LOCUS       sequence-378317-        4896 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Cp36 Donor with promoter for mcherry and puro expression.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4896)
  AUTHORS   Durrant MG, Fanton A, Tycko J, Hinks M, Chandrasekaran SS, Perry NT,
            Schaepe J, Du PP, Lotfy P, Bassik MC, Bintu L, Bhatt AS, Hsu PD
  TITLE     Systematic discovery of recombinases for efficient integration of 
            large DNA sequences into the human genome.
  JOURNAL   Nat Biotechnol. 2022 Oct 10. pii: 10.1038/s41587-022-01494-w. doi: 
            10.1038/s41587-022-01494-w.
  PUBMED    36217031
REFERENCE   2  (bases 1 to 4896)
  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: "Nat 
            Biotechnol. 2022 Oct 10. pii: 10.1038/s41587-022-01494-w. doi: 
            10.1038/s41587-022-01494-w."
COMMENT     Sequence Label: pAF0362 Ef1a-Cp36_attP-Puro-p2a-mCherry donor
FEATURES             Location/Qualifiers
     source          1..4896
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        32..136
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(137..205,206..997)
                     /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     complement(355..374)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      1168..1756
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     1657..1676
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     1910..1927
                     /label=L4440
                     /note="L4440 vector, forward primer"
     promoter        2101..3279
                     /label=EF-1-alpha promoter
                     /note="strong constitutive promoter for human elongation 
                     factor EF-1-alpha"
     intron          2332..3270
                     /label=EF-1-alpha intron A
                     /note="intron upstream of the start codon of human 
                     EF-1-alpha"
     primer_bind     3227..3247
                     /label=EF1a-F
                     /note="Human elongation factor-1a promoter, forward primer"
     CDS             3287..3883
                     /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(3287..3306)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     3783..3803
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     CDS             3884..3940
                     /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             3947..4657
                     /codon_start=1
                     /product="monomeric derivative of DsRed fluorescent protein
                     (Shaner et al., 2004)"
                     /label=mCherry
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEEDNMAIIKEFMRFKVHMEGSVNGHEFEIEGEGEGRPYEG
                     TQTAKLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNF
                     EDGGVVTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASSERMYPEDGALK
                     GEIKQRLKLKDGGHYDAEVKTTYKAKKPVQLPGAYNVNIKLDITSHNEDYTIVEQYERA
                     EGRHSTGGMDELYK"
     primer_bind     complement(4092..4110)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     4360..4379
