LOCUS       sequence-211556-        7186 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 7186)
  AUTHORS   Arking DE, Krebsova A, Macek M, Macek M, Arking A, Mian IS, Fried L,
            Hamosh A, Dey S, McIntosh I, Dietz HC
  TITLE     Association of human aging with a functional variant of klotho.
  JOURNAL   Proc Natl Acad Sci U S A. 2002 Jan 22. 99(2):856-61.
  PUBMED    11792841
REFERENCE   2  (bases 1 to 7186)
  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. 2002 Jan 22. 99(2):856-61."
COMMENT     Sequence Label: Klotho (secreted)
FEATURES             Location/Qualifiers
     source          1..7186
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(44..63)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        235..614
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        615..818
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     769..789
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     863..882
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        863..881
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             2684..2725
                     /codon_start=1
                     /product="epitope tag from simian virus 5"
                     /label=V5 tag
                     /translation="GKPIPNPLLGLDST"
     CDS             2735..2752
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(2775..2792)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    2781..3005
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      3051..3479
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3138..3157)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3348..3369
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(3488..3508)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        3493..3822
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      3673..3808
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     3735..3754
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             3889..4683
                     /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(3943..3962)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4553..4572
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    4859..4980
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4896..4915)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4950..4969
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(5029..5045)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5029..5045)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(5042..5064)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5053..5069
                     /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(5077..5083,5084..5101,5102..5107))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5122..5143
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5260..5277)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5431..6019)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5511..5530)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6190..6981,6982..7050))
                     /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     6813..6832
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7051..7155)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 gacggatcgg gagatctccc gatcccctat ggtgcactct cagtacaatc tgctctgatg
       61 ccgcatagtt aagccagtat ctgctccctg cttgtgtgtt ggaggtcgct gagtagtgcg
      121 cgagcaaaat ttaagctaca acaaggcaag gcttgaccga caattgcatg aagaatctgc
      181 ttagggttag gcgttttgcg ctgcttcgcg atgtacgggc cagatatacg cgttgacatt
      241 gattattgac tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata
      301 tggagttccg cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc
      361 cccgcccatt gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc
      421 attgacgtca atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt
      481 atcatatgcc aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt
      541 atgcccagta catgacctta tgggactttc ctacttggca gtacatctac gtattagtca
      601 tcgctattac catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg
