LOCUS       Exported                4975 bp ds-DNA     circular SYN 27-OCT-2025
DEFINITION  synthetic circular DNA
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4975)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Monday, Oct 27, 2025 from SnapGene 5.0.8
            https://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..4975
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     CDS             complement(336..1196)
                     /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(386..405)
                     /label=AmpStop
     primer_bind     1132..1151
                     /label=AmpStart
     promoter        complement(1197..1301)
                     /gene="bla"
                     /label=AmpR promoter
     gap             1238..1266
                     /label=amp prom|Unknown
     rep_origin      complement(1346..1801)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     rep_origin      complement(1478..1784)
                     /direction=LEFT
                     /label=F1 ori
     CDS             complement(1803..1871)
                     /label=LacZ alpha
     primer_bind     1891..1910
                     /label=pUCM13-52
     primer_bind     1923..1939
                     /label=M13-40
     primer_bind     1942..1959
                     /label=M13-fwd
     primer_bind     1942..1958
                     /label=M13
     primer_bind     1943..1959
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     1968..1988
                     /label=T7
     primer_bind     1969..1988
                     /label=T7
     promoter        1969..1987
                     /label=T7_promoter Promoter|promoter
     primer_bind     1969..1987
                     /label=Short-T7
     misc_feature    2012..2390
                     /label=P_TEF1
     primer_bind     2012..2030
                     /label=o5639
     primer_bind     2012..2030
                     /label=o5418
     primer_bind     2042..2061
                     /label=FRO351
     misc_feature    2046..2385
                     /label=pTEF-short
     gene            2047..3166
                     /label=natMX6
                     /note="yeast selectable marker conferring nourseothricin 
                     resistance"
     promoter        2047..2390
                     /label=TEF promoter
                     /note="Ashbya gossypii TEF promoter"
     misc_feature    2289..2296
                     /label=TATAbox
     primer_bind     2361..2390
                     /label=FRO4538
     primer_bind     complement(2361..2390)
                     /label=FRO4540
     primer_bind     2371..2411
                     /label=FRP373 FWD
     misc_feature    2391..2963
                     /label=NatMX
     misc_feature    2391..2963
                     /label=Resistance Marker
     CDS             2391..2963
                     /codon_start=1
                     /gene="Streptomyces noursei nat1"
                     /product="nourseothricin acetyltransferase"
                     /label=NrsR
                     /note="confers resistance to nourseothricin"
                     /translation="MGTTLDDTAYRYRTSVPGDAEAIEALDGSFTTDTVFRVTATGDGF
                     TLREVPVDPPLTKVFPDDESDDESDDGEDGDPDSRTFVAYGDDGDLAGFVVVSYSGWNR
                     RLTVEDIEVAPEHRGHGVGRALMGLATEFARERGAGHLWLEVTNVNAPAIHAYRRMGFT
                     LCGLDTALYDGTASDGEQALYMSMPCP"
     primer_bind     complement(2478..2497)
                     /label=FRO1851
     misc_feature    2964..3200
                     /label=term_TEF1
     misc_feature    2964..3164
                     /label=TermTef-Short
     primer_bind     2964..2993
                     /label=FRO4541
     primer_bind     complement(2964..2993)
                     /label=FRO4539
     terminator      2969..3166
                     /label=TEF terminator
                     /note="Ashbya gossypii TEF terminator"
     primer_bind     complement(3178..3201)
                     /label=FRO4760
     primer_bind     complement(3199..3222)
                     /label=FRO4299
     primer_bind     3202..3246
                     /label=FRO4300
     primer_bind     3220..3250
                     /label=FRO4770
     primer_bind     3222..3250
                     /label=o5604
     misc_feature    3223..3957
                     /label=P4Lacn.2
     misc_feature    3223..3956
                     /label=P4Lacn.2
     primer_bind     3223..3250
                     /label=FRO5081
     primer_bind     3223..3250
                     /label=FRO4507
     primer_bind     3223..3250
                     /label=O5766
     primer_bind     complement(3228..3249)
                     /label=FRO4368
     primer_bind     complement(3235..3254)
                     /label=FRO4445
     misc_feature    3321..3372
                     /label=duplication
     misc_feature    3336..3347
                     /label=RAP1binding
     misc_feature    3362..3372
                     /label=GCR1binding
     misc_feature    3384..3393
                     /label=grf2_binding
     misc_feature    3414..3424
                     /label=grf2_binding
     misc_feature    3443..3454
                     /label=RAP1binding
     misc_feature    3469..3479
                     /label=GCR1binding
     misc_feature    3507..3517
                     /label=GCR1binding
     primer_bind     3730..3753
                     /label=o5203
     primer_bind     complement(3730..3753)
                     /label=o5202
     primer_bind     complement(3775..3834)
                     /label=o5240
     misc_feature    3795..3813
                     /label=lacop
     protein_bind    3797..3813
                     /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)."