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     primer_bind     complement(4682..4699)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    4688..4895
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
ORIGIN
        1 gacgtcaggt ggcacttttc ggggaaatgt gcgcggaacc cctatttgtt tatttttcta
       61 aatacattca aatatgtatc cgctcatgag acaataaccc tgataaatgc ttcaataata
      121 ttgaaaaagg aagagtatga gtattcaaca tttccgtgtc gcccttattc ccttttttgc
      181 ggcattttgc cttcctgttt ttgctcaccc agaaacgctg gtgaaagtaa aagatgctga
      241 agatcagttg ggtgcacgag tgggttacat cgaactggat ctcaacagcg gtaagatcct
      301 tgagagtttt cgccccgaag aacgttttcc aatgatgagc acttttaaag ttctgctatg
      361 tggcgcggta ttatcccgta ttgacgccgg gcaagagcaa ctcggtcgcc gcatacacta
      421 ttctcagaat gacttggttg agtactcacc agtcacagaa aagcatctta cggatggcat
      481 gacagtaaga gaattatgca gtgctgccat aaccatgagt gataacactg cggccaactt
      541 acttctgaca acgatcggag gaccgaagga gctaaccgct tttttgcaca acatggggga
      601 tcatgtaact cgccttgatc gttgggaacc ggagctgaat gaagccatac caaacgacga
      661 gcgtgacacc acgatgcctg tagcaatggc aacaacgttg cgcaaactat taactggcga
      721 actacttact ctagcttccc ggcaacaatt aatagactgg atggaggcgg ataaagttgc
      781 aggaccactt ctgcgctcgg cccttccggc tggctggttt attgctgata aatctggagc
      841 cggtgagcgt gggtctcgcg gtatcattgc agcactgggg ccagatggta agccctcccg
      901 tatcgtagtt atctacacga cggggagtca ggcaactatg gatgaacgaa atagacagat
      961 cgctgagata ggtgcctcac tgattaagca ttggtaactg tcagaccaag tttactcata
     1021 tatactttag attgatttaa aacttcattt ttaatttaaa aggatctagg tgaagatcct
     1081 ttttgataat ctcatgacca aaatccctta acgtgagttt tcgttccact gagcgtcaga
     1141 ccccgtagaa aagatcaaag gatcttcttg agatcctttt tttctgcgcg taatctgctg
     1201 cttgcaaaca aaaaaaccac cgctaccagc ggtggtttgt ttgccggatc aagagctacc
     1261 aactcttttt ccgaaggtaa ctggcttcag cagagcgcag ataccaaata ctgttcttct
     1321 agtgtagccg tagttaggcc accacttcaa gaactctgta gcaccgccta catacctcgc
     1381 tctgctaatc ctgttaccag tggctgctgc cagtggcgat aagtcgtgtc ttaccgggtt
     1441 ggactcaaga cgatagttac cggataaggc gcagcggtcg ggctgaacgg ggggttcgtg
     1501 cacacagccc agcttggagc gaacgaccta caccgaactg agatacctac agcgtgagct
     1561 atgagaaagc gccacgcttc ccgaagggag aaaggcggac aggtatccgg taagcggcag
     1621 ggtcggaaca ggagagcgca cgagggagct tccaggggga aacgcctggt atctttatag
     1681 tcctgtcggg tttcgccacc tctgacttga gcgtcgattt ttgtgatgct cgtcaggggg
     1741 gcggagccta tggaaaaacg ccagcaacgc ggccttttta cggttcctgg ccttttgctg
     1801 gccttttgct cacatgttct ttcctgcgtt atcccctgat tctgtggata accgtattac
     1861 cgcctttgag tgagctgata ccgctcgccg cagccgaacg accgagcgca gcgagtcagt
     1921 gagcgaggaa gcggaagagc gagcttcgca cgattgcacc ttgggagcct taaccgcttt
     1981 tgaatgtccg tcttagttaa tccctaattg aacgctccaa aaggtgactt ccaataggga
     2041 tttattcctt ttaaaattaa cggcataatc gtgccgccga atgcaacctc gatcgagccg
     2101 ggctccggtg cccgtcagtg ggcagagcgc acatcgccca cagtccccga gaagttgggg
     2161 ggaggggtcg gcaattgaac cggtgcctag agaaggtggc gcggggtaaa ctgggaaagt
     2221 gatgtcgtgt actggctccg cctttttccc gagggtgggg gagaaccgta tataagtgca
     2281 gtagtcgccg tgaacgttct ttttcgcaac gggtttgccg ccagaacaca ggtaagtgcc
     2341 gtgtgtggtt cccgcgggcc tggcctcttt acgggttatg gcccttgcgt gccttgaatt
