LOCUS       sequence-220151-        8178 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses EGFP-Cre fusion under the control of the EF1alpha 
            promoter; EGFP is human codon-optimized..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8178)
  AUTHORS   Le Y, Miller JL, Sauer B
  TITLE     GFPcre fusion vectors with enhanced expression.
  JOURNAL   Anal Biochem. 1999 Jun 1. 270(2):334-6.
  PUBMED    10334853
REFERENCE   2  (bases 1 to 8178)
  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: "Anal 
            Biochem. 1999 Jun 1. 270(2):334-6."
COMMENT     Sequence Label: pBS598 EF1alpha-EGFPcre
FEATURES             Location/Qualifiers
     source          1..8178
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     1..17
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     promoter        107..1285
                     /label=EF-1-alpha promoter
                     /note="strong constitutive promoter for human elongation 
                     factor EF-1-alpha"
     intron          338..1276
                     /label=EF-1-alpha intron A
                     /note="intron upstream of the start codon of human 
                     EF-1-alpha"
     primer_bind     1233..1253
                     /label=EF1a-F
                     /note="Human elongation factor-1a promoter, forward primer"
     regulatory      1309..1318
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(1315..1317,1318..1320,1321..2031)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             join(1315..1317,1318..1320,1321..2031)
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(1621..1640)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1968..1989
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     CDS             2086..3117
                     /codon_start=1
                     /product="site-specific recombinase"
                     /label=Cre
                     /note="Cre is a site-specific recombinase from 
                     bacteriophage P1. Recombination occurs at loxP sequences."
                     /translation="MANLLTVHQNLPALPVDATSDEVRKNLMDMFRDRQAFSEHTWKML
                     LSVCRSWAAWCKLNNRKWFPAEPEDVRDYLLYLQARGLAVKTIQQHLGQLNMLHRRSGL
                     PRPSDSNAVSLVMRRIRKENVDAGERAKQALAFERTDFDQVRSLMENSDRCQDIRNLAF
                     LGIAYNTLLRIAEIARIRVKDISRTDGGRMLIHIGRTKTLVSTAGVEKALSLGVTKLVE
                     RWISVSGVADDPNNYLFCRVRKNGVAAPSATSQLSTRALEGIFEATHRLIYGAKDDSGQ
                     RYLAWSGHSARVGAARDMARAGVSIPEIMQAGGWTNVNIVMNYIRNLDSETGAMVRLLE
                     DGD"
     RBS             4769..4777
                     /label=Shine-Dalgarno sequence
                     /note="full consensus sequence for ribosome-binding sites 
                     upstream of start codons in E. coli; complementary to a 
                     region in the 3' end of the 16S rRNA (Chen et al., 1994)"
     primer_bind     complement(5579..5595)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5579..5595)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(5592..5614)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5603..5619
                     /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(5627..5633,5634..5651,5652..5657))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5672..5693
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5810..5827)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5981..6569)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(6061..6080)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6740..7531,7532..7600))
                     /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     7363..7382