     primer_bind     complement(3799..3813)
                     /label=FRO2169
     primer_bind     3805..3864
                     /label=o5241
     misc_feature    3816..3823
                     /label=TATAbox
     primer_bind     3825..3844
                     /label=o5204
     misc_feature    3826..3844
                     /label=lacop
     protein_bind    3826..3842
                     /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)."
     primer_bind     complement(3828..3844)
                     /label=FRO2169
     misc_feature    complement(3879..3886)
                     /label=TATAbox
     primer_bind     complement(3903..3958)
                     /label=o5610
     primer_bind     complement(3903..3958)
                     /label=o5609
     primer_bind     complement(3903..3958)
                     /label=o5608
     primer_bind     complement(3903..3958)
                     /label=o5605
     primer_bind     complement(3903..3957)
                     /label=O5640
     misc_feature    3916..3936
                     /label=lacop
     primer_bind     3916..3936
                     /label=o5204
     misc_feature    3916..3934
                     /label=lacop
     misc_feature    3918..3936
                     /label=lacop
     protein_bind    3918..3934
                     /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)."
     primer_bind     complement(3920..3936)
                     /label=FRO2169
     misc_feature    3937..3957
                     /label=lacop
     primer_bind     3937..3957
                     /label=o5204
     misc_feature    3937..3955
                     /label=lacop
     misc_feature    3939..3957
                     /label=lacop
     protein_bind    3939..3955
                     /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)."
     primer_bind     complement(3941..3964)
                     /label=FRO2169
     primer_bind     complement(3953..3972)
                     /label=o5335
     primer_bind     3958..3975
                     /label=o5391
     primer_bind     complement(3958..3975)
                     /label=o5390
     misc_feature    3958..3972
                     /label=good kozak
     primer_bind     3970..4035
                     /label=FRO4855
     primer_bind     3975..4011
                     /label=FRO4855
     gap             3976..4047
                     /label=FLAG TAG|Unknown
     CDS             3976..4047
                     /codon_start=1
                     /product="three tandem FLAG(R) epitope tags"
                     /label=3xFLAG
                     /translation="DYKDDDDKDYKDDDDKDYKDDDDK"
     primer_bind     complement(3985..4014)
                     /label=FRO4412
     primer_bind     complement(3985..4004)
                     /label=FRO4408
     primer_bind     complement(4009..4038)
                     /label=FRO4412
     primer_bind     complement(4009..4028)
                     /label=FRO4408
     primer_bind     complement(4016..4071)
                     /label=FRO4856
     primer_bind     4022..4112
                     /label=FRO4298
     primer_bind     complement(4033..4048)
                     /label=FRO4408
     primer_bind     complement(4033..4048)
                     /label=FRO4412
     gap             4048..4071
                     /label=8G linker|Unknown
     primer_bind     complement(4072..4095)
                     /label=FRO4532
     primer_bind     4072..4092
                     /label=S4 primer site|primer_bind
     primer_bind     complement(4093..4117)
                     /label=FRO4432
     primer_bind     complement(4109..4128)
                     /label=T3
     primer_bind     complement(4110..4129)
                     /label=FRO0114
     promoter        complement(4111..4129)
                     /label=T3 promoter
     primer_bind     complement(4111..4127)
                     /label=T3
     primer_bind     complement(4146..4166)
                     /label=M13-rev
     primer_bind     complement(4150..4166)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4150..4166)
                     /label=M13r
     primer_bind     4164..4188
