LOCUS       Exported                6300 bp ds-DNA     circular SYN 27-OCT-2025
DEFINITION  .
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6300)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Monday, Oct 27, 2025 from SnapGene 5.0.8
            https://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..6300
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     enhancer        235..614
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
                     /note="color: #ffffff"
     promoter        615..818
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
                     /note="color: #ffffff; direction: RIGHT"
     promoter        863..881
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
                     /note="color: #ffffff; direction: RIGHT"
     misc_feature    863..879
                     /label=T7 promoter primer
                     /note="color: #00ff00"
     regulatory      901..910
                     /regulatory_class="other"
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /note="color: #a6acb3"
     misc_feature    922..939
                     /label=His Tag
                     /note="color: #8080ff"
     CDS             922..939
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /note="color: #cc99b2"
                     /translation="HHHHHH"
     misc_feature    946..951
                     /label=NheI
                     /note="color: #ff00ff"
     misc_feature    952..990
                     /label=LAP
                     /note="color: #00ffff"
     misc_feature    993..998
                     /label=BamhI
                     /note="color: #0000ff"
     misc_feature    993..998
                     /label=BamHI
                     /note="color: #ff00ff"
     misc_feature    1006..1722
                     /label=BFP
                     /note="color: #0080ff"
     CDS             1006..1722
                     /codon_start=1
                     /product="blue variant of GFP"
                     /label=BFP
                     /note="mammalian codon-optimized"
                     /note="This feature has 3 segments:
                        1: 1006 .. 1008 / #3366ff
                        2: 1009 .. 1011 / #3366ff / 1a
                        3: 1012 .. 1722 / #3366ff"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLSHGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNFNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDSHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     misc_feature    1723..1788
                     /label=PM
                     /note="color: #808000"
     misc_feature    1723..1788
                     /label=Membrane targeting sequence
                     /note="color: #ffff80"
     misc_feature    1789..1791
                     /label=STOP codon
                     /note="color: #ff0000"
     misc_feature    1792..1797
                     /label=EcoRI
                     /note="color: #ff00ff"
     misc_feature    1825..1830
                     /label=XhoI
                     /note="color: #ff00ff"
     promoter        complement(1852..1870)
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
                     /note="color: #ffffff; direction: LEFT"
     polyA_signal    1896..2120
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
                     /note="color: #a6acb3"
     rep_origin      2166..2594
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
                     /note="color: #ffff00"
     promoter        2608..2938
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
                     /note="color: #ffffff; direction: RIGHT"
     rep_origin      2789..2924
                     /label=SV40 ori
                     /note="SV40 origin of replication"
                     /note="color: #ffff00"
     CDS             3005..3799
                     /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))"
                     /note="color: #ccffcc"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     polyA_signal    3973..4094
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
                     /note="color: #a6acb3"
     primer_bind     complement(4143..4159)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
                     /note="color: #a020f0; direction: LEFT"
     protein_bind    4167..4183
                     /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)."
