LOCUS       sequence-414368-        5394 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expression of the Cre recombinase linked to a puro selectable 
            marker.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5394)
  AUTHORS   Philip R, Sharma A, Matellan L, Erpf AC, Hsu WH, Tkach JM, Wyatt 
            HDM, Pelletier L
  TITLE     qTAG: an adaptable plasmid scaffold for CRISPR-based endogenous 
            tagging.
  JOURNAL   EMBO J. 2024 Dec 12. doi: 10.1038/s44318-024-00337-5.
  PUBMED    39668248
REFERENCE   2  (bases 1 to 5394)
  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: "EMBO J. 
            2024 Dec 12. doi: 10.1038/s44318-024-00337-5."
COMMENT     Sequence Label: pEF1-alpha-Cre-2A-Puro
FEATURES             Location/Qualifiers
     source          1..5394
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        79..290
                     /label=EF-1-alpha core promoter
                     /note="core promoter for human elongation factor 
                     EF-1-alpha"
     intron          291..1229
                     /label=EF-1-alpha intron A
                     /note="intron upstream of the start codon of human 
                     EF-1-alpha"
     primer_bind     1186..1206
                     /label=EF1a-F
                     /note="Human elongation factor-1a promoter, forward primer"
     CDS             1260..2288
                     /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="MSNLLTVHQNLPALPVDATSDEVRKNLMDMFRDRQAFSEHTWKML
                     LSVCRSWAAWCKLNNRKWFPAEPEDVRDYLLYLQARGLAVKTIQQHLGQLNMLHRRSGL
                     PRPSDSNAVSLVMRRIRKENVDAGERAKQALAFERTDFDQVRSLMENSDRCQDIRNLAF
                     LGIAYNTLLRIAEIARIRVKDISRTDGGRMLIHIGRTKTLVSTAGVEKALSLGVTKLVE
                     RWISVSGVADDPNNYLFCRVRKNGVAAPSATSQLSTRALEGIFEATHRLIYGAKDDSGQ
                     RYLAWSGHSARVGAARDMARAGVSIPEIMQAGGWTNVNIVMNYIRNLDSETGAMVRLLE
                     DGD"
     CDS             2304..2360
                     /codon_start=1
                     /product="2A peptide from porcine teschovirus-1 
                     polyprotein"
                     /label=P2A
                     /note="Eukaryotic ribosomes fail to insert a peptide bond 
                     between the Gly and Pro residues, yielding separate 
                     polypeptides."
                     /translation="ATNFSLLKQAGDVEENPGP"
     CDS             2376..2975
                     /codon_start=1
                     /gene="pac from Streptomyces alboniger"
                     /product="puromycin N-acetyltransferase"
                     /label=PuroR
                     /note="confers resistance to puromycin"
                     /translation="MTEYKPTVRLATRDDVPRAVRTLAAAFADYPATRHTVDPDRHIER
                     VTELQELFLTRVGLDIGKVWVADDGAAVAVWTTPESVEAGAVFAEIGPRMAELSGSRLA
                     AQQQMEGLLAPHRPKEPAWFLATVGVSPDHQGKGLGSAVVLPGVEAAERAGVPAFLETS
                     APRNLPFYERLGFTVTADVEVPEGPRTWCMTRKPGA"
     primer_bind     complement(2376..2395)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     2872..2892
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     polyA_signal    2997..3118
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3034..3053)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3088..3107
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(3186..3774)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3266..3285)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(3945..4736,4737..4805))
                     /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     4568..4587
