LOCUS       sequence-364695-        4732 bp    DNA     circular SYN 14-APR-2025
DEFINITION  Localization: N1 Cloning Vector, Excitation: 383, Emission: 448.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4732)
  AUTHORS   Davidson Lab
  TITLE     Michael Davidson Empty Backbones
  JOURNAL   unpublished
REFERENCE   2  (bases 1 to 4732)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Apr 14, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: 
            "unpublished"
COMMENT     Sequence Label: EBFP2-N1
FEATURES             Location/Qualifiers
     source          1..4732
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        1..304
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        305..508
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     459..479
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     misc_feature    531..611
                     /label=MCS
                     /note="multiple cloning site"
     regulatory      613..622
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(619..621,622..624,625..1338)
                     /codon_start=1
                     /product="enhanced blue variant of GFP (Ai et al., 2007)"
                     /label=EBFP2
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVRGEGEGDATNGKLTL
                     KFICTTGKLPVPWPTLVTTLSHGVQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GTYKTRAEVKFEGDTLVNRIELKGVDFKEDGNILGHKLEYNFNSHNIYIMAVKQKNGIK
                     VNFKIRHNVEDGSVQLADHYQQNTPIGDGPVLLPDSHYLSTQSVLSKDPNEKRDHMVLL
                     EFRTAAGITLGMDELYK"
     primer_bind     complement(664..685)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     polyA_signal    1460..1581
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1497..1516)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     1551..1570
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(1588..2043)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1725..1746)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     1937..1956
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        2070..2174
                     /gene="bla"
                     /label=AmpR promoter
     promoter        2176..2533
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        2176..2367
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(2199..2219)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      2384..2519
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             2568..3362
                     /codon_start=1
                     /product="aminoglycoside phosphotransferase"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     CDS             2568..3362
                     /codon_start=1
                     /gene="aph(3')-II (or nptII)"
                     /product="aminoglycoside phosphotransferase from Tn5"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(2622..2641)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     3232..3251
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(3550..3569)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    3594..3641
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      3970..4558
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4459..4478
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc cccgcccatt
       61 gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc attgacgtca
      121 atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt atcatatgcc
      181 aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt atgcccagta
      241 catgacctta tgggactttc ctacttggca gtacatctac gtattagtca tcgctattac
      301 catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg actcacgggg
      361 atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc aaaatcaacg
      421 ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg gtaggcgtgt
      481 acggtgggag gtctatataa gcagagctgg tttagtgaac cgtcagatcc gctagcgcta
      541 ccggactcag atctcgagct caagcttcga attctgcagt cgacggtacc gcgggcccgg
      601 gatccaccgg tcgccaccat ggtgagcaag ggcgaggagc tgttcaccgg ggtggtgccc
      661 atcctggtcg agctggacgg cgacgtaaac ggccacaagt tcagcgtgag gggcgagggc
      721 gagggcgatg ccaccaacgg caagctgacc ctgaagttca tctgcaccac cggcaagctg
      781 cccgtgccct ggcccaccct cgtgaccacc ctgagccacg gcgtgcagtg cttcgcccgc
      841 taccccgacc acatgaagca gcacgacttc ttcaagtccg ccatgcccga aggctacgtc
      901 caggagcgca ccatcttctt caaggacgac ggcacctaca agacccgcgc cgaggtgaag
      961 ttcgagggcg acaccctggt gaaccgcatc gagctgaagg gcgtcgactt caaggaggac
     1021 ggcaacatcc tggggcacaa gctggagtac aacttcaaca gccacaacat ctatatcatg