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7601..7705)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     7773..7791
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(7829..7851)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     7951..7970
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     join(8164..8178,1..8)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     join(8178,1..17)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
ORIGIN
        1 gtaaaacgac ggccagtgcc aagctttgca aagatggata aagttttaaa cagagaggaa
       61 tctttgcagc taatggacct tctaggtctt gaaaggagtg ggaattggct ccggtgcccg
      121 tcagtgggca gagcgcacat cgcccacagt ccccgagaag ttggggggag gggtcggcaa
      181 ttgaaccggt gcctagagaa ggtggcgcgg ggtaaactgg gaaagtgatg tcgtgtactg
      241 gctccgcctt tttcccgagg gtgggggaga accgtatata agtgcagtag tcgccgtgaa
      301 cgttcttttt cgcaacgggt ttgccgccag aacacaggta agtgccgtgt gtggttcccg
      361 cgggcctggc ctctttacgg gttatggccc ttgcgtgcct tgaattactt ccacctggct
      421 gcagtacgtg attcttgatc ccgagcttcg ggttggaagt gggtgggaga gttcgaggcc
      481 ttgcgcttaa ggagcccctt cgcctcgtgc ttgagttgag gcctggcctg ggcgctgggg
      541 ccgccgcgtg cgaatctggt ggcaccttcg cgcctgtctc gctgctttcg ataagtctct
      601 agccatttaa aatttttgat gacctgctgc gacgcttttt ttctggcaag atagtcttgt
      661 aaatgcgggc caagatctgc acactggtat ttcggttttt ggggccgcgg gcggcgacgg
      721 ggcccgtgcg tcccagcgca catgttcggc gaggcggggc ctgcgagcgc ggccaccgag
      781 aatcggacgg gggtagtctc aagctggccg gcctgctctg gtgcctggcc tcgcgccgcc
      841 gtgtatcgcc ccgccctggg cggcaaggct ggcccggtcg gcaccagttg cgtgagcgga
      901 aagatggccg cttcccggcc ctgctgcagg gagctcaaaa tggaggacgc ggcgctcggg
      961 agagcgggcg ggtgagtcac ccacacaaag gaaaagggcc tttccgtcct cagccgtcgc
     1021 ttcatgtgac tccacggagt accgggcgcc gtccaggcac ctcgattagt tctcgagctt
     1081 ttggagtacg tcgtctttag gttgggggga ggggttttat gcgatggagt ttccccacac
     1141 tgagtgggtg gagactgaag ttaggccagc ttggcacttg atgtaattct ccttggaatt
     1201 tgcccttttt gagtttggat cttggttcat tctcaagcct cagacagtgg ttcaaagttt
     1261 ttttcttcca tttcaggtgt cgtgaggaat tctctagcgc taccggtcgc caccatggtg
     1321 agcaagggcg aggagctgtt caccggggtg gtgcccatcc tggtcgagct ggacggcgac
     1381 gtaaacggcc acaagttcag cgtgtccggc gagggcgagg gcgatgccac ctacggcaag
     1441 ctgaccctga agttcatctg caccaccggc aagctgcccg tgccctggcc caccctcgtg
     1501 accaccctga cctacggcgt gcagtgcttc agccgctacc ccgaccacat gaagcagcac
     1561 gacttcttca agtccgccat gcccgaaggc tacgtccagg agcgcaccat cttcttcaag
     1621 gacgacggca actacaagac ccgcgccgag gtgaagttcg agggcgacac cctggtgaac
     1681 cgcatcgagc tgaagggcat cgacttcaag gaggacggca acatcctggg gcacaagctg
     1741 gagtacaact acaacagcca caacgtctat atcatggccg acaagcagaa gaacggcatc
     1801 aaggtgaact tcaagatccg ccacaacatc gaggacggca gcgtgcagct cgccgaccac
     1861 taccagcaga acacccccat cggcgacggc cccgtgctgc tgcccgacaa ccactacctg
     1921 agcacccagt ccgccctgag caaagacccc aacgagaagc gcgatcacat ggtcctgctg
     1981 gagttcgtga ccgccgccgg gatcactctc ggcatggacg agctgtacaa gtccggtaca
     2041 gctctcgacg gagaaagctc aggctctggc tcagagtctg actccatggc caatttactg
     2101 accgtacacc aaaatttgcc tgcattaccg gtcgatgcaa cgagtgatga ggttcgcaag
     2161 aacctgatgg acatgttcag ggatcgccag gcgttttctg agcatacctg gaaaatgctt
     2221 ctgtccgttt gccggtcgtg ggcggcatgg tgcaagttga ataaccggaa atggtttccc
     2281 gcagaacctg aagatgttcg cgattatctt ctatatcttc aggcgcgcgg tctggcagta
     2341 aaaactatcc agcaacattt gggccagcta aacatgcttc atcgtcggtc cgggctgcca
     2401 cgaccaagtg acagcaatgc tgtttcactg gttatgcggc ggatccgaaa agaaaacgtt