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(4806..4910)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      complement(4936..5391)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(5073..5094)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     5285..5304
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
ORIGIN
        1 attcgccatt caggctgcgc aactgttggg aagggcgatc ggtgcgggcc tcttcgctat
       61 tacgccagtc gattcagtgg gcagagcgca catcgcccac agtccccgag aagttggggg
      121 gaggggtcgg caattgaacc ggtgcctaga gaaggtggcg cggggtaaac tgggaaagtg
      181 atgtcgtgta ctggctccgc ctttttcccg agggtggggg agaaccgtat ataagtgcag
      241 tagtcgccgt gaacgttctt tttcgcaacg ggtttgccgc cagaacacag gtaagtgccg
      301 tgtgtggttc ccgcgggcct ggcctcttta cgggttatgg cccttgcgtg ccttgaatta
      361 cttccacctg gctgcagtac gtgattcttg atcccgagct tcgggttgga agtgggtggg
      421 agagttcgag gccttgcgct taaggagccc cttcgcctcg tgcttgagtt gaggcctggc
      481 ctgggcgctg gggccgccgc gtgcgaatct ggtggcacct tcgcgcctgt ctcgctgctt
      541 tcgataagtc tctagccatt taaaattttt gatgacctgc tgcgacgctt tttttctggc
      601 aagatagtct tgtaaatgcg ggccaagatc tgcacactgg tatttcggtt tttggggccg
      661 cgggcggcga cggggcccgt gcgtcccagc gcacatgttc ggcgaggcgg ggcctgcgag
      721 cgcggccacc gagaatcgga cgggggtagt ctcaagctgg ccggcctgct ctggtgcctg
      781 gcctcgcgcc gccgtgtatc gccccgccct gggcggcaag gctggcccgg tcggcaccag
      841 ttgcgtgagc ggaaagatgg ccgcttcccg gccctgctgc agggagctca aaatggagga
      901 cgcggcgctc gggagagcgg gcgggtgagt cacccacaca aaggaaaagg gcctttccgt
      961 cctcagccgt cgcttcatgt gactccacgg agtaccgggc gccgtccagg cacctcgatt
     1021 agttctcgag cttttggagt acgtcgtctt taggttgggg ggaggggttt tatgcgatgg
     1081 agtttcccca cactgagtgg gtggagactg aagttaggcc agcttggcac ttgatgtaat
     1141 tctccttgga atttgccctt tttgagtttg gatcttggtt cattctcaag cctcagacag
     1201 tggttcaaag tttttttctt ccatttcagg tgtcgtgagg atccgaattc acgcgtctaa
     1261 tgagcaattt actgaccgta caccaaaatt tgcctgcatt accggtcgat gcaacgagtg
     1321 atgaggttcg caagaacctg atggacatgt tcagggatcg ccaggcgttt tctgagcata
     1381 cctggaaaat gcttctgtcc gtttgccggt cgtgggcggc atggtgcaag ttgaataacc
     1441 ggaaatggtt tcccgcagaa cctgaagatg ttcgcgatta tcttctatat cttcaggcgc
     1501 gcggtctggc agtaaaaact atccagcaac atttgggcca gctaaacatg cttcatcgtc
     1561 ggtccgggct gccacgacca agtgacagca atgctgtttc actggttatg cggcggatcc
     1621 gaaaagaaaa cgttgatgcc ggtgaacgtg caaaacaggc tctagcgttc gaacgcactg
     1681 atttcgacca ggttcgttca ctcatggaaa atagcgatcg ctgccaggat atacgtaatc
     1741 tggcatttct ggggattgct tataacaccc tgttacgtat agccgaaatt gccaggatca
     1801 gggttaaaga tatctcacgt actgacggtg ggagaatgtt aatccatatt ggcagaacga
     1861 aaacgctggt tagcaccgca ggtgtagaga aggcacttag cctgggggta actaaactgg
     1921 tcgagcgatg gatttccgtc tctggtgtag ctgatgatcc gaataactac ctgttttgcc
     1981 gggtcagaaa aaatggtgtt gccgcgccat ctgccaccag ccagctatca actcgcgccc
     2041 tggaagggat ttttgaagca actcatcgat tgatttacgg cgctaaggat gactctggtc
     2101 agagatacct ggcctggtct ggacacagtg cccgtgtcgg agccgcgcga gatatggccc
     2161 gcgctggagt ttcaataccg gagatcatgc aagctggtgg ctggaccaat gtaaatattg
     2221 tcatgaacta tatccgtaac ctggatagtg aaacaggggc aatggtgcgc ctgctggaag
     2281 atggcgaccc cgggggatcc ggagctacta acttcagcct gctgaagcag gccggagacg
     2341 tggaggagaa ccctggacct actagaggaa gcggaatgac cgagtacaag cccacggtgc
     2401 gcctcgccac ccgcgacgac gtccccaggg ccgtacgcac cctcgccgcc gcgttcgccg
     2461 actaccccgc cacgcgccac accgtcgatc cggaccgcca catcgagcgg gtcaccgagc
     2521 tgcaagaact cttcctcacg cgcgtcgggc tcgacatcgg caaggtgtgg gtcgcggacg
