LOCUS       sequence-63663-e        8847 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 8847)
  AUTHORS   Phelps M, Stuelsatz P, Yablonka-Reuveni Z
  TITLE     Expression profile and overexpression outcome indicate a role for 
            betaKlotho in skeletal muscle fibro/adipogenesis.
  JOURNAL   FEBS J. 2016 May;283(9):1653-68. doi: 10.1111/febs.13682. Epub 2016 
            Apr 13.
  PUBMED    26881702
REFERENCE   2  (bases 1 to 8847)
  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: 
            "10.1111/febs.13682"; journalName: "FEBS J"; date: "2016-05"; 
            volume: "283"; issue: "9"; pages: "1653-68"
COMMENT     Sequence Label: pCMV-Klb-EGFP
FEATURES             Location/Qualifiers
     source          1..8847
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        10..389
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        390..593
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     544..564
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     638..657
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        638..656
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             join(3808..3810,3811..3813,3814..4527)
                     /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(3808..3810,3811..3813,3814..4524)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(3853..3874)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(4114..4133)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     4461..4482
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     promoter        complement(4562..4580)
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     complement(4563..4580)
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     primer_bind     complement(4600..4617)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    4606..4830
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      complement(4962..5550)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5042..5061)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     polyA_signal    5879..5926
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     primer_bind     5951..5970
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     CDS             complement(6158..6946)
                     /codon_start=1
                     /product="aminoglycoside phosphotransferase"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDQEHQGL
                     APAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIALA
                     TRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(6269..6288)
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     6873..6892
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     promoter        complement(6981..7338)
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      6995..7130
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     enhancer        7147..7338
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     7295..7315
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     CDS             complement(join(7398..8189,8190..8258))
                     /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     8021..8040
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(8259..8363)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      8390..8845
                     /direction=RIGHT
                     /label=M13 ori
                     /note="M13 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(8477..8496)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     8687..8708
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
ORIGIN
        1 gcgcgcgttg acattgatta ttgactagtt attaatagta atcaattacg gggtcattag
