LOCUS       sequence-222569-        4438 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  p3E entry vector for Three Multisite Gateway.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4438)
  TITLE     p3E_he1a constructs
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 4438)
  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: 
            "Unpublished"
COMMENT     Sequence Label: p3E_he1a:ECFP
FEATURES             Location/Qualifiers
     source          1..4438
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     protein_bind    complement(62..186)
                     /gene="mutant version of attR"
                     /label=attR2
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     regulatory      665..674
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(671..673,674..676,677..1390)
                     /codon_start=1
                     /product="enhanced CFP"
                     /label=ECFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTWGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYISHNVYITADKQKNGIK
                     ANFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(716..737)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(977..996)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1324..1345
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     complement(1753..1770)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    1759..1984
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     protein_bind    complement(1985..2080)
                     /gene="mutant version of attL"
                     /label=attL3
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     primer_bind     complement(2117..2136)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(2118..2136)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(2141..2157)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2141..2157)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(2259..2278)
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     CDS             2270..3079
                     /codon_start=1
                     /gene="aph(3')-Ia"
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYGKP
                     DAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGKTA
                     FQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDASD
                     FDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGIAD
                     RYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     primer_bind     complement(2337..2356)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     rep_origin      3226..3814
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3715..3734
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     3968..3985
                     /label=L4440
                     /note="L4440 vector, forward primer"
     terminator      complement(4144..4171)
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB 
                     gene"
     terminator      complement(4263..4349)
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     primer_bind     4398..4420
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     4412..4429
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     4413..4429
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
ORIGIN
        1 gggccctgca gctctagagc tcgaattcta caggtcacta ataccatcta agtagttggt
       61 tcatagtgac tgcatatgtt gtgttttaca gtattatgta gtctgttttt tatgcaaaat
      121 ctaatttaat atattgatat ttatatcatt ttacgtttct cgttcaactt tcttgtacaa
      181 agtggtcaac cactccaggc atagctagag ccactgatgc atctatagag ctgaagtaaa
      241 gatagggttg atgcaaatcg atttagaatt ttcaatgtag aaaaaacaaa ataaaaagtc
      301 tattcacttg atggatagat tagtcaaaca aacatagagt aggcacttcc cttgactctg
      361 taaacttttg gatttaacag attgtgcact ccttttagtt aaacacatta gtccctgaaa
      421 actaagaccc taaccataat cccattgtag acccacctac tcgggtgaat ataaagctcg
      481 cacagatcat ctcacagaca tcagaccttg gcaaaggagc atcctgaaca tctctacaca
      541 ctgaggcaac cctagggccg ccaccatgct gtgttgtatc agaagaacaa agcctgtgga
      601 gaagaatgag gaggctgacc aggaactgca gagcacggta ccgcgggccc gggatccacc
      661 ggccgccacc atggtgagca agggcgagga gctgttcacc ggggtggtgc ccatcctggt
      721 cgagctggac ggcgacgtaa acggccacaa gttcagcgtg tccggcgagg gcgagggcga
      781 tgccacctac ggcaagctga ccctgaagtt catctgcacc accggcaagc tgcccgtgcc
      841 ctggcccacc ctcgtgacca ccctgacctg gggcgtgcag tgcttcagcc gctaccccga
