LOCUS       sequence-325851-        7158 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Introduce Cre-inducible (EF1a promoter followed by lox-stop-lox) GFP
            cDNA into (ES) cells by recombination-mediated cassette exchange.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7158)
  TITLE     Karreth lab plasmids
REFERENCE   2  (bases 1 to 7158)
  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"
COMMENT     Sequence Label: cEF1a-LSL-GFP
FEATURES             Location/Qualifiers
     source          1..7158
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        48..547
                     /label=PGK promoter
                     /note="mouse phosphoglycerate kinase 1 promoter"
     primer_bind     complement(83..106)
                     /label=MSCV-rev
                     /note="Murine stem cell virus, reverse primer"
     primer_bind     480..499
                     /label=mPGK-F
                     /note="Mouse PGK promoter, forward primer"
     protein_bind    593..640
                     /label=FRT
                     /bound_moiety="FLP recombinase from the Saccharomyces 
                     cerevisiae 2u plasmid"
                     /note="FLP-mediated recombination occurs in the 8-bp core 
                     sequence TCTAGAAA (Turan and Bode, 2011)."
     misc_feature    join(673..738,739..776)
                     /label=Ad SA
                     /note="adenovirus major late transcript splice acceptor 
                     (Friedrich and Soriano, 1991)"
     polyA_signal    945..1079
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     polyA_signal    1199..1333
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     promoter        1349..1560
                     /label=EF-1-alpha core promoter
                     /note="core promoter for human elongation factor 
                     EF-1-alpha"
     protein_bind    complement(1591..1624)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     intron          2162..2227
                     /label=small t intron
                     /note="SV40 (simian virus 40) small t antigen intron"
     CDS             2357..2377
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     polyA_signal    2802..2936
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     protein_bind    complement(2978..3011)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     regulatory      3057..3066
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(3063..3065,3066..3068,3069..3782)
                     /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(3063..3065,3066..3068,3069..3779)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(3108..3129)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(3369..3388)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     3716..3737
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     intron          3825..4397
                     /label=beta-globin intron
                     /note="intron from rabbit beta-globin gene"
     rep_origin      complement(4725..5313)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4805..4824)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5484..6275,6276..6344))
                     /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     6107..6126