      661 actcacgggg atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc
      721 aaaatcaacg ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg
      781 gtaggcgtgt acggtgggag gtctatataa gcagagctct ctggctaact agagaaccca
      841 ctgcttactg gcttatcgaa attaatacga ctcactatag ggagacccaa gctggctagt
      901 taagcttggt accgagctcg gatccactag tccagtgtgg tggaattgcc cttcctggct
      961 cccgcgcagc atgcccgcca gcgccccgcc gcgccgcccg cggccgccgc cgccgtcgct
     1021 gtcgctgctg ctggtgctgc tgggcctggg cggccgccgc ctgcgtgcgg agccgggcga
     1081 cggcgcgcag acctgggccc gtttctcgcg gcctcctgcc cccgaggccg cgggcctctt
     1141 ccagggcacc ttccccgacg gcttcctctg ggccgtgggc agcgccgcct accagaccga
     1201 gggcggctgg cagcagcacg gcaagggtgc gtccatctgg gacacgttca cccaccaccc
     1261 cctggcaccc ccgggagact cccggaacgc cagtctgccg ttgggcgccc cgtcgccgct
     1321 gcagcccgcc accggggacg tagccagcga cagctacaac aacgtcttcc gcgacacgga
     1381 ggcgctgcgc gagctcgggg tcactcacta ccgcttctcc atctcgtggg cgcgagtgct
     1441 ccccaatggc agcgcgggcg tccccaaccg cgaggggctg cgctactacc ggcgcctgct
     1501 ggagcggctg cgggagctgg gcgtgcagcc cgtggtcacc ctgtaccact gggacctgcc
     1561 ccagcgcctg caggacgcct acggcggctg ggccaaccgc gccctggccg accacttcag
     1621 ggattacgcg gagctctgct tccgccactt cggcggtcag gtcaagtact ggatcaccat
     1681 cgacaacccc tacgtggtgg cctggcacgg ctacgccacc gggcgcctgg cccccggcat
     1741 ccggggcagc ccgcggctcg ggtacctggt ggcgcacaac ctcctcctgg ctcatgccaa
     1801 agtctggcat ctctacaata cttctttccg tcccactcag ggaggtcagg tgtccattgc
     1861 cctaagctct cactggatca atcctcgaag aatgaccgac cacagcatca aagaatgtca
     1921 aaaatctctg gactttgtac taggttggtt tgccaaaccc gtatttattg atggtgacta
     1981 tcccgagagc atgaagaata acctttcatc tattctgcct gattttactg aatctgagaa
     2041 aaagttcatc aaaggaactg ctgacttttt tgctctttgc tttggaccca ccttgagttt
     2101 tcaacttttg gaccctcaca tgaagttccg ccaattggaa tctcccaacc tgaggcaact
     2161 gctttcctgg attgaccttg aatttaacca tcctcaaata tttattgtgg aaaatggctg
     2221 gtttgtctca gggaccacca agagagatga tgccaaatat atgtattacc tcaaaaagtt
     2281 catcatggaa accttaaaag ccatcaagct ggatggggtg gatgtcatcg ggtataccgc
     2341 atggtccctc atggatggtt tcgagtggca cagaggttac agcatcaggc gtggactctt
     2401 ctatgttgac tttctaagcc aggacaagat gttgttgcca aagtcttcag ccttgttcta
     2461 ccaaaagctg atagagaaaa atggcttccc tcccttacct gaaaatcagc ccctagaagg
     2521 gacatttccc tgtgactttg cttggggagt tgttgacaac tacattcaag taagtcagct
     2581 gacaaaacca atcagcagtc tcaccaagcc ctatcacaag ggcaattctg cagatatcca
     2641 gcacagtggc ggccgctcga gtctagaggg cccgcggttc gaaggtaagc ctatccctaa
     2701 ccctctcctc ggtctcgatt ctacgcgtac cggtcatcat caccatcacc attgagttta
     2761 aacccgctga tcagcctcga ctgtgccttc tagttgccag ccatctgttg tttgcccctc
     2821 ccccgtgcct tccttgaccc tggaaggtgc cactcccact gtcctttcct aataaaatga
     2881 ggaaattgca tcgcattgtc tgagtaggtg tcattctatt ctggggggtg gggtggggca
     2941 ggacagcaag ggggaggatt gggaagacaa tagcaggcat gctggggatg cggtgggctc
     3001 tatggcttct gaggcggaaa gaaccagctg gggctctagg gggtatcccc acgcgccctg
     3061 tagcggcgca ttaagcgcgg cgggtgtggt ggttacgcgc agcgtgaccg ctacacttgc
     3121 cagcgcccta gcgcccgctc ctttcgcttt cttcccttcc tttctcgcca cgttcgccgg
     3181 ctttccccgt caagctctaa atcgggggct ccctttaggg ttccgattta gtgctttacg
     3241 gcacctcgac cccaaaaaac ttgattaggg tgatggttca cgtagtgggc catcgccctg
     3301 atagacggtt tttcgccctt tgacgttgga gtccacgttc tttaatagtg gactcttgtt
     3361 ccaaactgga acaacactca accctatctc ggtctattct tttgatttat aagggatttt
     3421 gccgatttcg gcctattggt taaaaaatga gctgatttaa caaaaattta acgcgaatta
     3481 attctgtgga atgtgtgtca gttagggtgt ggaaagtccc caggctcccc agcaggcaga
     3541 agtatgcaaa gcatgcatct caattagtca gcaaccaggt gtggaaagtc cccaggctcc
     3601 ccagcaggca gaagtatgca aagcatgcat ctcaattagt cagcaaccat agtcccgccc
     3661 ctaactccgc ccatcccgcc cctaactccg cccagttccg cccattctcc gccccatggc
     3721 tgactaattt tttttattta tgcagaggcc gaggccgcct ctgcctctga gctattccag
     3781 aagtagtgag gaggcttttt tggaggccta ggcttttgca aaaagctccc gggagcttgt
     3841 atatccattt tcggatctga tcaagagaca ggatgaggat cgtttcgcat gattgaacaa
     3901 gatggattgc acgcaggttc tccggccgct tgggtggaga ggctattcgg ctatgactgg