     2581 acggcgccgc ggtggcggtc tggaccacgc cggagagcgt cgaagcgggg gcggtgttcg
     2641 ccgagatcgg cccgcgcatg gccgagttga gcggttcccg gctggccgcg cagcaacaga
     2701 tggaaggcct cctggcgccg caccggccca aggagcccgc gtggttcctg gccaccgtcg
     2761 gcgtctcgcc cgaccaccag ggcaagggtc tgggcagcgc cgtcgtgctc cccggagtgg
     2821 aggcggccga gcgcgccggg gtgcccgcct tcctggagac ctccgcgccc cgcaacctcc
     2881 ccttctacga gcggctcggc ttcaccgtca ccgccgacgt cgaggtgccc gaaggaccgc
     2941 gcacctggtg catgacccgc aagcccggtg cctgaggcgc gcctattatg gtacctaact
     3001 tgtttattgc agcttataat ggttacaaat aaagcaatag catcacaaat ttcacaaata
     3061 aagcattttt ttcactgcat tctagttgtg gtttgtccaa actcatcaat gtatcttaca
     3121 tatgacatgt gagcaaaagg ccagcaaaag gccaggaacc gtaaaaaggc cgcgttgctg
     3181 gcgtttttcc ataggctccg cccccctgac gagcatcaca aaaatcgacg ctcaagtcag
     3241 aggtggcgaa acccgacagg actataaaga taccaggcgt ttccccctgg aagctccctc
     3301 gtgcgctctc ctgttccgac cctgccgctt accggatacc tgtccgcctt tctcccttcg
     3361 ggaagcgtgg cgctttctca tagctcacgc tgtaggtatc tcagttcggt gtaggtcgtt
     3421 cgctccaagc tgggctgtgt gcacgaaccc cccgttcagc ccgaccgctg cgccttatcc
     3481 ggtaactatc gtcttgagtc caacccggta agacacgact tatcgccact ggcagcagcc
     3541 actggtaaca ggattagcag agcgaggtat gtaggcggtg ctacagagtt cttgaagtgg
     3601 tggcctaact acggctacac tagaagaaca gtatttggta tctgcgctct gctgaagcca
     3661 gttaccttcg gaaaaagagt tggtagctct tgatccggca aacaaaccac cgctggtagc
     3721 ggtggttttt ttgtttgcaa gcagcagatt acgcgcagaa aaaaaggatc tcaagaagat
     3781 cctttgatct tttctacggg gtctgacgct cagtggaacg aaaactcacg ttaagggatt
     3841 ttggtcatga gattatcaaa aaggatcttc acctagatcc ttttaaatta aaaatgaagt
     3901 tttaaatcaa tctaaagtat atatgagtaa acttggtctg acagttacca atgcttaatc
     3961 agtgaggcac ctatctcagc gatctgtcta tttcgttcat ccatagttgc ctgactcccc
     4021 gtcgtgtaga taactacgat acgggagggc ttaccatctg gccccagtgc tgcaatgata
     4081 ccgcgagacc cacgctcacc ggctccagat ttatcagcaa taaaccagcc agccggaagg
     4141 gccgagcgca gaagtggtcc tgcaacttta tccgcctcca tccagtctat taattgttgc
     4201 cgggaagcta gagtaagtag ttcgccagtt aatagtttgc gcaacgttgt tgccattgct
     4261 acaggcatcg tggtgtcacg ctcgtcgttt ggtatggctt cattcagctc cggttcccaa
     4321 cgatcaaggc gagttacatg atcccccatg ttgtgcaaaa aagcggttag ctccttcggt
     4381 cctccgatcg ttgtcagaag taagttggcc gcagtgttat cactcatggt tatggcagca
     4441 ctgcataatt ctcttactgt catgccatcc gtaagatgct tttctgtgac tggtgagtac
     4501 tcaaccaagt cattctgaga atagtgtatg cggcgaccga gttgctcttg cccggcgtca
     4561 atacgggata ataccgcgcc acatagcaga actttaaaag tgctcatcat tggaaaacgt
     4621 tcttcggggc gaaaactctc aaggatctta ccgctgttga gatccagttc gatgtaaccc
     4681 actcgtgcac ccaactgatc ttcagcatct tttactttca ccagcgtttc tgggtgagca
     4741 aaaacaggaa ggcaaaatgc cgcaaaaaag ggaataaggg cgacacggaa atgttgaata
     4801 ctcatactct tcctttttca atattattga agcatttatc agggttattg tctcatgagc
     4861 ggatacatat ttgaatgtat ttagaaaaat aaacaaatag gggttccgcg cacatttccc
     4921 cgaaaagtgc cacctaaatt gtaagcgtta atattttgtt aaaattcgcg ttaaattttt
     4981 gttaaatcag ctcatttttt aaccaatagg ccgaaatcgg caaaatccct tataaatcaa
     5041 aagaatagac cgagataggg ttgagtgttg ttccagtttg gaacaagagt ccactattaa
     5101 agaacgtgga ctccaacgtc aaagggcgaa aaaccgtcta tcagggcgat ggcccactac
     5161 gtgaaccatc accctaatca agttttttgg ggtcgaggtg ccgtaaagca ctaaatcgga
     5221 accctaaagg gagcccccga tttagagctt gacggggaaa gccggcgaac gtggcgagaa
     5281 aggaagggaa gaaagcgaaa ggagcgggcg ctagggcgct ggcaagtgta gcggtcacgc
     5341 tgcgcgtaac caccacaccc gccgcgctta atgcgccgct acagggcgcg tccc
//