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6345..6449)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      complement(6475..6930)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(6612..6633)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     6824..6843
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     7057..7079
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     7071..7088
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     7072..7088
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     7095..7114
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        7095..7113
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     join(7146..7158,1..4)
                     /label=SK primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     join(7146..7158,1..4)
                     /label=pBluescriptSK
                     /note="For pBluescript vector"
ORIGIN
        1 gatcccccgg gctgcaggaa ttaattcgtc gacctcgaaa ttctaccggg taggggaggc
       61 gcttttccca aggcagtctg gagcatgcgc tttagcagcc ccgctgggca cttggcgcta
      121 cacaagtggc ctctggcctc gcacacattc cacatccacc ggtaggcgcc aaccggctcc
      181 gttctttggt ggccccttcg cgccaccttc tactcctccc ctagtcagga agttcccccc
      241 cgccccgcag ctcgcgtcgt gcaggacgtg acaaatggaa gtagcacgtc tcactagtct
      301 cgtgcagatg gacagcaccg ctgagcaatg gaagcgggta ggcctttggg gcagcggcca
      361 atagcagctt tgctccttcg ctttctgggc tcagaggctg ggaaggggtg ggtccggggg
      421 cgggctcagg ggcgggctca ggggcggggc gggcgcccga aggtcctccg gaggcccggc
      481 attctgcacg cttcaaaagc gcacgtctgc cgcgctgttc tcctcttcct catctccggg
      541 cctttcgacc tgcatccatc tagatctcga tcgagcagct gaagcttatg gagaagttac
      601 tattccgaag ttcctattct ctagaaagta taggaacttc gatccccctc gatcccccgg
      661 gctgcagatc tgtagggcgc agtagtccag ggtttccttg atgatgtcat acttatcctg
      721 tccctttttt ttccacagct cgcggttgag gacaaactct tcgcggtctt tccagtgggg
      781 atcgacggta tcgataagct tgcagatctg cgactctaga ggatctgcga ctctagagga
      841 tcataatcag ccataccaca tttgtagagg ttttacttgc tttaaaaaac ctcccacacc
      901 tccccctgaa cctgaaacat aaaatgaatg caattgttgt tgttaacttg tttattgcag
      961 cttataatgg ttacaaataa agcaatagca tcacaaattt cacaaataaa gcattttttt
     1021 cactgcattc tagttgtggt ttgtccaaac tcatcaatgt atcttatcat gtctggatct
     1081 gcgactctag aggatcataa tcagccatac cacatttgta gaggttttac ttgctttaaa
     1141 aaacctccca cacctccccc tgaacctgaa acataaaatg aatgcaattg ttgttgttaa
     1201 cttgtttatt gcagcttata atggttacaa ataaagcaat agcatcacaa atttcacaaa
     1261 taaagcattt ttttcactgc attctagttg tggtttgtcc aaactcatca atgtatctta
     1321 tcatgtctgg atcgagttta ccatgcatgg gcagagcgca catcgcccac agtccccgag
     1381 aagttggggg gaggggtcgg caattgatcc ggtgcctaga gaaggtggcg cggggtaaac
     1441 tgggaaagtg atgtcgtgta ctggctccgc ctttttcccg agggtggggg agaaccgtat
     1501 ataagtgcag tagtcgccgt gaacgttctt tttcgcaacg ggtttgccgc cagaacacag
     1561 tccacgctcg agacctcttc gagggaccta ataacttcgt atagcataca ttatacgaag
     1621 ttatattaag ggttccggat cctcggggac accaaatatg gcgatctcgg ccttttcgtt
     1681 tcttggagct gggacatgtt tgccatcgat ccatctacca ccagaacggc cgttagatct
     1741 gctgccaccg ttgtttccac cgaagaaacc accgttgccg taaccaccac gacggttgtt
     1801 gctaaagaag ctgccaccgc cacggccacc gttgtagccg ccgttgttgt tattgtagtt
     1861 gctactgtta tttctggcac ttcttggttt tcctcttaag tgaggaggaa cataaccatt
