LOCUS       Exported                7372 bp ds-DNA     circular SYN 18-JUL-2024
DEFINITION  synthetic circular DNA
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7372)
  AUTHORS   SnapGene License
  TITLE     Direct Submission
  JOURNAL   Exported Monday, Jul 7, 2025 from SnapGene 5.0.8
            https://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..7372
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     promoter        475..1653
                     /label=EF-1-alpha promoter
                     /note="strong constitutive promoter for human elongation 
                     factor EF-1-alpha"
     intron          706..1644
                     /label=EF-1-alpha intron A
                     /note="intron upstream of the start codon of human 
                     EF-1-alpha"
     promoter        1670..1688
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     1720..1768
                     /label=Primer 1
     protein_bind    1720..1744
                     /gene="mutant version of attB"
                     /label=attB1
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     misc_feature    1745..1767
                     /label=MCS (fragment)
                     /note="pLannotate"
                     /note="/database=snapgene"
                     /note="/fragment=True"
                     /note="/identity=100.0"
                     /note="/match_length=28.4"
                     /note="/other=misc_feature"
     CDS             1782..2274
                     /codon_start=1
                     /label=EGFP
                     /label=EGFP 5' half
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     V"
     regulatory      1782..1785
                     /regulatory_class="other"
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
     intron          2275..2294
                     /label=Artificial intron and MCS
     misc_feature    2301..2800
                     /label=human Chr 9 / EGFL7
     ncRNA           2536..2608
                     /label=mir-126
                     /note="pLannotate"
                     /note="/database=Rfam"
                     /note="/fragment=False"
                     /note="/identity=100.0"
                     /note="/match_length=100.0"
                     /note="/other=ncRNA"
     intron          2801..3050
                     /label=Artificial intron and MCS
     misc_feature    2819..2839
                     /label=critical T stretch
     CDS             3051..3279
                     /codon_start=1
                     /label=EGFP
                     /label=EGFP 5' half
                     /translation="LQDPPQHRGRQRAARRPLPAEHPHRRRPRAAARQPLPEHPVRPEQ
                     RPQREARSHGPAGVRDRRRDHSRHGRAVQV"
     primer_bind     complement(3266..3332)
                     /label=Primer 2
     protein_bind    complement(3308..3332)
                     /gene="mutant version of attB"
                     /label=attB2
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     CDS             3358..3399
                     /codon_start=1
                     /product="epitope tag from simian virus 5"
                     /label=V5 tag
                     /translation="GKPIPNPLLGLDST"
     CDS             3409..3426
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     misc_feature    3449..3466
                     /label=BGH rev primers
     polyA_signal    3455..3679
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      3725..4153
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     promoter        4167..4497
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     promoter        4545..4592
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             4611..5009
                     /codon_start=1
                     /gene="Aspergillus terreus BSD"
                     /product="blasticidin S deaminase"
                     /label=BSD
                     /note="confers resistance to blasticidin"
                     /translation="MAKPLSQEESTLIERATATINSIPISEDYSVASAALSSDGRIFTG
                     VNVYHFTGGPCAELVVLGTAAAAAAGNLTCIVAIGNENRGILSPCGRCRQVLLDLHPGI
                     KAIVKDSDGQPTAVGIRELLPSGYVWEG"
     polyA_signal    5167..5288
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     misc_feature    complement(5204..5223)
                     /label=SV40pA REVSeq
     primer_bind     complement(5333..5353)
                     /label=M13-rev
     protein_bind    5361..5377
                     /label=lac operator
                     /bound_moiety="lac repressor encoded by lacI"
                     /note="The lac repressor binds to the lac operator to 
                     inhibit transcription in E. coli. This inhibition can be 
                     relieved by adding lactose or 
                     isopropyl-beta-D-thiogalactopyranoside (IPTG)."