      901 ccacatgaag cagcacgact tcttcaagtc cgccatgccc gaaggctacg tccaggagcg
      961 caccatcttc ttcaaggacg acggcaacta caagacccgc gccgaggtga agttcgaggg
     1021 cgacaccctg gtgaaccgca tcgagctgaa gggcatcgac ttcaaggagg acggcaacat
     1081 cctggggcac aagctggagt acaactacat cagccacaac gtctatatca ccgccgacaa
     1141 gcagaagaac ggcatcaagg ccaacttcaa gatccgccac aacatcgagg acggcagcgt
     1201 gcagctcgcc gaccactacc agcagaacac ccccatcggc gacggccccg tgctgctgcc
     1261 cgacaaccac tacctgagca cccagtccgc cctgagcaaa gaccccaacg agaagcgcga
     1321 tcacatggtc ctgctggagt tcgtgaccgc cgccgggatc actctcggca tggacgagct
     1381 gtacaagtaa agcggccgcg actctagatc ataatcagcc ataccacatt tgtagaggtt
     1441 ttacttgctt taaaaaacct cccacacctc cccctgaacc tgaaacataa aatgaatgca
     1501 attgttgttg ttatcgaatt ctgcaggaat tgggtggcat ccctgtgacc cctccccagt
     1561 gcctctcctg gccctggaag ttgccactcc agtgcccacc agccttgtcc taataaaatt
     1621 aagttgcatc attttgtctg actaggtgtc cttctataat attatggggt ggaggggggt
     1681 ggtatggagc aaggggcaag ttgggaagac aacctgtagg gcacgagggg gggcccgaaa
     1741 cccgctgatc agcctcgact gtgccttcta gttgccagcc atctgttgtt tgcccctccc
     1801 ccgtgccttc cttgaccctg gaaggtgcca ctcccactgt cctttcctaa taaaatgagg
     1861 aaattgcatc gcattgtctg agtaggtgtc attctattct ggggggtggg gttgggcagg
     1921 acagcaaggg ggaggattgg gaagacaata gcaaggcatg ctggggatgc ggtgggctct
     1981 atggcaactt tattatacaa agttggcatt ataaaaaagc attgcttatc aatttgttgc
     2041 aacgaacagg tcactatcag tcaaaataaa atcattattt ggagctccat ggtagcgtta
     2101 acgcggccgc gatatcccct atagtgagtc gtattacatg gtcatagctg tttcctggca
     2161 gctctggccc gtgtctcaaa atctctgatg ttacattgca caagataaaa atatatcatc
     2221 atgaacaata aaactgtctg cttacataaa cagtaataca aggggtgtta tgagccatat
     2281 tcaacgggaa acgtcgaggc cgcgattaaa ttccaacatg gatgctgatt tatatgggta
     2341 taaatgggct cgcgataatg tcgggcaatc aggtgcgaca atctatcgct tgtatgggaa
     2401 gcccgatgcg ccagagttgt ttctgaaaca tggcaaaggt agcgttgcca atgatgttac
     2461 agatgagatg gtcagactaa actggctgac ggaatttatg cctcttccga ccatcaagca
     2521 ttttatccgt actcctgatg atgcatggtt actcaccact gcgatccccg gaaaaacagc
     2581 attccaggta ttagaagaat atcctgattc aggtgaaaat attgttgatg cgctggcagt
     2641 gttcctgcgc cggttgcatt cgattcctgt ttgtaattgt ccttttaaca gcgatcgcgt
     2701 atttcgtctc gctcaggcgc aatcacgaat gaataacggt ttggttgatg cgagtgattt
     2761 tgatgacgag cgtaatggct ggcctgttga acaagtctgg aaagaaatgc ataaactttt
     2821 gccattctca ccggattcag tcgtcactca tggtgatttc tcacttgata accttatttt
     2881 tgacgagggg aaattaatag gttgtattga tgttggacga gtcggaatcg cagaccgata
     2941 ccaggatctt gccatcctat ggaactgcct cggtgagttt tctccttcat tacagaaacg
     3001 gctttttcaa aaatatggta ttgataatcc tgatatgaat aaattgcagt ttcatttgat
     3061 gctcgatgag tttttctaat cagaattggt taattggttg taacactggc agagcattac
     3121 gctgacttga cgggacggcg caagctcatg accaaaatcc cttaacgtga gttacgcgtc
     3181 gttccactga gcgtcagacc ccgtagaaaa gatcaaagga tcttcttgag atcctttttt
     3241 tctgcgcgta atctgctgct tgcaaacaaa aaaaccaccg ctaccagcgg tggttttttt
     3301 gccggatcaa gagctaccaa ctctttttcc gaaggtaact ggcttcagca gagcgcagat
     3361 accaaatact gttcttctag tgtagccgta gttaggccac cacttcaaga actctgtagc
     3421 accgcctaca tacctcgctc tgctaatcct gttaccagtg gctgctgcca gtggcgataa
     3481 gtcgtgtctt accgggttgg actcaagacg atagttaccg gataaggcgc agcggtcggg
     3541 ctgaacgggg ggttcgtgca cacagcccag cttggagcga acgacctaca ccgaactgag
     3601 atacctacag cgtgagctat gagaaagcgc cacgcttccc gaagggagaa aggcggacag
     3661 gtatccggta agcggcaggg tcggaacagg agagcgcacg agggagcttc cagggggaaa
     3721 cgcctggtat ctttatagtc ctgtcgggtt tcgccacctc tgacttgagc gtcgattttt
     3781 gtgatgctcg tcaggggggc ggagcctatg gaaaaacgcc agcaacgcgg cctttttacg
     3841 gttcctggcc ttttgctggc cttttgctca catgttcttt cctgcgttat cccctgattc
     3901 tgtggataac cgtattaccg cctttgagtg agctgatacc gctcgccgca gccgaacgac
     3961 cgagcgcagc gagtcagtga gcgaggaagc ggaagagcgc ccaatacgca aaccgcctct
     4021 ccccgcgcgt tggccgattc attaatgcag ctggcacgac aggtttcccg actggaaagc
     4081 gggcagtgag cgcaacgcaa ttaatacgcg taccgctagc caggaagagt ttgtagaaac
     4141 gcaaaaaggc catccgtcag gatggccttc tgcttagttt gatgcctggc agtttatggc
     4201 gggcgtcctg cccgccaccc tccgggccgt tgcttcacaa cgttcaaatc cgctcccggc
     4261 ggatttgtcc tactcaggag agcgttcacc gacaaacaac agataaaacg aaaggcccag
     4321 tcttccgact gagcctttcg ttttatttga tgcctggcag ttccctactc tcgcgttaac
     4381 gctagcatgg atgttttccc agtcacgacg ttgtaaaacg acggccagtc ttaagctc
//