     1921 ctcgttgttg tcgttgatgc ttaaattttg cacttgttcg ctcagttcag ccataatatg
     1981 aaatgctttt cttgttgttc ttacggaata ccacttgcca cctatcacca caactaactt
     2041 tttcccgttc ctccatctct tttatatttt ttttctcgat cgagggatct ttgtgaagga
     2101 accttacttc tgtggtgtga cataattgga caaactacct acagagattt aaagctctaa
     2161 ggtaaatata aaatttttaa gtgtataatg tgttaaacta ctgattctaa ttgtttgtgt
     2221 attttagatt ccaacctatg gaactgatga atgggagcag tggtggaatg cctttaatga
     2281 ggaaaacctg ttttgctcag aagaaatgcc atctagtgat gatgaggcta ctgctgactc
     2341 tcaacattct actcctccaa aaaagaagag aaaggtagaa gaccccaagg actttccttc
     2401 agaattgcta agttttttga gtcatgctgt gtttagtaat agaactcttg cttgctttgc
     2461 tatttacacc acaaaggaaa aagctgcact gctatacaag aaaattatgg aaaaatattc
     2521 tgtaaccttt ataagtaggc ataacagtta taatcataac atactgtttt ttcttactcc
     2581 acacaggcat agagtgtctg ctattaataa ctatgctcaa aaattgtgta cctttagctt
     2641 tttaatttgt aaaggggtta ataaggaata tttgatgtat agtgccttga ctagagatca
     2701 taatcagcca taccacattt gtagaggttt tacttgcttt aaaaaacctc ccacacctcc
     2761 ccctgaacct gaaacataaa atgaatgcaa ttgttgttgt taacttgttt attgcagctt
     2821 ataatggtta caaataaagc aatagcatca caaatttcac aaataaagca tttttttcac
     2881 tgcattctag ttgtggtttg tccaaactca tcaatgtatc ttatcatgtc tggatctgac
     2941 atggtaagta agcttgggct gcaggtcgag ggacctaata acttcgtata gcatacatta
     3001 tacgaagtta tattaagggt tccggatcct aagcttgatg gaattctcga gatatcgcca
     3061 ccatggtgag caagggcgag gagctgttca ccggggtggt gcccatcctg gtcgagctgg
     3121 acggcgacgt aaacggccac aagttcagcg tgtccggcga gggcgagggc gatgccacct
     3181 acggcaagct gaccctgaag ttcatctgca ccaccggcaa gctgcccgtg ccctggccca
     3241 ccctcgtgac caccctgacc tacggcgtgc agtgcttcag ccgctacccc gaccacatga
     3301 agcagcacga cttcttcaag tccgccatgc ccgaaggcta cgtccaggag cgcaccatct
     3361 tcttcaagga cgacggcaac tacaagaccc gcgccgaggt gaagttcgag ggcgacaccc
     3421 tggtgaaccg catcgagctg aagggcatcg acttcaagga ggacggcaac atcctggggc
     3481 acaagctgga gtacaactac aacagccaca acgtctatat catggccgac aagcagaaga
     3541 acggcatcaa ggtgaacttc aagatccgcc acaacatcga ggacggcagc gtgcagctcg
     3601 ccgaccacta ccagcagaac acccccatcg gcgacggccc cgtgctgctg cccgacaacc
     3661 actacctgag cacccagtcc gccctgagca aagaccccaa cgagaagcgc gatcacatgg
     3721 tcctgctgga gttcgtgacc gccgccggga tcactctcgg catggacgag ctgtacaagt
     3781 aactcgagtt catgaattcg cagtcgagcg ttgagaactt cagggtgagt ttggggaccc
     3841 ttgattgttc tttctttttc gctattgtaa aattcatgtt atatggaggg ggcaaagttt
     3901 tcagggtgtt gtttagaatg ggaagatgtc ccttgtatca ccatggaccc tcatgataat
     3961 tttgtttctt tcactttcta ctctgttgac aaccattgtc tcctcttatt ttcttttcat
     4021 tttctgtaac tttttcgtta aactttagct tgcatttgta acgaattttt aaattcactt
     4081 ttgtttattt gtcagattgt aagtactttc tctaatcact tttttttcaa ggcaatcagg
     4141 gtatattata ttgtacttca gcacagtttt agagaacaat tgttataatt aaatgataag
     4201 gtagaatatt tctgcatata aattctggct ggcgtggaaa tattcttatt ggtagaaaca
     4261 actacaccct ggtcatcatc ctgcctttct ctttatggtt acaatgatat acactgtttg
     4321 agatgaggat aaaatactct gagtccaaac cgggcccctc tgctaaccat gttcatgcct
     4381 tcttctcttt cctacagctc ctgggcaacg tgctggttgt tgtgctgtct catcattttg
     4441 gcaaaggatt cactcctcag gtgcaggctg cctatcagaa ggtggtggct ggtgtggcca
     4501 atgccctggc tcacaaatac cactgagatc gttttccctc tgccaaaaat tatggggaca
