctt atcatgtctg tataccgtcg acctctagct agagcttggc
     3781 gtaatcatgg tcatagctgt ttcctgtgtg aaattgttat ccgctcacaa ttccacacaa
     3841 catacgagcc ggaagcataa agtgtaaagc ctggggtgcc taatgagtga gctaactcac
     3901 attaattgcg ttgcgctcac tgcccgcttt ccagtcggga aacctgtcgt gccagctgca
     3961 ttaatgaatc ggccaacgcg cggggagagg cggtttgcgt attgggcgct cttccgcttc
     4021 ctcgctcact gactcgctgc gctcggtcgt tcggctgcgg cgagcggtat cagctcactc
     4081 aaaggcggta atacggttat ccacagaatc aggggataac gcaggaaaga acatgtgagc
     4141 aaaaggccag caaaaggcca ggaaccgtaa aaaggccgcg ttgctggcgt ttttccatag
     4201 gctccgcccc cctgacgagc atcacaaaaa tcgacgctca agtcagaggt ggcgaaaccc
     4261 gacaggacta taaagatacc aggcgtttcc ccctggaagc tccctcgtgc gctctcctgt
     4321 tccgaccctg ccgcttaccg gatacctgtc cgcctttctc ccttcgggaa gcgtggcgct
     4381 ttctcatagc tcacgctgta ggtatctcag ttcggtgtag gtcgttcgct ccaagctggg
     4441 ctgtgtgcac gaaccccccg ttcagcccga ccgctgcgcc ttatccggta actatcgtct
     4501 tgagtccaac ccggtaagac acgacttatc gccactggca gcagccactg gtaacaggat
     4561 tagcagagcg aggtatgtag gcggtgctac agagttcttg aagtggtggc ctaactacgg
     4621 ctacactaga agaacagtat ttggtatctg cgctctgctg aagccagtta ccttcggaaa
     4681 aagagttggt agctcttgat ccggcaaaca aaccaccgct ggtagcggtg gtttttttgt
     4741 ttgcaagcag cagattacgc gcagaaaaaa aggatctcaa gaagatcctt tgatcttttc
     4801 tacggggtct gacgctcagt ggaacgaaaa ctcacgttaa gggattttgg tcatgagatt
     4861 atcaaaaagg atcttcacct agatcctttt aaattaaaaa tgaagtttta aatcaatcta
     4921 aagtatatat gagtaaactt ggtctgacag ttaccaatgc ttaatcagtg aggcacctat
     4981 ctcagcgatc tgtctatttc gttcatccat agttgcctga ctccccgtcg tgtagataac
     5041 tacgatacgg gagggcttac catctggccc cagtgctgca atgataccgc gagacccacg
     5101 ctcaccggct ccagatttat cagcaataaa ccagccagcc ggaagggccg agcgcagaag
     5161 tggtcctgca actttatccg cctccatcca gtctattaat tgttgccggg aagctagagt
     5221 aagtagttcg ccagttaata gtttgcgcaa cgttgttgcc attgctacag gcatcgtggt
     5281 gtcacgctcg tcgtttggta tggcttcatt cagctccggt tcccaacgat caaggcgagt
     5341 tacatgatcc cccatgttgt gcaaaaaagc ggttagctcc ttcggtcctc cgatcgttgt
     5401 cagaagtaag ttggccgcag tgttatcact catggttatg gcagcactgc ataattctct
     5461 tactgtcatg ccatccgtaa gatgcttttc tgtgactggt gagtactcaa ccaagtcatt
     5521 ctgagaatag tgtatgcggc gaccgagttg ctcttgcccg gcgtcaatac gggataatac
     5581 cgcgccacat agcagaactt taaaagtgct catcattgga aaacgttctt cggggcgaaa
     5641 actctcaagg atcttaccgc tgttgagatc cagttcgatg taacccactc gtgcacccaa
     5701 ctgatcttca gcatctttta ctttcaccag cgtttctggg tgagcaaaaa caggaaggca
     5761 aaatgccgca aaaaagggaa taagggcgac acggaaatgt tgaatactca tactcttcct
     5821 ttttcaatat tattgaagca tttatcaggg ttattgtctc atgagcggat acatatttga
     5881 atgtatttag aaaaataaac aaataggggt tccgcgcaca tttccccgaa aagtgccacc
     5941 tgacgtcgac ggatcgggag atctcccgat cccctatggt gcactctcag tacaatctgc
     6001 tctgatgccg catagttaag ccagtatctg ctccctgctt gtgtgttgga ggtcgctgag
     6061 tagtgcgcga gcaaaattta agctacaaca aggcaaggct tgaccgacaa ttgcatgaag
     6121 aatctgctta gggttaggcg ttttgcgctg cttcgcgatg tacgggccag atatacgcgt
     6181 t
//