                     /note="color: #31849b"
     promoter        complement(4191..4221)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
                     /note="This reverse directional feature has 3 segments:
                        1: 4191 .. 4197 / #ffffff / -10
                        2: 4198 .. 4215 / #ffffff
                        3: 4216 .. 4221 / #ffffff / -35"
     protein_bind    4236..4257
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
                     /note="color: #31849b"
     rep_origin      complement(4545..5133)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
                     /note="color: #ffff00"
     CDS             complement(5304..6164)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /note="This feature has 2 segments:
                        1: 5304 .. 6095 / #ccffcc
                        2: 6096 .. 6164 / #ccffcc / signal sequence
                     Cleavage site after base 6095"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     promoter        complement(6165..6269)
                     /gene="bla"
                     /label=AmpR promoter
                     /note="color: #ffffff; direction: LEFT"
ORIGIN
        1 gacggatcgg gagatctccc gatcccctat ggtcgactct cagtacaatc tgctctgatg
       61 ccgcatagtt aagccagtat ctgctccctg cttgtgtgtt ggaggtcgct gagtagtgcg
      121 cgagcaaaat ttaagctaca acaaggcaag gcttgaccga caattgcatg aagaatctgc
      181 ttagggttag gcgttttgcg ctgcttcgcg atgtacgggc cagatatacg cgttgacatt
      241 gattattgac tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata
      301 tggagttccg cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc
      361 cccgcccatt gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc
      421 attgacgtca atgggtggac tatttacggt aaactgccca cttggcagta catcaagtgt
      481 atcatatgcc aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt
      541 atgcccagta catgacctta tgggactttc ctacttggca gtacatctac gtattagtca
      601 tcgctattac catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg
      661 actcacgggg atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc
      721 aaaatcaacg ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg
      781 gtaggcgtgt acggtgggag gtctatataa gcagagctct ctggctaact agagaaccca
      841 ctgcttactg gcttatcgaa attaatacga ctcactatag ggagacccaa gcttgcggcc
      901 gccaccatgg tgcggggttc tcatcatcat catcatcatg gtatggctag cggcttcgag
      961 atcgacaagg tgtggtacga cctggacgcc aaggatccga gctcgatggt gagcaagggc
     1021 gaggagctgt tcaccggggt ggtgcccatc ctggtcgagc tggacggcga cgtaaacggc
     1081 cacaagttca gcgtgtccgg cgagggcgag ggcgatgcca cctacggcaa gctgaccctg
     1141 aagttcatct gcaccaccgg caagctgccc gtgccctggc ccaccctcgt gaccaccctg
     1201 agccacggcg tccagtgctt cagccgctac cccgaccaca tgaagcagca cgacttcttc
     1261 aagtccgcca tgcccgaagg ctacgtccag gagcgcacca tcttcttcaa ggacgacggc
     1321 aactacaaga cccgcgccga ggtgaagttc gagggcgaca ccctggtgaa ccgcatcgag
     1381 ctgaagggca tcgacttcaa ggaggacggc aacatcctgg ggcacaagct ggagtacaac
