LOCUS       sequence-378589-        7778 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Mammalian expression plasmid for human CBS.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7778)
  AUTHORS   Switzer CH, Kasamatsu S, Ihara H, Eaton P
  TITLE     SOD1 is an essential H(2)S detoxifying enzyme.
  JOURNAL   Proc Natl Acad Sci U S A. 2023 Jan 17;120(3):e2205044120. doi: 
            10.1073/pnas.2205044120. Epub 2023 Jan 11.
  PUBMED    36630448
REFERENCE   2  (bases 1 to 7778)
  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: "Proc Natl 
            Acad Sci U S A."; date: "2023-01-17"; volume: "120(3):e2205044120. 
            doi"; pages: " 10.1073/pnas.2205044120. Epub 2023 Jan 11"
COMMENT     Sequence Label: GFP-CBS (WT)
FEATURES             Location/Qualifiers
     source          1..7778
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        53..432
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        433..636
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     587..607
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     681..700
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        681..699
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             741..1454
                     /codon_start=1
                     /product="green fluorescent protein"
                     /label=GFP
                     /translation="MSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKV
                     NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLLE
                     FVTAAGITHGMDELYK"
     CDS             join(741..743,744..1454)
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="MSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKV
                     NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLLE
                     FVTAAGITHGMDELYK"
     primer_bind     complement(783..804)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(1044..1063)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1391..1412
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     complement(3187..3204)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    3193..3417
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      3463..3891
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3550..3569)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3760..3781
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(3900..3920)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        3905..4234
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      4085..4220
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     4147..4166
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             4301..5095
                     /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(4355..4374)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4965..4984
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    5269..5390
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5306..5325)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     5360..5379
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(5439..5455)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5439..5455)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(5452..5474)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5463..5479
                     /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(5487..5493,5494..5511,5512..5517))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5532..5553