     promoter        complement(5385..5415)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5430..5451
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     rep_origin      complement(5721..6403)
                     /direction=LEFT
                     /label=ColE1 origin
     CDS             complement(6498..7358)
                     /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"
     promoter        complement(join(7359..7372,1..91))
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 aatattattg aagcatttat cagggttatt gtctcatgag cggatacata tttgaatgta
       61 tttagaaaaa taaacaaata ggggttccgc gcacatttcc ccgaaaagtg ccacctgacg
      121 tcgacggatc gggagatctc ccgatcccct atggtcgact ctcagtacaa tctgctctga
      181 tgccgcatag ttaagccagt atctgctccc tgcttgtgtg ttggaggtcg ctgagtagtg
      241 cgcgagcaaa atttaagcta caacaaggca aggcttgacc gacaattgca tgaagaatct
      301 gcttagggtt aggcgttttg cgctgcttcg cgatgtacgg gccagatata cgcgttgaca
      361 ttgattattg actaggcttt tgcaaaaagc tttgcaaaga tggataaagt tttaaacaga
      421 gaggaatctt tgcagctaat ggaccttcta ggtcttgaaa ggagtgcctc gtgaggctcc
      481 ggtgcccgtc agtgggcaga gcgcacatcg cccacagtcc ccgagaagtt ggggggaggg
      541 gtcggcaatt gaaccggtgc ctagagaagg tggcgcgggg taaactggga aagtgatgtc
      601 gtgtactggc tccgcctttt tcccgagggt gggggagaac cgtatataag tgcagtagtc
      661 gccgtgaacg ttctttttcg caacgggttt gccgccagaa cacaggtaag tgccgtgtgt
      721 ggttcccgcg ggcctggcct ctttacgggt tatggccctt gcgtgccttg aattacttcc
      781 acctggctgc agtacgtgat tcttgatccc gagcttcggg ttggaagtgg gtgggagagt
      841 tcgaggcctt gcgcttaagg agccccttcg cctcgtgctt gagttgaggc ctggcctggg
      901 cgctggggcc gccgcgtgcg aatctggtgg caccttcgcg cctgtctcgc tgctttcgat
      961 aagtctctag ccatttaaaa tttttgatga cctgctgcga cgcttttttt ctggcaagat
     1021 agtcttgtaa atgcgggcca agatctgcac actggtattt cggtttttgg ggccgcgggc
     1081 ggcgacgggg cccgtgcgtc ccagcgcaca tgttcggcga ggcggggcct gcgagcgcgg
     1141 ccaccgagaa tcggacgggg gtagtctcaa gctggccggc ctgctctggt gcctggcctc
     1201 gcgccgccgt gtatcgcccc gccctgggcg gcaaggctgg cccggtcggc accagttgcg
     1261 tgagcggaaa gatggccgct tcccggccct gctgcaggga gctcaaaatg gaggacgcgg
     1321 cgctcgggag agcgggcggg tgagtcaccc acacaaagga aaagggcctt tccgtcctca
     1381 gccgtcgctt catgtgactc cacggagtac cgggcgccgt ccaggcacct cgattagttc
     1441 tcgagctttt ggagtacgtc gtctttaggt tggggggagg ggttttatgc gatggagttt
     1501 ccccacactg agtgggtgga gactgaagtt aggccagctt ggcacttgat gtaattctcc
     1561 ttggaatttg ccctttttga gtttggatct tggttcattc tcaagcctca gacagtggtt
     1621 caaagttttt ttcttccatt tcaggtgtcg tgaggaatta gcttggtact aatacgactc
     1681 actataggga gacccaagct ggctaggtaa gcttgatcaa caagtttgta caaaaaagca
     1741 ggcagctagc gctaccggac tcagatccac cggtcgccac catggtgagc aagggcgagg
     1801 agctgttcac cggggtggtg cccatcctgg tcgagctgga cggcgacgta aacggccaca
     1861 agttcagcgt gtccggcgag ggcgagggcg atgccaccta cggcaagctg accctgaagt
     1921 tcatctgcac caccggcaag ctgcccgtgc cctggcccac cctcgtgacc accctgacct
     1981 acggcgtgca gtgcttcagc cgctaccccg accacatgaa gcagcacgac ttcttcaagt
     2041 ccgccatgcc cgaaggctac gtccaggagc gcaccatctt cttcaaggac gacggcaact
     2101 acaagacccg cgccgaggtg aagttcgagg gcgacaccct ggtgaaccgc atcgagctga
     2161 agggcatcga cttcaaggag gacggcaaca tcctggggca caagctggag tacaactaca