     2461 gatgccggtg aacgtgcaaa acaggctcta gcgttcgaac gcactgattt cgaccaggtt
     2521 cgttcactca tggaaaatag cgatcgctgc caggatatac gtaatctggc atttctgggg
     2581 attgcttata acaccctgtt acgtatagcc gaaattgcca ggatcagggt taaagatatc
     2641 tcacgtactg acggtgggag aatgttaatc catattggca gaacgaaaac gctggttagc
     2701 accgcaggtg tagagaaggc acttagcctg ggggtaacta aactggtcga gcgatggatt
     2761 tccgtctctg gtgtagctga tgatccgaat aactacctgt tttgccgggt cagaaaaaat
     2821 ggtgttgccg cgccatctgc caccagccag ctatcaactc gcgccctgga agggattttt
     2881 gaagcaactc atcgattgat ttacggcgct aaggatgact ctggtcagag atacctggcc
     2941 tggtctggac acagtgcccg tgtcggagcc gcgcgagata tggcccgcgc tggagtttca
     3001 ataccggaga tcatgcaagc tggtggctgg accaatgtaa atattgtcat gaactatatc
     3061 cgtaacctgg atagtgaaac aggggcaatg gtgcgcctgc tggaagatgg cgattagcca
     3121 ttaacgcgta aatgattgct ataattattt gatatttatg gtgacatatg agaaaggatt
     3181 tcaacatcga cggaaaatat gtagtgctgt ctgtaagcac taatattcag tcgccagccg
     3241 tcattgtcac tgtaaagctg agcgatagaa tgcctgatat tgactcaata tccgttgcgt
     3301 ttcctgtcaa aagtatgcgt agtgctgaac atttcgtgat gaatgccacc gaggaagaag
     3361 cacggcgcgg ttttgctaaa gtgatgtctg agtttggcga actcttgggt aaggttggaa
     3421 ttgtcgaggg atctcggaat ggaccccaac tgctcctgct ccaccggtaa gactcccgat
     3481 ccttggtctt tagaatacca agttgggacc gcagagcgga atccccgagt tgtagaggct
     3541 tggcgggaat aggcaccttt agttggcgat tcattccggt tctttctaga atccgctctt
     3601 gcaaaagcct tcattagtta cgagtattgt cgaacgggtc ctttggcggg gttggggcta
     3661 ggatttagac gcgcaaatgt ccggttcctg atcacccagt tagtggggac atctgggttg
     3721 agtcccaggc attactaaac ttactgtgaa ttgcttgaat taagaaagag gtgaaggacc
     3781 tttatgtctt gggactcaaa gacataatcc ctgacttaac ctgtgaggag aaaagtgggg
     3841 ctaggctccc tgcagctccg aggaggactt agtgaactga gccgggactc gtggtgttgg
     3901 ccactgctgt aatgctgcct ccctcatgct gtcttctttc tcctccccag gcggctcctg
     3961 cacttgcacc agctcctgcg cctgcaagaa ctgcaagtgc acctcctgca agaagagtga
     4021 gttgggacac cttgggtggc ggctaaggct aggggcgggg aactcctaca aaactggctc
     4081 tgagaaatgt cctttgcttc ccggaggcca ttgtattgtc tcggggacag aactatacag
     4141 agaactattt aaaaaaaccg aggtcttctc tgttggggac aggaagcaga ggtcttcagc
     4201 caggctgacc tcttcctcct cctttctagg ctgctgctcc tgctgtcccg tgggctgctc
     4261 caaatgtgcc cagggctgtg tctgcaaagg cgccgcggac aagtgcacgt gctgtgcctg
     4321 atgtgacgaa cagcgctgcc accacgtgta aatagtatcg gaccaaccca gcgtcttcct
     4381 atacagttcc accctgttta ctaaaccccc gttttctacc gagtacgtga ataataaaag
     4441 cctgtttgag tctaactctg gttttcttgg tgtggtttgg caataagaaa ctggggtgac
     4501 ttgatagtct ggggatctgg ttttggaccc cctcgtgcct ttacctcccg cctctggccc
     4561 tcacagaggg gtaatgtctt tgggtaaagc caagctatcc cataagcttc ctcatggaaa
     4621 acagctgtgt gcctggtcag gtcttgtgtt aggggaaggg agggagggag ggtggtcctg
     4681 gaagacttga tgtcccatcc tggaagcaca gaaaatctta tcatttatat tctctttgga
     4741 ttctggtttc ctctcacatt gttgggtata aggaggtggg tatgaaaatt aacatagggt
     4801 gtctcgtatg ctaggcaggg actaacactg aactagattc cagaggggct ttcttggctg
     4861 ttttgacaca agttgcttac cttgtcctag aactcattct gcagtccagt ctgaccttga
     4921 gtttttgatt ctcctgcctc agcttcccaa gtacctggga ttacaggagt gactgtgtca
     4981 ccaggccatc tcctattcag ccaccaaatg cagacactat tgcatatgcc atcaagattt
     5041 tgctgaaagg accctgataa agctgtctcc tgtgaggctt tgccagtgcc cagcaaacac
     5101 agaagtggat gctcacagtc agctattgga tggaacacag ggcccccaat ggaggagcta
     5161 gagaaagtac ccaaggagct aaagggatct gcaaccctat aggtggaaca gcaatctgaa
     5221 ctaaccagta ccccctgaac tcgtgtatct agcttcgtat gtagcagaag atggcctagt
     5281 tggccatcac tgggaagaga ggacccttgg tctagcaaac tatatgcccc agttcagggg