     4561 tcatgaagcc ccttgagcat ctgacttctg gctaataaag gaaatttatt ttcattgcaa
     4621 tagtgtgttg gaatttctcg atcgctagca tcctaggcgc gccacatgtg agcaaaaggc
     4681 cagcaaaagg ccaggaaccg taaaaaggcc gcgttgctgg cgtttttcca taggctccgc
     4741 ccccctgacg agcatcacaa aaatcgacgc tcaagtcaga ggtggcgaaa cccgacagga
     4801 ctataaagat accaggcgtt tccccctgga agctccctcg tgcgctctcc tgttccgacc
     4861 ctgccgctta ccggatacct gtccgccttt ctcccttcgg gaagcgtggc gctttctcat
     4921 agctcacgct gtaggtatct cagttcggtg taggtcgttc gctccaagct gggctgtgtg
     4981 cacgaacccc ccgttcagcc cgaccgctgc gccttatccg gtaactatcg tcttgagtcc
     5041 aacccggtaa gacacgactt atcgccactg gcagcagcca ctggtaacag gattagcaga
     5101 gcgaggtatg taggcggtgc tacagagttc ttgaagtggt ggcctaacta cggctacact
     5161 agaagaacag tatttggtat ctgcgctctg ctgaagccag ttaccttcgg aaaaagagtt
     5221 ggtagctctt gatccggcaa acaaaccacc gctggtagcg gtggtttttt tgtttgcaag
     5281 cagcagatta cgcgcagaaa aaaaggatct caagaagatc ctttgatctt ttctacgggg
     5341 tctgacgctc agtggaacga aaactcacgt taagggattt tggtcatgag attatcaaaa
     5401 aggatcttca cctagatcct tttaaattaa aaatgaagtt ttaaatcaat ctaaagtata
     5461 tatgagtaaa cttggtctga cagttaccaa tgcttaatca gtgaggcacc tatctcagcg
     5521 atctgtctat ttcgttcatc catagttgcc tgactccccg tcgtgtagat aactacgata
     5581 cgggagggct taccatctgg ccccagtgct gcaatgatac cgcgagaccc acgctcaccg
     5641 gctccagatt tatcagcaat aaaccagcca gccggaaggg ccgagcgcag aagtggtcct
     5701 gcaactttat ccgcctccat ccagtctatt aattgttgcc gggaagctag agtaagtagt
     5761 tcgccagtta atagtttgcg caacgttgtt gccattgcta caggcatcgt ggtgtcacgc
     5821 tcgtcgtttg gtatggcttc attcagctcc ggttcccaac gatcaaggcg agttacatga
     5881 tcccccatgt tgtgcaaaaa agcggttagc tccttcggtc ctccgatcgt tgtcagaagt
     5941 aagttggccg cagtgttatc actcatggtt atggcagcac tgcataattc tcttactgtc
     6001 atgccatccg taagatgctt ttctgtgact ggtgagtact caaccaagtc attctgagaa
     6061 tagtgtatgc ggcgaccgag ttgctcttgc ccggcgtcaa tacgggataa taccgcgcca
     6121 catagcagaa ctttaaaagt gctcatcatt ggaaaacgtt cttcggggcg aaaactctca
     6181 aggatcttac cgctgttgag atccagttcg atgtaaccca ctcgtgcacc caactgatct
     6241 tcagcatctt ttactttcac cagcgtttct gggtgagcaa aaacaggaag gcaaaatgcc
     6301 gcaaaaaagg gaataagggc gacacggaaa tgttgaatac tcatactctt cctttttcaa
     6361 tattattgaa gcatttatca gggttattgt ctcatgagcg gatacatatt tgaatgtatt
     6421 tagaaaaata aacaaatagg ggttccgcgc acatttcccc gaaaagtgcc acctaaattg
     6481 taagcgttaa tattttgtta aaattcgcgt taaatttttg ttaaatcagc tcatttttta
     6541 accaataggc cgaaatcggc aaaatccctt ataaatcaaa agaatagacc gagatagggt
     6601 tgagtgttgt tccagtttgg aacaagagtc cactattaaa gaacgtggac tccaacgtca
     6661 aagggcgaaa aaccgtctat cagggcgatg gcccactacg tgaaccatca ccctaatcaa
     6721 gttttttggg gtcgaggtgc cgtaaagcac taaatcggaa ccctaaaggg agcccccgat
     6781 ttagagcttg acggggaaag ccggcgaacg tggcgagaaa ggaagggaag aaagcgaaag
     6841 gagcgggcgc tagggcgctg gcaagtgtag cggtcacgct gcgcgtaacc accacacccg
     6901 ccgcgcttaa tgcgccgcta cagggcgcgt cccattcgcc attcaggctg cgcaactgtt
     6961 gggaagggcg atcggtgcgg gcctcttcgc tattacgcca gctggcgaaa gggggatgtg
     7021 ctgcaaggcg attaagttgg gtaacgccag ggttttccca gtcacgacgt tgtaaaacga
     7081 cggccagtga attgtaatac gactcactat agggcgaatt ggagctccac cgcggtggcg
     7141 gccgctctag aactagtg
//