     2221 acagccacaa cgtctatatc atggccgaca agcagaagaa cggcatcaag gtgagtggat
     2281 cgggtaccga attcgccctt agtggacatt gccgtgtggc tgttaggcat ggtggggggc
     2341 actggaatct gggcggaagg cggtggggac tccctctcca gggagggagg atggggaggg
     2401 aggataggtg ggttcccgag aactgggggc aggttgcccg gagcctcata tcagccaaga
     2461 aggcagaagt gccccgtccc ggggtcctgt ctgcatccag cgcagcattc tggaagacgc
     2521 cacgcctccg ctggcgacgg gacattatta cttttggtac gcgctgcgac acttcaaact
     2581 cgtaccgtga gtaataatgc gccgtccacg gcaccgcatc gaaaacgccg ctgagacctc
     2641 agccttgacc tccctcagcg tggccgggac cctgagcctc tgcgcagagc cacccgcccc
     2701 gacgtactta ggcggcatag ccctgagacc tctggccagc gccaggcagg cagcgggggc
     2761 ggcagaggcc tgggcctgag tcttctggct ctgcctctcc aagggcgaat tccatgatct
     2821 tttttttttt tttttttttc ccatgatcat cccggggatc ctgcagatgc agtcagattt
     2881 ctgaactgtc tcagcaaatg gtaagtaact tagagctagt aattaataac ctgtcctttg
     2941 atttctgatt cagccaagaa tggccatatt tgggaaaggc agatctggag agtaaccacg
     3001 ttttcatgca gtctagactg ttaacgcagt gtttctctaa ctctcaggtg aacttcaaga
     3061 tccgccacaa catcgaggac ggcagcgtgc agctcgccga ccactaccag cagaacaccc
     3121 ccatcggcga cggccccgtg ctgctgcccg acaaccacta cctgagcacc cagtccgccc
     3181 tgagcaaaga ccccaacgag aagcgcgatc acatggtcct gctggagttc gtgaccgccg
     3241 ccgggatcac tctcggcatg gacgagctgt acaagtaaag cggccgcgac tctagacgcg
     3301 tactagtgcc cagctttctt gtacaaagtg gttgatctag agggcccgcg gttcgaaggt
     3361 aagcctatcc ctaaccctct cctcggtctc gattctacgc gtaccggtca tcatcaccat
     3421 caccattgag tttaaacccg ctgatcagcc tcgactgtgc cttctagttg ccagccatct
     3481 gttgtttgcc cctcccccgt gccttccttg accctggaag gtgccactcc cactgtcctt
     3541 tcctaataaa atgaggaaat tgcatcgcat tgtctgagta ggtgtcattc tattctgggg
     3601 ggtggggtgg ggcaggacag caagggggag gattgggaag acaatagcag gcatgctggg
     3661 gatgcggtgg gctctatggc ttctgaggcg gaaagaacca gctggggctc tagggggtat
     3721 ccccacgcgc cctgtagcgg cgcattaagc gcggcgggtg tggtggttac gcgcagcgtg
     3781 accgctacac ttgccagcgc cctagcgccc gctcctttcg ctttcttccc ttcctttctc
     3841 gccacgttcg ccggctttcc ccgtcaagct ctaaatcggg gcatcccttt agggttccga
     3901 tttagtgctt tacggcacct cgaccccaaa aaacttgatt agggtgatgg ttcacgtagt
     3961 gggccatcgc cctgatagac ggtttttcgc cctttgacgt tggagtccac gttctttaat
     4021 agtggactct tgttccaaac tggaacaaca ctcaacccta tctcggtcta ttcttttgat
     4081 ttataaggga ttttggggat ttcggcctat tggttaaaaa atgagctgat ttaacaaaaa
     4141 tttaacgcga attaattctg tggaatgtgt gtcagttagg gtgtggaaag tccccaggct
     4201 ccccaggcag gcagaagtat gcaaagcatg catctcaatt agtcagcaac caggtgtgga
     4261 aagtccccag gctccccagc aggcagaagt atgcaaagca tgcatctcaa ttagtcagca
     4321 accatagtcc cgcccctaac tccgcccatc ccgcccctaa ctccgcccag ttccgcccat
     4381 tctccgcccc atggctgact aatttttttt atttatgcag aggccgaggc cgcctctgcc
     4441 tctgagctat tccagaagta gtgaggaggc ttttttggag gcctaggctt ttgcaaaaag
     4501 ctcccgggag cttgtatatc cattttcgga tctgatcagc acgtgttgac aattaatcat
     4561 cggcatagta tatcggcata gtataatacg acaaggtgag gaactaaacc atggccaagc
     4621 ctttgtctca agaagaatcc accctcattg aaagagcaac ggctacaatc aacagcatcc
     4681 ccatctctga agactacagc gtcgccagcg cagctctctc tagcgacggc cgcatcttca
     4741 ctggtgtcaa tgtatatcat tttactgggg gaccttgtgc agaactcgtg gtgctgggca