     2401 acttccacct ggctgcagta cgtgattctt gatcccgagc ttcgggttgg aagtgggtgg
     2461 gagagttcga ggccttgcgc ttaaggagcc ccttcgcctc gtgcttgagt tgaggcctgg
     2521 cctgggcgct ggggccgccg cgtgcgaatc tggtggcacc ttcgcgcctg tctcgctgct
     2581 ttcgataagt ctctagccat ttaaaatttt tgatgacctg ctgcgacgct ttttttctgg
     2641 caagatagtc ttgtaaatgc gggccaagat ctgcacactg gtatttcggt ttttggggcc
     2701 gcgggcggcg acggggcccg tgcgtcccag cgcacatgtt cggcgaggcg gggcctgcga
     2761 gcgcggccac cgagaatcgg acgggggtag tctcaagctg gccggcctgc tctggtgcct
     2821 ggcctcgcgc cgccgtgtat cgccccgccc tgggcggcaa ggctggcccg gtcggcacca
     2881 gttgcgtgag cggaaagatg gccgcttccc ggccctgctg cagggagctc aaaatggagg
     2941 acgcggcgct cgggagagcg ggcgggtgag tcacccacac aaaggaaaag ggcctttccg
     3001 tcctcagccg tcgcttcatg tgactccacg gagtaccggg cgccgtccag gcacctcgat
     3061 tagttctcga gcttttggag tacgtcgtct ttaggttggg gggaggggtt ttatgcgatg
     3121 gagtttcccc acactgagtg ggtggagact gaagttaggc cagcttggca cttgatgtaa
     3181 ttctccttgg aatttgccct ttttgagttt ggatcttggt tcattctcaa gcctcagaca
     3241 gtggttcaaa gtttttttct tccatttcag gtgtcgtgaa gccaccatga ccgagtacaa
     3301 gcccacggtg cgcctcgcca cccgcgacga cgtccccagg gccgtacgca ccctcgccgc
     3361 cgcgttcgcc gactaccccg ccacgcgcca caccgtcgat ccggaccgcc acatcgagcg
     3421 ggtcaccgag ctgcaagaac tcttcctcac gcgcgtcggg ctcgacatcg gcaaggtgtg
     3481 ggtcgcggac gacggcgccg ccgtggcggt ctggaccacg ccggagagcg tcgaagcggg
     3541 ggcggtgttc gccgagatcg gcccgcgcat ggccgagttg agcggttccc ggctggccgc
     3601 gcagcaacag atggaaggcc tcctggcgcc gcaccggccc aaggagcccg cgtggttcct
     3661 ggccaccgtc ggagtctcgc ccgaccacca gggcaagggt ctgggcagcg ccgtcgtgct
     3721 ccccggagtg gaggcggccg agcgcgccgg ggtgcccgcc ttcctggaga cctccgcgcc
     3781 ccgcaacctc cccttctacg agcggctcgg cttcaccgtc accgccgacg tcgaggtgcc
     3841 cgaaggaccg cgcacctggt gcatgacccg caagcccggt gccgccacga acttctctct
     3901 gttaaagcaa gcaggtgacg tggaagaaaa ccccggtccc ggatccatgg tgagcaaggg
     3961 cgaggaggat aacatggcca tcatcaagga gttcatgcgc ttcaaggtgc acatggaggg
     4021 ctccgtgaac ggccacgagt tcgagatcga gggcgagggc gagggccgcc cctacgaggg
     4081 cacccagacc gccaagctga aggtgaccaa gggtggcccc ctgcccttcg cctgggacat
     4141 cctgtcccct cagttcatgt acggctccaa ggcctacgtg aagcaccccg ccgacatccc
     4201 cgactacttg aagctgtcct tccccgaggg cttcaagtgg gagcgcgtga tgaacttcga
     4261 ggacggcggc gtggtgaccg tgacccagga ctcctccctg caggacggcg agttcatcta
     4321 caaggtgaag ctgcgcggca ccaacttccc ctccgacggc cccgtaatgc agaagaagac
     4381 catgggctgg gaggcctcct ccgagcggat gtaccccgag gacggcgccc tgaagggcga
     4441 gatcaagcag aggctgaagc tgaaggacgg cggccactac gacgctgagg tcaagaccac
     4501 ctacaaggcc aagaagcccg tgcagctgcc cggcgcctac aacgtcaaca tcaagttgga
     4561 catcacctcc cacaacgagg actacaccat cgtggaacag tacgaacgcg ccgagggccg
     4621 ccactccacc ggcggcatgg acgagctgta caagtaactc gagctagagc tcgctgatca
     4681 gcctcgactg tgccttctag ttgccagcca tctgttgttt gcccctcccc cgtgccttcc
     4741 ttgaccctgg aaggtgccac tcccactgtc ctttcctaat aaaatgagga aattgcatcg
     4801 cattgtctga gtaggtgtca ttctattctg gggggtgggg tggggcagga cagcaagggg
     4861 gaggattggg aagacaatag caggcatgct ggggac
//