     1441 ttcaacagcc acaacgtcta tatcatggcc gacaagcaga agaacggcat caaggtgaac
     1501 ttcaagatcc gccacaacat cgaggacggc agcgtgcagc tcgccgacca ctaccagcag
     1561 aacaccccca tcggcgacgg ccccgtgctg ctgcccgaca gccactacct gagcacccag
     1621 tccgccctga gcaaagaccc caacgagaag cgcgatcaca tggtcctgct ggagttcgtg
     1681 accgccgccg ggatcactct cggcatggac gagctgtaca agcgcagcaa gctgaacccc
     1741 cccgacgaga gcggccccgg ctgcatgagc tgcaagtgcg tgctgagcta agaattctgc
     1801 agatatccat cacactggcg gccgctcgag catgcatcta gagggcccta ttctatagtg
     1861 tcacctaaat gctagagctc gctgatcagc ctcgactgtg ccttctagtt gccagccatc
     1921 tgttgtttgc ccctcccccg tgccttcctt gaccctggaa ggtgccactc ccactgtcct
     1981 ttcctaataa aatgaggaaa ttgcatcgca ttgtctgagt aggtgtcatt ctattctggg
     2041 gggtggggtg gggcaggaca gcaaggggga ggattgggaa gacaatagca ggcatgctgg
     2101 ggatgcggtg ggctctatgg cttctgaggc ggaaagaacc agctggggct ctagggggta
     2161 tccccacgcg ccctgtagcg gcgcattaag cgcggcgggt gtggtggtta cgcgcagcgt
     2221 gaccgctaca cttgccagcg ccctagcgcc cgctcctttc gctttcttcc cttcctttct
     2281 cgccacgttc gccggctttc cccgtcaagc tctaaatcgg ggcatccctt tagggttccg
     2341 atttagtgct ttacggcacc tcgaccccaa aaaacttgat tagggtgatg gttcacgtag
     2401 tgggccatcg ccctgataga cggtttttcg ccctttgacg ttggagtcca cgttctttaa
     2461 tagtggactc ttgttccaaa ctggaacaac actcaaccct atctcggtct attcttttga
     2521 tttataaggg attttgggga tttcggccta ttggttaaaa aatgagctga tttaacaaaa
     2581 atttaacgcg aattaattct gtggaatgtg tgtcagttag ggtgtggaaa gtccccaggc
     2641 tccccaggca ggcagaagta tgcaaagcat gcatctcaat tagtcagcaa ccaggtgtgg
     2701 aaagtcccca ggctccccag caggcagaag tatgcaaagc atgcatctca attagtcagc
     2761 aaccatagtc ccgcccctaa ctccgcccat cccgccccta actccgccca gttccgccca
     2821 ttctccgccc catggctgac taattttttt tatttatgca gaggccgagg ccgcctctgc
     2881 ctctgagcta ttccagaagt agtgaggagg cttttttgga ggcctaggct tttgcaaaaa
     2941 gctcccggga gcttgtatat ccattttcgg atctgatcaa gagacaggat gaggatcgtt
     3001 tcgcatgatt gaacaagatg gattgcacgc aggttctccg gccgcttggg tggagaggct
     3061 attcggctat gactgggcac aacagacaat cggctgctct gatgccgccg tgttccggct
     3121 gtcagcgcag gggcgcccgg ttctttttgt caagaccgac ctgtccggtg ccctgaatga
     3181 actgcaggac gaggcagcgc ggctatcgtg gctggccacg acgggcgttc cttgcgcagc
     3241 tgtgctcgac gttgtcactg aagcgggaag ggactggctg ctattgggcg aagtgccggg
     3301 gcaggatctc ctgtcatctc accttgctcc tgccgagaaa gtatccatca tggctgatgc
     3361 aatgcggcgg ctgcatacgc ttgatccggc tacctgccca ttcgaccacc aagcgaaaca
     3421 tcgcatcgag cgagcacgta ctcggatgga agccggtctt gtcgatcagg atgatctgga
     3481 cgaagagcat caggggctcg cgccagccga actgttcgcc aggctcaagg cgcgcatgcc
     3541 cgacggcgag gatctcgtcg tgacccatgg cgatgcctgc ttgccgaata tcatggtgga
     3601 aaatggccgc ttttctggat tcatcgactg tggccggctg ggtgtggcgg accgctatca
     3661 ggacatagcg ttggctaccc gtgatattgc tgaagagctt ggcggcgaat gggctgaccg
     3721 cttcctcgtg ctttacggta tcgccgctcc cgattcgcag cgcatcgcct tctatcgcct
     3781 tcttgacgag ttcttctgag cgggactctg gggttcgaaa tgaccgacca agcgacgccc
     3841 aacctgccat cacgagattt cgattccacc gccgccttct atgaaaggtt gggcttcgga