     3961 gcacaacaga caatcggctg ctctgatgcc gccgtgttcc ggctgtcagc gcaggggcgc
     4021 ccggttcttt ttgtcaagac cgacctgtcc ggtgccctga atgaactgca ggacgaggca
     4081 gcgcggctat cgtggctggc cacgacgggc gttccttgcg cagctgtgct cgacgttgtc
     4141 actgaagcgg gaagggactg gctgctattg ggcgaagtgc cggggcagga tctcctgtca
     4201 tctcaccttg ctcctgccga gaaagtatcc atcatggctg atgcaatgcg gcggctgcat
     4261 acgcttgatc cggctacctg cccattcgac caccaagcga aacatcgcat cgagcgagca
     4321 cgtactcgga tggaagccgg tcttgtcgat caggatgatc tggacgaaga gcatcagggg
     4381 ctcgcgccag ccgaactgtt cgccaggctc aaggcgcgca tgcccgacgg cgaggatctc
     4441 gtcgtgaccc atggcgatgc ctgcttgccg aatatcatgg tggaaaatgg ccgcttttct
     4501 ggattcatcg actgtggccg gctgggtgtg gcggaccgct atcaggacat agcgttggct
     4561 acccgtgata ttgctgaaga gcttggcggc gaatgggctg accgcttcct cgtgctttac
     4621 ggtatcgccg ctcccgattc gcagcgcatc gccttctatc gccttcttga cgagttcttc
     4681 tgagcgggac tctggggttc gcgaaatgac cgaccaagcg acgcccaacc tgccatcacg
     4741 agatttcgat tccaccgccg ccttctatga aaggttgggc ttcggaatcg ttttccggga
     4801 cgccggctgg atgatcctcc agcgcgggga tctcatgctg gagttcttcg cccaccccaa
     4861 cttgtttatt gcagcttata atggttacaa ataaagcaat agcatcacaa atttcacaaa
     4921 taaagcattt ttttcactgc attctagttg tggtttgtcc aaactcatca atgtatctta
     4981 tcatgtctgt ataccgtcga cctctagcta gagcttggcg taatcatggt catagctgtt
     5041 tcctgtgtga aattgttatc cgctcacaat tccacacaac atacgagccg gaagcataaa
     5101 gtgtaaagcc tggggtgcct aatgagtgag ctaactcaca ttaattgcgt tgcgctcact
     5161 gcccgctttc cagtcgggaa acctgtcgtg ccagctgcat taatgaatcg gccaacgcgc
     5221 ggggagaggc ggtttgcgta ttgggcgctc ttccgcttcc tcgctcactg actcgctgcg
     5281 ctcggtcgtt cggctgcggc gagcggtatc agctcactca aaggcggtaa tacggttatc
     5341 cacagaatca ggggataacg caggaaagaa catgtgagca aaaggccagc aaaaggccag
     5401 gaaccgtaaa aaggccgcgt tgctggcgtt tttccatagg ctccgccccc ctgacgagca
     5461 tcacaaaaat cgacgctcaa gtcagaggtg gcgaaacccg acaggactat aaagatacca
     5521 ggcgtttccc cctggaagct ccctcgtgcg ctctcctgtt ccgaccctgc cgcttaccgg
     5581 atacctgtcc gcctttctcc cttcgggaag cgtggcgctt tctcatagct cacgctgtag
     5641 gtatctcagt tcggtgtagg tcgttcgctc caagctgggc tgtgtgcacg aaccccccgt
     5701 tcagcccgac cgctgcgcct tatccggtaa ctatcgtctt gagtccaacc cggtaagaca
     5761 cgacttatcg ccactggcag cagccactgg taacaggatt agcagagcga ggtatgtagg
     5821 cggtgctaca gagttcttga agtggtggcc taactacggc tacactagaa gaacagtatt
     5881 tggtatctgc gctctgctga agccagttac cttcggaaaa agagttggta gctcttgatc
     5941 cggcaaacaa accaccgctg gtagcggtgg tttttttgtt tgcaagcagc agattacgcg
     6001 cagaaaaaaa ggatctcaag aagatccttt gatcttttct acggggtctg acgctcagtg
     6061 gaacgaaaac tcacgttaag ggattttggt catgagatta tcaaaaagga tcttcaccta
     6121 gatcctttta aattaaaaat gaagttttaa atcaatctaa agtatatatg agtaaacttg
     6181 gtctgacagt taccaatgct taatcagtga ggcacctatc tcagcgatct gtctatttcg
     6241 ttcatccata gttgcctgac tccccgtcgt gtagataact acgatacggg agggcttacc
     6301 atctggcccc agtgctgcaa tgataccgcg agacccacgc tcaccggctc cagatttatc
     6361 agcaataaac cagccagccg gaagggccga gcgcagaagt ggtcctgcaa ctttatccgc
     6421 ctccatccag tctattaatt gttgccggga agctagagta agtagttcgc cagttaatag
     6481 tttgcgcaac gttgttgcca ttgctacagg catcgtggtg tcacgctcgt cgtttggtat
     6541 ggcttcattc agctccggtt cccaacgatc aaggcgagtt acatgatccc ccatgttgtg
     6601 caaaaaagcg gttagctcct tcggtcctcc gatcgttgtc agaagtaagt tggccgcagt
     6661 gttatcactc atggttatgg cagcactgca taattctctt actgtcatgc catccgtaag
     6721 atgcttttct gtgactggtg agtactcaac caagtcattc tgagaatagt gtatgcggcg
     6781 accgagttgc tcttgcccgg cgtcaatacg ggataatacc gcgccacata gcagaacttt
     6841 aaaagtgctc atcattggaa aacgttcttc ggggcgaaaa ctctcaagga tcttaccgct
     6901 gttgagatcc agttcgatgt aacccactcg tgcacccaac tgatcttcag catcttttac
     6961 tttcaccagc gtttctgggt gagcaaaaac aggaaggcaa aatgccgcaa aaaagggaat
     7021 aagggcgaca cggaaatgtt gaatactcat actcttcctt tttcaatatt attgaagcat
     7081 ttatcagggt tattgtctca tgagcggata catatttgaa tgtatttaga aaaataaaca
     7141 aataggggtt ccgcgcacat ttccccgaaa agtgccacct gacgtc
//