       61 ttcatagccc atatatggag ttccgcgtta cataacttac ggtaaatggc ccgcctggct
      121 gaccgcccaa cgacccccgc ccattgacgt caataatgac gtatgttccc atagtaacgc
      181 caatagggac tttccattga cgtcaatggg tggactattt acggtaaact gcccacttgg
      241 cagtacatca agtgtatcat atgccaagta cgccccctat tgacgtcaat gacggtaaat
      301 ggcccgcctg gcattatgcc cagtacatga ccttatggga ctttcctact tggcagtaca
      361 tctacgtatt agtcatcgct attaccatgg tgatgcggtt ttggcagtac atcaatgggc
      421 gtggatagcg gtttgactca cggggatttc caagtctcca ccccattgac gtcaatggga
      481 gtttgttttg gcaccaaaat caacgggact ttccaaaatg tcgtaacaac tccgccccat
      541 tgacgcaaat gggcggtagg cgtgtacggt gggaggtcta tataagcaga gctctctggc
      601 taactagaga acccactgct tactggctta tcgaaattaa tacgactcac tatagggaga
      661 cccaagcttt aaatgaagac aggctgtgca gcagggtctc cggggaatga atggattttc
      721 ttcagctctg atgaaagaaa cacacgctct aggaaaacaa tgtccaacag ggcactgcaa
      781 agatctgccg tgctgtctgc gtttgttctg ctgcgagctg ttaccggctt ctccggagac
      841 gggaaagcaa tatgggataa aaaacagtac gtgagtccgg taaacccaag tcagctgttc
      901 ctctatgaca ctttccctaa aaacttttcc tggggcgttg ggaccggagc atttcaagtg
      961 gaagggagtt ggaagacaga tggaagagga ccctcgatct gggatcggta cgtctactca
     1021 cacctgagag gtgtcaacgg cacagacaga tccactgaca gttacatctt tctggaaaaa
     1081 gacttgttgg ctctggattt tttaggagtt tctttttatc agttctcaat ctcctggcca
     1141 cggttgtttc ccaatggaac agtagcagca gtgaatgcgc aaggtctccg gtactaccgt
     1201 gcacttctgg actcgctggt acttaggaat atcgagccca ttgttacctt gtaccattgg
     1261 gatttgcctc tgacgctcca ggaagaatat gggggctgga aaaatgcaac tatgatagat
     1321 ctcttcaacg actatgccac atactgcttc cagacctttg gagaccgtgt caaatattgg
     1381 attacaattc acaaccctta ccttgttgct tggcatgggt ttggcacagg tatgcatgca
     1441 ccaggagaga agggaaattt aacagctgtc tacactgtgg gacacaacct gatcaaggca
     1501 cattcgaaag tgtggcataa ctacgacaaa aacttccgcc ctcatcagaa gggttggctc
     1561 tccatcacct tggggtccca ttggatagag ccaaacagaa cagacaacat ggaggacgtg
     1621 atcaactgcc agcactccat gtcctctgtg cttggatggt tcgccaaccc catccacggg
     1681 gacggcgact accctgagtt catgaagacg ggcgccatga tccccgagtt ctctgaggca
     1741 gagaaggagg aggtgagggg cacggctgat ttctttgcct tttccttcgg gcccaacaac
     1801 ttcaggccct caaacaccgt ggtgaaaatg ggacaaaatg tatcactcaa cttaaggcag
     1861 gtgctgaact ggattaaact ggaatacgat gaccctcaaa tcttgatttc ggagaacggc
     1921 tggttcacag atagctatat aaagacagag gacaccacgg ccatctacat gatgaagaat
     1981 ttcctaaacc aggttcttca agcaataaaa tttgatgaaa tccgcgtgtt tggttatacg
     2041 gcctggactc tcctggatgg ctttgagtgg caggatgcct atacgacccg acgagggctg
     2101 ttttatgtgg actttaacag tgagcagaaa gagaggaaac ccaagtcctc ggctcattac
     2161 tacaagcaga tcatacaaga caacggcttc cctttgaaag agtccacgcc agacatgaag
     2221 ggtcggttcc cctgtgattt ctcttgggga gtcactgagt ctgttcttaa gcccgagttt
     2281 acggtctcct ccccgcagtt taccgatcct cacctgtatg tgtggaatgt cactggcaac
     2341 agattgctct accgagtgga aggggtaagg ctgaaaacaa gaccatccca gtgcacagat
     2401 tatgtgagca tcaaaaaacg agttgaaatg ttggcaaaaa tgaaagtcac ccactaccag
     2461 tttgctctgg actggacctc tatccttccc actggcaatc tgtccaaagt taacagacaa
     2521 gtgttaaggt actataggtg tgtggtgagc gaaggactga agctgggcgt cttccccatg
     2581 gtgacgttgt accacccaac ccactcccat ctcggcctcc ccctgccact tctgagcagt
     2641 ggggggtggc taaacatgaa cacagccaag gccttccagg actacgctga gctgtgcttc
     2701 cgggagttgg gggacttggt gaagctctgg atcaccatca atgagcctaa caggctgagt
     2761 gacatgtaca accgcacgag taatgacacc taccgtgcag cccacaacct gatgatcgcc
     2821 catgcccagg tctggcacct ctatgatagg cagtataggc cggtccagca tggggctgtg
     2881 tcgctgtcct tacattgcga ctgggcagaa cctgccaacc cctttgtgga ttcacactgg
     2941 aaggcagccg agcgcttcct ccagtttgag atcgcctggt ttgcagatcc gctcttcaag