     3901 atcgttttcc gggacgccgg ctggatgatc ctccagcgcg gggatctcat gctggagttc
     3961 ttcgcccacc ccaacttgtt tattgcagct tataatggtt acaaataaag caatagcatc
     4021 acaaatttca caaataaagc atttttttca ctgcattcta gttgtggttt gtccaaactc
     4081 atcaatgtat cttatcatgt ctgtataccg tcgacctcta gctagagctt ggcgtaatca
     4141 tggtcatagc tgtttcctgt gtgaaattgt tatccgctca caattccaca caacatacga
     4201 gccggaagca taaagtgtaa agcctggggt gcctaatgag tgagctaact cacattaatt
     4261 gcgttgcgct cactgcccgc tttccagtcg ggaaacctgt cgtgccagct gcattaatga
     4321 atcggccaac gcgcggggag aggcggtttg cgtattgggc gctcttccgc ttcctcgctc
     4381 actgactcgc tgcgctcggt cgttcggctg cggcgagcgg tatcagctca ctcaaaggcg
     4441 gtaatacggt tatccacaga atcaggggat aacgcaggaa agaacatgtg agcaaaaggc
     4501 cagcaaaagg ccaggaaccg taaaaaggcc gcgttgctgg cgtttttcca taggctccgc
     4561 ccccctgacg agcatcacaa aaatcgacgc tcaagtcaga ggtggcgaaa cccgacagga
     4621 ctataaagat accaggcgtt tccccctgga agctccctcg tgcgctctcc tgttccgacc
     4681 ctgccgctta ccggatacct gtccgccttt ctcccttcgg gaagcgtggc gctttctcaa
     4741 tgctcacgct gtaggtatct cagttcggtg taggtcgttc gctccaagct gggctgtgtg
     4801 cacgaacccc ccgttcagcc cgaccgctgc gccttatccg gtaactatcg tcttgagtcc
     4861 aacccggtaa gacacgactt atcgccactg gcagcagcca ctggtaacag gattagcaga
     4921 gcgaggtatg taggcggtgc tacagagttc ttgaagtggt ggcctaacta cggctacact
     4981 agaaggacag tatttggtat ctgcgctctg ctgaagccag ttaccttcgg aaaaagagtt
     5041 ggtagctctt gatccggcaa acaaaccacc gctggtagcg gtggtttttt tgtttgcaag
     5101 cagcagatta cgcgcagaaa aaaaggatct caagaagatc ctttgatctt ttctacgggg
     5161 tctgacgctc agtggaacga aaactcacgt taagggattt tggtcatgag attatcaaaa
     5221 aggatcttca cctagatcct tttaaattaa aaatgaagtt ttaaatcaat ctaaagtata
     5281 tatgagtaaa cttggtctga cagttaccaa tgcttaatca gtgaggcacc tatctcagcg
     5341 atctgtctat ttcgttcatc catagttgcc tgactccccg tcgtgtagat aactacgata
     5401 cgggagggct taccatctgg ccccagtgct gcaatgatac cgcgagaccc acgctcaccg
     5461 gctccagatt tatcagcaat aaaccagcca gccggaaggg ccgagcgcag aagtggtcct
     5521 gcaactttat ccgcctccat ccagtctatt aattgttgcc gggaagctag agtaagtagt
     5581 tcgccagtta atagtttgcg caacgttgtt gccattgcta caggcatcgt ggtgtcacgc
     5641 tcgtcgtttg gtatggcttc attcagctcc ggttcccaac gatcaaggcg agttacatga
     5701 tcccccatgt tgtgcaaaaa agcggttagc tccttcggtc ctccgatcgt tgtcagaagt
     5761 aagttggccg cagtgttatc actcatggtt atggcagcac tgcataattc tcttactgtc
     5821 atgccatccg taagatgctt ttctgtgact ggtgagtact caaccaagtc attctgagaa
     5881 tagtgtatgc ggcgaccgag ttgctcttgc ccggcgtcaa tacgggataa taccgcgcca
     5941 catagcagaa ctttaaaagt gctcatcatt ggaaaacgtt cttcggggcg aaaactctca
     6001 aggatcttac cgctgttgag atccagttcg atgtaaccca ctcgtgcacc caactgatct
     6061 tcagcatctt ttactttcac cagcgtttct gggtgagcaa aaacaggaag gcaaaatgcc
     6121 gcaaaaaagg gaataagggc gacacggaaa tgttgaatac tcatactctt cctttttcaa
     6181 tattattgaa gcatttatca gggttattgt ctcatgagcg gatacatatt tgaatgtatt
     6241 tagaaaaata aacaaatagg ggttccgcgc acatttcccc gaaaagtgcc acctgacgtc
//