     1081 gccgtcaagc agaagaacgg catcaaggtg aacttcaaga tccgccacaa cgtggaggac
     1141 ggcagcgtgc agctcgccga ccactaccag cagaacaccc ccatcggcga cggccccgtg
     1201 ctgctgcccg acagccacta cctgagcacc cagtccgtgc tgagcaaaga ccccaacgag
     1261 aagcgcgatc acatggtcct gctggagttc cgcaccgccg ccgggatcac tctcggcatg
     1321 gacgagctgt acaagtaagc ggccgcgact ctagatcata atcagccata ccacatttgt
     1381 agaggtttta cttgctttaa aaaacctccc acacctcccc ctgaacctga aacataaaat
     1441 gaatgcaatt gttgttgtta acttgtttat tgcagcttat aatggttaca aataaagcaa
     1501 tagcatcaca aatttcacaa ataaagcatt tttttcactg cattctagtt gtggtttgtc
     1561 caaactcatc aatgtatctt aaggcgtaaa ttgtaagcgt taatattttg ttaaaattcg
     1621 cgttaaattt ttgttaaatc agctcatttt ttaaccaata ggccgaaatc ggcaaaatcc
     1681 cttataaatc aaaagaatag accgagatag ggttgagtgt tgttccagtt tggaacaaga
     1741 gtccactatt aaagaacgtg gactccaacg tcaaagggcg aaaaaccgtc tatcagggcg
     1801 atggcccact acgtgaacca tcaccctaat caagtttttt ggggtcgagg tgccgtaaag
     1861 cactaaatcg gaaccctaaa gggagccccc gatttagagc ttgacgggga aagccggcga
     1921 acgtggcgag aaaggaaggg aagaaagcga aaggagcggg cgctagggcg ctggcaagtg
     1981 tagcggtcac gctgcgcgta accaccacac ccgccgcgct taatgcgccg ctacagggcg
     2041 cgtcaggtgg cacttttcgg ggaaatgtgc gcggaacccc tatttgttta tttttctaaa
     2101 tacattcaaa tatgtatccg ctcatgagac aataaccctg ataaatgctt caataatatt
     2161 gaaaaaggaa gagtcctgag gcggaaagaa ccagctgtgg aatgtgtgtc agttagggtg
     2221 tggaaagtcc ccaggctccc cagcaggcag aagtatgcaa agcatgcatc tcaattagtc
     2281 agcaaccagg tgtggaaagt ccccaggctc cccagcaggc agaagtatgc aaagcatgca
     2341 tctcaattag tcagcaacca tagtcccgcc cctaactccg cccatcccgc ccctaactcc
     2401 gcccagttcc gcccattctc cgccccatgg ctgactaatt ttttttattt atgcagaggc
     2461 cgaggccgcc tcggcctctg agctattcca gaagtagtga ggaggctttt ttggaggcct
     2521 aggcttttgc aaagatcgat caagagacag gatgaggatc gtttcgcatg attgaacaag
     2581 atggattgca cgcaggttct ccggccgctt gggtggagag gctattcggc tatgactggg
     2641 cacaacagac aatcggctgc tctgatgccg ccgtgttccg gctgtcagcg caggggcgcc
     2701 cggttctttt tgtcaagacc gacctgtccg gtgccctgaa tgaactgcaa gacgaggcag
     2761 cgcggctatc gtggctggcc acgacgggcg ttccttgcgc agctgtgctc gacgttgtca
     2821 ctgaagcggg aagggactgg ctgctattgg gcgaagtgcc ggggcaggat ctcctgtcat
     2881 ctcaccttgc tcctgccgag aaagtatcca tcatggctga tgcaatgcgg cggctgcata
     2941 cgcttgatcc ggctacctgc ccattcgacc accaagcgaa acatcgcatc gagcgagcac
     3001 gtactcggat ggaagccggt cttgtcgatc aggatgatct ggacgaagag catcaggggc
     3061 tcgcgccagc cgaactgttc gccaggctca aggcgagcat gcccgacggc gaggatctcg
     3121 tcgtgaccca tggcgatgcc tgcttgccga atatcatggt ggaaaatggc cgcttttctg
     3181 gattcatcga ctgtggccgg ctgggtgtgg cggaccgcta tcaggacata gcgttggcta
     3241 cccgtgatat tgctgaagag cttggcggcg aatgggctga ccgcttcctc gtgctttacg
     3301 gtatcgccgc tcccgattcg cagcgcatcg ccttctatcg ccttcttgac gagttcttct
     3361 gagcgggact ctggggttcg aaatgaccga ccaagcgacg cccaacctgc catcacgaga
     3421 tttcgattcc accgccgcct tctatgaaag gttgggcttc ggaatcgttt tccgggacgc
     3481 cggctggatg atcctccagc gcggggatct catgctggag ttcttcgccc accctagggg
     3541 gaggctaact gaaacacgga aggagacaat accggaagga acccgcgcta tgacggcaat
     3601 aaaaagacag aataaaacgc acggtgttgg gtcgtttgtt cataaacgcg gggttcggtc
     3661 ccagggctgg cactctgtcg ataccccacc gagaccccat tggggccaat acgcccgcgt
     3721 ttcttccttt tccccacccc accccccaag ttcgggtgaa ggcccagggc tcgcagccaa
     3781 cgtcggggcg gcaggccctg ccatagcctc aggttactca tatatacttt agattgattt
     3841 aaaacttcat ttttaattta aaaggatcta ggtgaagatc ctttttgata atctcatgac
     3901 caaaatccct taacgtgagt tttcgttcca ctgagcgtca gaccccgtag aaaagatcaa
     3961 aggatcttct tgagatcctt tttttctgcg cgtaatctgc tgcttgcaaa caaaaaaacc
     4021 accgctacca gcggtggttt gtttgccgga tcaagagcta ccaactcttt ttccgaaggt
     4081 aactggcttc agcagagcgc agataccaaa tactgttctt ctagtgtagc cgtagttagg
     4141 ccaccacttc aagaactctg tagcaccgcc tacatacctc gctctgctaa tcctgttacc
     4201 agtggctgct gccagtggcg ataagtcgtg tcttaccggg ttggactcaa gacgatagtt
     4261 accggataag gcgcagcggt cgggctgaac ggggggttcg tgcacacagc ccagcttgga
     4321 gcgaacgacc tacaccgaac tgagatacct acagcgtgag ctatgagaaa gcgccacgct
     4381 tcccgaaggg agaaaggcgg acaggtatcc ggtaagcggc agggtcggaa caggagagcg
     4441 cacgagggag cttccagggg gaaacgcctg gtatctttat agtcctgtcg ggtttcgcca
     4501 cctctgactt gagcgtcgat ttttgtgatg ctcgtcaggg gggcggagcc tatggaaaaa
     4561 cgccagcaac gcggcctttt tacggttcct ggccttttgc tggccttttg ctcacatgtt
     4621 ctttcctgcg ttatcccctg attctgtgga taaccgtatt accgccatgc attagttatt
     4681 aatagtaatc aattacgggg tcattagttc atagcccata tatggagttc cg
//