     3001 actggcgact atccatcggt tatgaaggaa tacatcgcct ccaagaacca gcgagggctg
     3061 tctagctcag tcctgccgcg cttcaccgcg aaggagagca ggctggtgaa gggtaccgtc
     3121 gacttctacg cactgaacca cttcactacg aggttcgtga tacacaagca gctgaacacc
     3181 aaccgctcag ttgcagacag ggacgtccag ttcctgcagg acatcacccg cctaagctcg
     3241 cccagccgcc tggctgtaac accctgggga gtgcgcaagc tccttgcgtg gatccggagg
     3301 aactacagag acagggatat ctacatcaca gccaatggca tcgatgacct ggctctagag
     3361 gatgatcaga tccgaaagta ctacttggag aagtatgtcc aggaggctct gaaagcatat
     3421 ctcattgaca aggtcaaaat caaaggctac tatgcattca aactgactga agagaaatct
     3481 aagcctagat ttggattttt cacctctgac ttcagagcta agtcctctgt ccagttttac
     3541 agcaagctga tcagcagcag tggcctcccc gctgagaaca gaagtcctgc gtgtggtcag
     3601 cctgcggaag acacagactg caccatttgc tcatttctcg tggagaagaa accactcatc
     3661 ttcttcggtt gctgcttcat ctccactctg gctgtactgc tatccatcac cgtttttcat
     3721 catcaaaaga gaagaaaatt ccagaaagca aggaacttac aaaatatacc attgaagaaa
     3781 ggccacagca gagttttcag caccggtatg gtgagcaagg gcgaggagct gttcaccggg
     3841 gtggtgccca tcctggtcga gctggacggc gacgtaaacg gccacaagtt cagcgtgtcc
     3901 ggcgagggcg agggcgatgc cacctacggc aagctgaccc tgaagttcat ctgcaccacc
     3961 ggcaagctgc ccgtgccctg gcccaccctc gtgaccaccc tgacctacgg cgtgcagtgc
     4021 ttcagccgct accccgacca catgaagcag cacgacttct tcaagtccgc catgcccgaa
     4081 ggctacgtcc aggagcgcac catcttcttc aaggacgacg gcaactacaa gacccgcgcc
     4141 gaggtgaagt tcgagggcga caccctggtg aaccgcatcg agctgaaggg catcgacttc
     4201 aaggaggacg gcaacatcct ggggcacaag ctggagtaca actacaacag ccacaacgtc
     4261 tatatcatgg ccgacaagca gaagaacggc atcaaggtga acttcaagat ccgccacaac
     4321 atcgaggacg gcagcgtgca gctcgccgac cactaccagc agaacacccc catcggcgac
     4381 ggccccgtgc tgctgcccga caaccactac ctgagcaccc agtccgccct gagcaaagac
     4441 cccaacgaga agcgcgatca catggtcctg ctggagttcg tgaccgccgc cgggatcact
     4501 ctcggcatgg acgagctgta caagtaagcg gccgctcgag catgcatcta gagggcccta
     4561 ttctatagtg tcacctaaat gctagagctc gctgatcagc ctcgactgtg ccttctagtt
     4621 gccagccatc tgttgtttgc ccctcccccg tgccttcctt gaccctggaa ggtgccactc
     4681 ccactgtcct ttcctaataa aatgaggaaa ttgcatcgca ttgtctgagt aggtgtcatt
     4741 ctattctggg gggtggggtg gggcaggaca gcaaggggga ggattgggaa gacaatagca
     4801 ggcatgctgg ggatgcggtg ggctctatgg cttctgaggc ggaaagaacc agtggcggta
     4861 atacggttat ccacagaatc aggggataac gcaggaaaga acatgtgagc aaaaggccag
     4921 caaaaggcca ggaaccgtaa aaaggccgcg ttgctggcgt ttttccatag gctccgcccc
     4981 cctgacgagc atcacaaaaa tcgacgctca agtcagaggt ggcgaaaccc gacaggacta
     5041 taaagatacc aggcgtttcc ccctggaagc tccctcgtgc gctctcctgt tccgaccctg
     5101 ccgcttaccg gatacctgtc cgcctttctc ccttcgggaa gcgtggcgct ttctcatagc
     5161 tcacgctgta ggtatctcag ttcggtgtag gtcgttcgct ccaagctggg ctgtgtgcac
     5221 gaaccccccg ttcagcccga ccgctgcgcc ttatccggta actatcgtct tgagtccaac
     5281 ccggtaagac acgacttatc gccactggca gcagccactg gtaacaggat tagcagagcg
     5341 aggtatgtag gcggtgctac agagttcttg aagtggtggc ctaactacgg ctacactaga
     5401 aggacagtat ttggtatctg cgctctgctg aagccagtta ccttcggaaa aagagttggt
     5461 agctcttgat ccggcaaaca aaccaccgct ggtagcggtg gtttttttgt ttgcaagcag
     5521 cagattacgc gcagaaaaaa aggatctcaa gaagatcctt tgatcttttc tacggggtct
     5581 gacgctcagt ggaacgaaaa ctcacgttaa gggattttgg tcatgagatt atcaaaaagg
     5641 atcttcacct agatcctttt aaattaaaaa tgaagtttta aatcaatcta aagtatatat
     5701 gagtaacctg aggctatggc agggcctgcc gccccgacgt tggctgcgag ccctgggcct
     5761 tcacccgaac ttggggggtg gggtggggaa aaggaagaaa cgcgggcgta ttggccccaa
     5821 tggggtctcg gtggggtatc gacagagtgc cagccctggg accgaacccc gcgtttatga
     5881 acaaacgacc caacaccgtg cgttttattc tgtcttttta ttgccgtcat agcgcgggtt