     4801 ctgctgctgc tgcggcagct ggcaacctga cttgtatcgt cgcgatcgga aatgagaaca
     4861 ggggcatctt gagcccctgc ggacggtgtc gacaggtgct tctcgatctg catcctggga
     4921 tcaaagcgat agtgaaggac agtgatggac agccgacggc agttgggatt cgtgaattgc
     4981 tgccctctgg ttatgtgtgg gagggctaag cacttcgtgg ccgaggagca ggactgacac
     5041 gtgctacgag atttcgattc caccgccgcc ttctatgaaa ggttgggctt cggaatcgtt
     5101 ttccgggacg ccggctggat gatcctccag cgcggggatc tcatgctgga gttcttcgcc
     5161 caccccaact tgtttattgc agcttataat ggttacaaat aaagcaatag catcacaaat
     5221 ttcacaaata aagcattttt ttcactgcat tctagttgtg gtttgtccaa actcatcaat
     5281 gtatcttatc atgtctgtat accgtcgacc tctagctaga gcttggcgta atcatggtca
     5341 tagctgtttc ctgtgtgaaa ttgttatccg ctcacaattc cacacaacat acgagccgga
     5401 agcataaagt gtaaagcctg gggtgcctaa tgagtgagct aactcacatt aattgcgttg
     5461 cgctcactgc ccgctttcca gtcgggaaac ctgtcgtgcc agctgcatta atgaatcggc
     5521 caacgcgcgg ggagaggcgg tttgcgtatt gggcgctctt ccgcttcctc gctcactgac
     5581 tcgctgcgct cggtcgttcg gctgcggcga gcggtatcag ctcactcaaa ggcggtaata
     5641 cggttatcca cagaatcagg ggataacgca ggaaagaaca tgtgagcaaa aggccagcaa
     5701 aaggccagga accgtaaaaa ggccgcgttg ctggcgtttt tccataggct ccgcccccct
     5761 gacgagcatc acaaaaatcg acgctcaagt cagaggtggc gaaacccgac aggactataa
     5821 agataccagg cgtttccccc tggaagctcc ctcgtgcgct ctcctgttcc gaccctgccg
     5881 cttaccggat acctgtccgc ctttctccct tcgggaagcg tggcgctttc tcaatgctca
     5941 cgctgtaggt atctcagttc ggtgtaggtc gttcgctcca agctgggctg tgtgcacgaa
     6001 ccccccgttc agcccgaccg ctgcgcctta tccggtaact atcgtcttga gtccaacccg
     6061 gtaagacacg acttatcgcc actggcagca gccactggta acaggattag cagagcgagg
     6121 tatgtaggcg gtgctacaga gttcttgaag tggtggccta actacggcta cactagaagg
     6181 acagtatttg gtatctgcgc tctgctgaag ccagttacct tcggaaaaag agttggtagc
     6241 tcttgatccg gcaaacaaac caccgctggt agcggtggtt tttttgtttg caagcagcag
     6301 attacgcgca gaaaaaaagg atctcaagaa gatcctttga tcttttctac ggggtctgac
     6361 gctcagtgga acgaaaactc acgttaaggg attttggtca tgagattatc aaaaaggatc
     6421 ttcacctaga tccttttaaa ttaaaaatga agttttaaat caatctaaag tatatatgag
     6481 taaacttggt ctgacagtta ccaatgctta atcagtgagg cacctatctc agcgatctgt
     6541 ctatttcgtt catccatagt tgcctgactc cccgtcgtgt agataactac gatacgggag
     6601 ggcttaccat ctggccccag tgctgcaatg ataccgcgag acccacgctc accggctcca
     6661 gatttatcag caataaacca gccagccgga agggccgagc gcagaagtgg tcctgcaact
     6721 ttatccgcct ccatccagtc tattaattgt tgccgggaag ctagagtaag tagttcgcca
     6781 gttaatagtt tgcgcaacgt tgttgccatt gctacaggca tcgtggtgtc acgctcgtcg
     6841 tttggtatgg cttcattcag ctccggttcc caacgatcaa ggcgagttac atgatccccc
     6901 atgttgtgca aaaaagcggt tagctccttc ggtcctccga tcgttgtcag aagtaagttg
     6961 gccgcagtgt tatcactcat ggttatggca gcactgcata attctcttac tgtcatgcca
     7021 tccgtaagat gcttttctgt gactggtgag tactcaacca agtcattctg agaatagtgt
     7081 atgcggcgac cgagttgctc ttgcccggcg tcaatacggg ataataccgc gccacatagc
     7141 agaactttaa aagtgctcat cattggaaaa cgttcttcgg ggcgaaaact ctcaaggatc
     7201 ttaccgctgt tgagatccag ttcgatgtaa cccactcgtg cacccaactg atcttcagca
     7261 tcttttactt tcaccagcgt ttctgggtga gcaaaaacag gaaggcaaaa tgccgcaaaa
     7321 aagggaataa gggcgacacg gaaatgttga atactcatac tcttcctttt tc
//