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5670..5687)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5841..6429)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5921..5940)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6600..7391,7392..7460))
                     /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     7223..7242
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7461..7565)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(7640..7659)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
ORIGIN
        1 agggttaggc gttttgcgct gcttcgcgat gtacgggcca gatatacgcg ttgacattga
       61 ttattgacta gttattaata gtaatcaatt acggggtcat tagttcatag cccatatatg
      121 gagttccgcg ttacataact tacggtaaat ggcccgcctg gctgaccgcc caacgacccc
      181 cgcccattga cgtcaataat gacgtatgtt cccatagtaa cgccaatagg gactttccat
      241 tgacgtcaat gggtggagta tttacggtaa actgcccact tggcagtaca tcaagtgtat
      301 catatgccaa gtacgccccc tattgacgtc aatgacggta aatggcccgc ctggcattat
      361 gcccagtaca tgaccttatg ggactttcct acttggcagt acatctacgt attagtcatc
      421 gctattacca tggtgatgcg gttttggcag tacatcaatg ggcgtggata gcggtttgac
      481 tcacggggat ttccaagtct ccaccccatt gacgtcaatg ggagtttgtt ttggcaccaa
      541 aatcaacggg actttccaaa atgtcgtaac aactccgccc cattgacgca aatgggcggt
      601 aggcgtgtac ggtgggaggt ctatataagc agagctctct ggctaactag agaacccact
      661 gcttactggc ttatcgaaat taatacgact cactataggg agacccaagc tggctagcgt
      721 ttaaacttaa gcttgccacc atgagcaagg gcgaggagct gttcaccggg gtggtgccca
      781 tcctggtcga gctggacggc gacgtaaacg gccacaagtt cagcgtgtcc ggcgagggcg
      841 agggcgatgc cacctacggc aagctgaccc tgaagttcat ctgcaccacc ggcaagctgc
      901 ccgtgccctg gcccaccctc gtgaccaccc tgacctacgg cgtgcagtgc ttcagccgct
      961 accccgacca catgaagcag cacgacttct tcaagtccgc catgcccgaa ggctacgtcc
     1021 aggagcgcac catcttcttc aaggacgacg gcaactacaa gacccgcgcc gaggtgaagt
     1081 tcgagggcga caccctggtg aaccgcatcg agctgaaggg catcgacttc aaggaggacg
     1141 gcaacatcct ggggcacaag ctggagtaca actacaacag ccacaacgtc tatatcatgg
     1201 ccgacaagca gaagaacggc atcaaggtga acttcaagat ccgccacaac atcgaggacg
     1261 gcagcgtgca gctcgccgac cactaccagc agaacacccc catcggcgac ggccccgtgc
     1321 tgctgcccga caaccactac ctgagcaccc agtccgccct gagcaaagac cccaacgaga
     1381 agcgcgatca catggtcctg ctggagttcg tgaccgccgc cgggatcact cacggcatgg
     1441 acgagctgta caagggtacc atgccttctg agacccccca ggcagaagtg gggcccacag
     1501 gctgccccca ccgctcaggg ccacactcgg cgaaggggag cctggagaag gggtccccag
     1561 aggataagga agccaaggag cccctgtgga tccggcccga tgctccgagc aggtgcacct
     1621 ggcagctggg ccggcctgcc tccgagtccc cacatcacca cactgccccg gcaaaatctc
     1681 caaaaatctt gccagatatt ctgaagaaaa tcggggacac ccctatggtc agaatcaaca
     1741 agattgggaa gaagttcggc ctgaagtgtg agctcttggc caagtgtgag ttcttcaacg
     1801 cgggcgggag cgtgaaggac cgcatcagcc tgcggatgat tgaggatgct gagcgcgacg
     1861 ggacgctgaa gcccggggac acgattatcg agccgacatc cgggaacacc gggatcgggc
     1921 tggccctggc tgcggcagtg aggggctatc gctgcatcat cgtgatgcca gagaagatga
     1981 gctccgagaa ggtggacgtg ctgcgggcac tgggggctga gattgtgagg acgcccacca
     2041 atgccaggtt cgactccccg gagtcacacg tgggggtggc ctggcggctg aagaacgaaa
     2101 tccccaattc tcacatccta gaccagtacc gcaacgccag caaccccctg gctcactacg
     2161 acaccaccgc tgatgagatc ctgcagcagt gtgatgggaa gctggacatg ctggtggctt
     2221 cagtgggcac gggcggcacc atcacgggca ttgccaggaa gctgaaggag aagtgtcctg