     5941 ccttccggta ttgtctcctt ccgtgtttca gttagcctcc ccctagggtg ggcgaagaac
     6001 tccagcatga gatccccgcg ctggaggatc atccagccgg cgtcccggaa aacgattccg
     6061 aagcccaacc tttcatagaa ggcggcggtg gaatcgaaat ctcgtgatgg caggttgggc
     6121 gtcgcttggt cggtcatttc gaaccccaga gtcccgctca gaagaactcg tcaagaaggc
     6181 gatagaaggc gatgcgctgc gaatcgggag cggcgatacc gtaaagcacg aggaagcggt
     6241 cagcccattc gccgccaagc tcttcagcaa tatcacgggt agccaacgct atgtcctgat
     6301 agcggtccgc cacacccagc cggccacagt cgatgaatcc agaaaagcgg ccattttcca
     6361 ccatgatatt cggcaagcag gcatcgccat gggtcacgac gagatcctcg ccgtcgggca
     6421 tgctcgcctt gagcctggcg aacagttcgg ctggcgcgag cccctgatgc tcttgatcat
     6481 cctgatcgac aagaccggct tccatccgag tacgtgctcg ctcgatgcga tgtttcgctt
     6541 ggtggtcgaa tgggcaggta gccggatcaa gcgtatgcag ccgccgcatt gcatcagcca
     6601 tgatggatac tttctcggca ggagcaaggt gagatgacag gagatcctgc cccggcactt
     6661 cgcccaatag cagccagtcc cttcccgctt cagtgacaac gtcgagcaca gctgcgcaag
     6721 gaacgcccgt cgtggccagc cacgatagcc gcgctgcctc gtcttgcagt tcattcaggg
     6781 caccggacag gtcggtcttg acaaaaagaa ccgggcgccc ctgcgctgac agccggaaca
     6841 cggcggcatc agagcagccg attgtctgtt gtgcccagtc atagccgaat agcctctcca
     6901 cccaagcggc cggagaacct gcgtgcaatc catcttgttc aatcatgcga aacgatcctc
     6961 atcctgtctc ttgatcgatc tttgcaaaag cctaggcctc caaaaaagcc tcctcactac
     7021 ttctggaata gctcagaggc cgaggcggcc tcggcctctg cataaataaa aaaaattagt
     7081 cagccatggg gcggagaatg ggcggaactg ggcggagtta ggggcgggat gggcggagtt
     7141 aggggcggga ctatggttgc tgactaattg agatgcatgc tttgcatact tctgcctgct
     7201 ggggagcctg gggactttcc acacctggtt gctgactaat tgagatgcat gctttgcata
     7261 cttctgcctg ctggggagcc tggggacttt ccacacccta actgacacac attccacagc
     7321 tggttctttc cgcctcagga ctcttccttt ttcaataaat caatctaaag tatatatgag
     7381 taaacttggt ctgacagtta ccaatgctta atcagtgagg cacctatctc agcgatctgt
     7441 ctatttcgtt catccatagt tgcctgactc cccgtcgtgt agataactac gatacgggag
     7501 ggcttaccat ctggccccag tgctgcaatg ataccgcgag acccacgctc accggctcca
     7561 gatttatcag caataaacca gccagccgga agggccgagc gcagaagtgg tcctgcaact
     7621 ttatccgcct ccatccagtc tattaattgt tgccgggaag ctagagtaag tagttcgcca
     7681 gttaatagtt tgcgcaacgt tgttgccatt gctacaggca tcgtggtgtc acgctcgtcg
     7741 tttggtatgg cttcattcag ctccggttcc caacgatcaa ggcgagttac atgatccccc
     7801 atgttgtgca aaaaagcggt tagctccttc ggtcctccga tcgttgtcag aagtaagttg
     7861 gccgcagtgt tatcactcat ggttatggca gcactgcata attctcttac tgtcatgcca
     7921 tccgtaagat gcttttctgt gactggtgag tactcaacca agtcattctg agaatagtgt
     7981 atgcggcgac cgagttgctc ttgcccggcg tcaatacggg ataataccgc gccacatagc
     8041 agaactttaa aagtgctcat cattggaaaa cgttcttcgg ggcgaaaact ctcaaggatc
     8101 ttaccgctgt tgagatccag ttcgatgtaa cccactcgtg cacccaactg atcttcagca
     8161 tcttttactt tcaccagcgt ttctgggtga gcaaaaacag gaaggcaaaa tgccgcaaaa
     8221 aagggaataa gggcgacacg gaaatgttga atactcatac tcttcctttt tcaatattat
     8281 tgaagcattt atcagggtta ttgtctcatg agcggataca tatttgaatg tatttagaaa
     8341 aataaacaaa taggggttcc gcgcacattt ccccgaaaag tgccacctga cgcgccctgt
     8401 agcggcgcat taagcgcggc gggtgtggtg gttacgcgca gcgtgaccgc tacacttgcc
     8461 agcgccctag cgcccgctcc tttcgctttc ttcccttcct ttctcgccac gttcgccggc
     8521 tttccccgtc aagctctaaa tcgggggctc cctttagggt tccgatttag tgctttacgg
     8581 cacctcgacc ccaaaaaact tgattagggt gatggttcac gtagtgggcc atcgccctga
     8641 tagacggttt ttcgcccttt gacgttggag tccacgttct ttaatagtgg actcttgttc
     8701 caaactggaa caacactcaa ccctatctcg gtctattctt ttgatttata agggattttg
     8761 ccgatttcgg cctattggtt aaaaaatgag ctgatttaac aaaaatttaa cgcgaatttt
     8821 aacaaaatat taacgcttac aatttac
//