     5341 aaagccaggg ccaagaagtg ggagtgagtg ggtaggggag cggggtgggg gtaggggtgg
     5401 ggtatagggg acttttggga tagcatttga aatataaacg aagaaaatat ccaaaaaaaa
     5461 aaagatggaa aaaaaagtga taaagagcgt gggtcctgcc tgtaatcaag atatttgaga
     5521 tgtagcagca ggaaggccag gaattggtcg agggtaccga gctcgaattc gtaatcatgt
     5581 catagctgtt tcctgtgtga aattgttatc cgctcacaat tccacacaac atacgagccg
     5641 gaagcataaa gtgtaaagcc tggggtgcct aatgagtgag ctaactcaca ttaattgcgt
     5701 tgcgctcact gcccgctttc cagtcgggaa acctgtcgtg ccagctgcat taatgaatcg
     5761 gccaacgcgc ggggagaggc ggtttgcgta ttgggcgctc ttccgcttcc tcgctcactg
     5821 actcgctgcg ctcggtcgtt cggctgcggc gagcggtatc agctcactca aaggcggtaa
     5881 tacggttatc cacagaatca ggggataacg caggaaagaa catgtgagca aaaggccagc
     5941 aaaaggccag gaaccgtaaa aaggccgcgt tgctggcgtt tttccatagg ctccgccccc
     6001 ctgacgagca tcacaaaaat cgacgctcaa gtcagaggtg gcgaaacccg acaggactat
     6061 aaagatacca ggcgtttccc cctggaagct ccctcgtgcg ctctcctgtt ccgaccctgc
     6121 cgcttaccgg atacctgtcc gcctttctcc cttcgggaag cgtggcgctt tctcatagct
     6181 cacgctgtag gtatctcagt tcggtgtagg tcgttcgctc caagctgggc tgtgtgcacg
     6241 aaccccccgt tcagcccgac cgctgcgcct tatccggtaa ctatcgtctt gagtccaacc
     6301 cggtaagaca cgacttatcg ccactggcag cagccactgg taacaggatt agcagagcga
     6361 ggtatgtagg cggtgctaca gagttcttga agtggtggcc taactacggc tacactagaa
     6421 gaacagtatt tggtatctgc gctctgctga agccagttac cttcggaaaa agagttggta
     6481 gctcttgatc cggcaaacaa accaccgctg gtagcggtgg tttttttgtt tgcaagcagc
     6541 agattacgcg cagaaaaaaa ggatctcaag aagatccttt gatcttttct acggggtctg
     6601 acgctcagtg gaacgaaaac tcacgttaag ggattttggt catgagatta tcaaaaagga
     6661 tcttcaccta gatcctttta aattaaaaat gaagttttaa atcaatctaa agtatatatg
     6721 agtaaacttg gtctgacagt taccaatgct taatcagtga ggcacctatc tcagcgatct
     6781 gtctatttcg ttcatccata gttgcctgac tccccgtcgt gtagataact acgatacggg
     6841 agggcttacc atctggcccc agtgctgcaa tgataccgcg agacccacgc tcaccggctc
     6901 cagatttatc agcaataaac cagccagccg gaagggccga gcgcagaagt ggtcctgcaa
     6961 ctttatccgc ctccatccag tctattaatt gttgccggga agctagagta agtagttcgc
     7021 cagttaatag tttgcgcaac gttgttgcca ttgctacagg catcgtggtg tcacgctcgt
     7081 cgtttggtat ggcttcattc agctccggtt cccaacgatc aaggcgagtt acatgatccc
     7141 ccatgttgtg caaaaaagcg gttagctcct tcggtcctcc gatcgttgtc agaagtaagt
     7201 tggccgcagt gttatcactc atggttatgg cagcactgca taattctctt actgtcatgc
     7261 catccgtaag atgcttttct gtgactggtg agtactcaac caagtcattc tgagaatagt
     7321 gtatgcggcg accgagttgc tcttgcccgg cgtcaatacg ggataatacc gcgccacata
     7381 gcagaacttt aaaagtgctc atcattggaa aacgttcttc ggggcgaaaa ctctcaagga
     7441 tcttaccgct gttgagatcc agttcgatgt aacccactcg tgcacccaac tgatcttcag
     7501 catcttttac tttcaccagc gtttctgggt gagcaaaaac aggaaggcaa aatgccgcaa
     7561 aaaagggaat aagggcgaca cggaaatgtt gaatactcat actcttcctt tttcaatatt
     7621 attgaagcat ttatcagggt tattgtctca tgagcggata catatttgaa tgtatttaga
     7681 aaaataaaca aataggggtt ccgcgcacat ttccccgaaa agtgccacct gacgtctaag
     7741 aaaccattat tatcatgaca ttaacctata aaaataggcg tatcacgagg ccctttcgtc
     7801 tcgcgcgttt cggtgatgac ggtgaaaacc tctgacacat gcagctcccg gagacggtca
     7861 cagcttgtct gtaagcggat gccgggagca gacaagcccg tcagggcgcg tcagcgggtg
     7921 ttggcgggtg tcggggctgg cttaactatg cggcatcaga gcagattgta ctgagagtgc
     7981 accatatgcg gtgtgaaata ccgcacagat gcgtaaggag aaaataccgc atcaggcgcc
     8041 attcgccatt caggctgcgc aactgttggg aagggcgatc ggtgcgggcc tcttcgctat
     8101 tacgccagct ggcgaaaggg ggatgtgctg caaggcgatt aagttgggta acgccagggt
     8161 tttcccagtc acgacgtt
//