     2281 gatgcaggat cattggggtg gatcccgaag ggtccatcct cgcagagccg gaggagctga
     2341 accagacgga gcagacaacc tacgaggtgg aagggatcgg ctacgacttc atccccacgg
     2401 tgctggacag gacggtggtg gacaagtggt tcaagagcaa cgatgaggag gcgttcacct
     2461 ttgcccgcat gctgatcgcg caagaggggc tgctgtgcgg tggcagtgct ggcagcacgg
     2521 tggcggtggc cgtgaaggcc gcgcaggagc tgcaggaggg ccagcgctgc gtggtcattc
     2581 tgcccgactc agtgcggaac tacatgacca agttcctgag cgacaggtgg atgctgcaga
     2641 agggctttct gaaggaggag gacctcacgg agaagaagcc ctggtggtgg cacctccgtg
     2701 ttcaggagct gggcctgtca gccccgctga ccgtgctccc gaccatcacc tgtgggcaca
     2761 ccatcgagat cctccgggag aagggcttcg accaggcgcc cgtggtggat gaggcggggg
     2821 taatcctggg aatggtgacg cttgggaaca tgctctcgtc cctgcttgcc gggaaggtgc
     2881 agccgtcaga ccaagttggc aaagtcatct acaagcagtt caaacagatc cgcctcacgg
     2941 acacgctggg caggctctcg cacatcctgg agatggacca cttcgccctg gtggtgcacg
     3001 agcagatcca gtaccacagc accgggaagt ccagtcagcg gcagatggtg ttcggggtgg
     3061 tcaccgccat tgacttgctg aacttcgtgg ccgcccagga gcgggaccag aagtgagaat
     3121 tctgcagata tccagcacag tggcggccgc tcgagtctag agggcccgtt taaacccgct
     3181 gatcagcctc gactgtgcct tctagttgcc agccatctgt tgtttgcccc tcccccgtgc
     3241 cttccttgac cctggaaggt gccactccca ctgtcctttc ctaataaaat gaggaaattg
     3301 catcgcattg tctgagtagg tgtcattcta ttctgggggg tggggtgggg caggacagca
     3361 agggggagga ttgggaagac aatagcaggc atgctgggga tgcggtgggc tctatggctt
     3421 ctgaggcgga aagaaccagc tggggctcta gggggtatcc ccacgcgccc tgtagcggcg
     3481 cattaagcgc ggcgggtgtg gtggttacgc gcagcgtgac cgctacactt gccagcgccc
     3541 tagcgcccgc tcctttcgct ttcttccctt cctttctcgc cacgttcgcc ggctttcccc
     3601 gtcaagctct aaatcggggg ctccctttag ggttccgatt tagtgcttta cggcacctcg
     3661 accccaaaaa acttgattag ggtgatggtt cacgtagtgg gccatcgccc tgatagacgg
     3721 tttttcgccc tttgacgttg gagtccacgt tctttaatag tggactcttg ttccaaactg
     3781 gaacaacact caaccctatc tcggtctatt cttttgattt ataagggatt ttgccgattt
     3841 cggcctattg gttaaaaaat gagctgattt aacaaaaatt taacgcgaat taattctgtg
     3901 gaatgtgtgt cagttagggt gtggaaagtc cccaggctcc ccagcaggca gaagtatgca
     3961 aagcatgcat ctcaattagt cagcaaccag gtgtggaaag tccccaggct ccccagcagg
     4021 cagaagtatg caaagcatgc atctcaatta gtcagcaacc atagtcccgc ccctaactcc
     4081 gcccatcccg cccctaactc cgcccagttc cgcccattct ccgccccatg gctgactaat
     4141 tttttttatt tatgcagagg ccgaggccgc ctctgcctct gagctattcc agaagtagtg
     4201 aggaggcttt tttggaggcc taggcttttg caaaaagctc ccgggagctt gtatatccat
     4261 tttcggatct gatcaagaga caggatgagg atcgtttcgc atgattgaac aagatggatt
     4321 gcacgcaggt tctccggccg cttgggtgga gaggctattc ggctatgact gggcacaaca
     4381 gacaatcggc tgctctgatg ccgccgtgtt ccggctgtca gcgcaggggc gcccggttct
     4441 ttttgtcaag accgacctgt ccggtgccct gaatgaactg caggacgagg cagcgcggct
     4501 atcgtggctg gccacgacgg gcgttccttg cgcagctgtg ctcgacgttg tcactgaagc
     4561 gggaagggac tggctgctat tgggcgaagt gccggggcag gatctcctgt catctcacct
     4621 tgctcctgcc gagaaagtat ccatcatggc tgatgcaatg cggcggctgc atacgcttga
     4681 tccggctacc tgcccattcg accaccaagc gaaacatcgc atcgagcgag cacgtactcg
     4741 gatggaagcc ggtcttgtcg atcaggatga tctggacgaa gagcatcagg ggctcgcgcc
     4801 agccgaactg ttcgccaggc tcaaggcgcg catgcccgac ggcgaggatc tcgtcgtgac
     4861 ccatggcgat gcctgcttgc cgaatatcat ggtggaaaat ggccgctttt ctggattcat
     4921 cgactgtggc cggctgggtg tggcggaccg ctatcaggac atagcgttgg ctacccgtga
     4981 tattgctgaa gagcttggcg gcgaatgggc tgaccgcttc ctcgtgcttt acggtatcgc
     5041 cgctcccgat tcgcagcgca tcgccttcta tcgccttctt gacgagttct tctgagcggg
     5101 actctggggt tcgaaatgac cgaccaagcg acgcccaacc tgccatcacg agatttcgat
     5161 tccaccgccg ccttctatga aaggttgggc ttcggaatcg ttttccggga cgccggctgg
     5221 atgatcctcc agcgcgggga tctcatgctg gagttcttcg cccaccccaa cttgtttatt
     5281 gcagcttata atggttacaa ataaagcaat agcatcacaa atttcacaaa taaagcattt
     5341 ttttcactgc attctagttg tggtttgtcc aaactcatca atgtatctta tcatgtctgt
     5401 ataccgtcga cctctagcta gagcttggcg taatcatggt catagctgtt tcctgtgtga
     5461 aattgttatc cgctcacaat tccacacaac atacgagccg gaagcataaa gtgtaaagcc
     5521 tggggtgcct aatgagtgag ctaactcaca ttaattgcgt tgcgctcact gcccgctttc
     5581 cagtcgggaa acctgtcgtg ccagctgcat taatgaatcg gccaacgcgc ggggagaggc
     5641 ggtttgcgta ttgggcgctc ttccgcttcc tcgctcactg actcgctgcg ctcggtcgtt
     5701 cggctgcggc gagcggtatc agctcactca aaggcggtaa tacggttatc cacagaatca
     5761 ggggataacg caggaaagaa catgtgagca aaaggccagc aaaaggccag gaaccgtaaa
     5821 aaggccgcgt tgctggcgtt tttccatagg ctccgccccc ctgacgagca tcacaaaaat
     5881 cgacgctcaa gtcagaggtg gcgaaacccg acaggactat aaagatacca ggcgtttccc
     5941 cctggaagct ccctcgtgcg ctctcctgtt ccgaccctgc cgcttaccgg atacctgtcc
     6001 gcctttctcc cttcgggaag cgtggcgctt tctcatagct cacgctgtag gtatctcagt
     6061 tcggtgtagg tcgttcgctc caagctgggc tgtgtgcacg aaccccccgt tcagcccgac
     6121 cgctgcgcct tatccggtaa ctatcgtctt gagtccaacc cggtaagaca cgacttatcg
     6181 ccactggcag cagccactgg taacaggatt agcagagcga ggtatgtagg cggtgctaca
     6241 gagttcttga agtggtggcc taactacggc tacactagaa gaacagtatt tggtatctgc
     6301 gctctgctga agccagttac cttcggaaaa agagttggta gctcttgatc cggcaaacaa
     6361 accaccgctg gtagcggtgg tttttttgtt tgcaagcagc agattacgcg cagaaaaaaa
     6421 ggatctcaag aagatccttt gatcttttct acggggtctg acgctcagtg gaacgaaaac
     6481 tcacgttaag ggattttggt catgagatta tcaaaaagga tcttcaccta gatcctttta
     6541 aattaaaaat gaagttttaa atcaatctaa agtatatatg agtaaacttg gtctgacagt
     6601 taccaatgct taatcagtga ggcacctatc tcagcgatct gtctatttcg ttcatccata
     6661 gttgcctgac tccccgtcgt gtagataact acgatacggg agggcttacc atctggcccc
     6721 agtgctgcaa tgataccgcg agacccacgc tcaccggctc cagatttatc agcaataaac
     6781 cagccagccg gaagggccga gcgcagaagt ggtcctgcaa ctttatccgc ctccatccag
     6841 tctattaatt gttgccggga agctagagta agtagttcgc cagttaatag tttgcgcaac
     6901 gttgttgcca ttgctacagg catcgtggtg tcacgctcgt cgtttggtat ggcttcattc
     6961 agctccggtt cccaacgatc aaggcgagtt acatgatccc ccatgttgtg caaaaaagcg
     7021 gttagctcct tcggtcctcc gatcgttgtc agaagtaagt tggccgcagt gttatcactc
     7081 atggttatgg cagcactgca taattctctt actgtcatgc catccgtaag atgcttttct
     7141 gtgactggtg agtactcaac caagtcattc tgagaatagt gtatgcggcg accgagttgc
     7201 tcttgcccgg cgtcaatacg ggataatacc gcgccacata gcagaacttt aaaagtgctc
     7261 atcattggaa aacgttcttc ggggcgaaaa ctctcaagga tcttaccgct gttgagatcc
     7321 agttcgatgt aacccactcg tgcacccaac tgatcttcag catcttttac tttcaccagc
     7381 gtttctgggt gagcaaaaac aggaaggcaa aatgccgcaa aaaagggaat aagggcgaca
     7441 cggaaatgtt gaatactcat actcttcctt tttcaatatt attgaagcat ttatcagggt
     7501 tattgtctca tgagcggata catatttgaa tgtatttaga aaaataaaca aataggggtt
     7561 ccgcgcacat ttccccgaaa agtgccacct gacgtcgacg gatcgggaga tctcccgatc
     7621 ccctatggtg cactctcagt acaatctgct ctgatgccgc atagttaagc cagtatctgc
     7681 tccctgcttg tgtgttggag gtcgctgagt agtgcgcgag caaaatttaa gctacaacaa
     7741 ggcaaggctt gaccgacaat tgcatgaaga atctgctt
//