                     /label=FRO2169
     primer_bind     complement(4164..4183)
                     /label=LacOp-for
     misc_binding    complement(4172..4194)
                     /label=LacO
     misc_feature    complement(4172..4192)
                     /label=lacop
     primer_bind     complement(4172..4192)
                     /label=o5204
     misc_feature    complement(4172..4190)
                     /label=lacop
     misc_feature    complement(4174..4192)
                     /label=lacop
     protein_bind    4174..4190
                     /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(4198..4228)
                     /label=lac promoter
     primer_bind     complement(4198..4217)
                     /label=pUCM13-rev-157
     protein_bind    4243..4264
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     rep_origin      complement(join(4534..4975,1..241))
                     /direction=LEFT
                     /label=ColE1 origin
ORIGIN
        1 tacggctaca ctagaaggac agtatttggt atctgcgctc tgctgaagcc agttaccttc
       61 ggaaaaagag ttggtagctc ttgatccggc aaacaaacca ccgctggtag cggtggtttt
      121 tttgtttgca agcagcagat tacgcgcaga aaaaaaggat ctcaagaaga tcctttgatc
      181 ttttctacgg ggtctgacgc tcagtggaac gaaaactcac gttaagggat tttggtcatg
      241 agattatcaa aaaggatctt cacctagatc cttttaaatt aaaaatgaag ttttaaatca
      301 atctaaagta tatatgagta aacttggtct gacagttacc aatgcttaat cagtgaggca
      361 cctatctcag cgatctgtct atttcgttca tccatagttg cctgactccc cgtcgtgtag
      421 ataactacga tacgggaggg cttaccatct ggccccagtg ctgcaatgat accgcgagac
      481 ccacgctcac cggctccaga tttatcagca ataaaccagc cagccggaag ggccgagcgc
      541 agaagtggtc ctgcaacttt atccgcctcc atccagtcta ttaattgttg ccgggaagct
      601 agagtaagta gttcgccagt taatagtttg cgcaacgttg ttgccattgc tacaggcatc
      661 gtggtgtcac gctcgtcgtt tggtatggct tcattcagct ccggttccca acgatcaagg
      721 cgagttacat gatcccccat gttgtgcaaa aaagcggtta gctccttcgg tcctccgatc
      781 gttgtcagaa gtaagttggc cgcagtgtta tcactcatgg ttatggcagc actgcataat
      841 tctcttactg tcatgccatc cgtaagatgc ttttctgtga ctggtgagta ctcaaccaag
      901 tcattctgag aatagtgtat gcggcgaccg agttgctctt gcccggcgtc aatacgggat
      961 aataccgcgc cacatagcag aactttaaaa gtgctcatca ttggaaaacg ttcttcgggg
     1021 cgaaaactct caaggatctt accgctgttg agatccagtt cgatgtaacc cactcgtgca
     1081 cccaactgat cttcagcatc ttttactttc accagcgttt ctgggtgagc aaaaacagga
     1141 aggcaaaatg ccgcaaaaaa gggaataagg gcgacacgga aatgttgaat actcatactc
     1201 ttcctttttc aatattattg aagcatttat cagggttatt gtctcatgag cggatacata
     1261 tttgaatgta tttagaaaaa taaacaaata ggggttccgc gcacatttcc ccgaaaagtg
     1321 ccaccaatac ccgactctat atgctaaatt gtaagcgtta atattttgtt aaaattcgcg
     1381 ttaaattttt gttaaatcag ctcatttttt aaccaatagg ccgaaatcgg caaaatccct
     1441 tataaatcaa aagaatagac cgagataggg ttgagtgttg ttccagtttg gaacaagagt
     1501 ccactattaa agaacgtgga ctccaacgtc aaagggcgaa aaaccgtcta tcagggcgat
     1561 ggcccactac gtgaaccatc accctaatca agttttttgg ggtcgaggtg ccgtaaagca
     1621 ctaaatcgga accctaaagg gagcccccga tttagagctt gacggggaaa gccggcgaac
     1681 gtggcgagaa aggaagggaa gaaagcgaaa ggagcgggcg ctagggcgct ggcaagtgta
     1741 gcggtcacgc tgcgcgtaac caccacaccc gccgcgctta atgcgccgct acagggcgcg
     1801 tcccattcgc cattcaggct gcgcaactgt tgggaagggc gatcggtgcg ggcctcttcg
     1861 ctattacgcc agctggcgaa agggggatgt gctgcaaggc gattaagttg ggtaacgcca
     1921 gggttttccc agtcacgacg ttgtaaaacg acggccagtg agcgcgcgta atacgactca
     1981 ctatagggcg aattggagct cgatcctcga gagcttgcct tgtccccgcc gggtcacccg
     2041 gccagcgaca tggaggccca gaataccctc cttgacagtc ttgacgtgcg cagctcaggg
     2101 gcatgatgtg actgtcgccc gtacatttag cccatacatc cccatgtata atcatttgca
     2161 tccatacatt ttgatggccg cacggcgcga agcaaaaatt acggctcctc gctgcggacc
     2221 tgcgagcagg gaaacgctcc cctcacagac gcgttgaatt gtccccacgc cgcgcccctg
     2281 tagagaaata taaaaggtta ggatttgcca ctgaggttct tctttcatat acttcctttt
     2341 aaaatcttgc taggatacag ttctcacatc acatccgaac ataaacaacc atgggtacca
     2401 ctcttgacga cacggcttac cggtaccgca ccagtgtccc gggggacgcc gaggccatcg
     2461 aggcactgga tgggtccttc accaccgaca ccgtcttccg cgtcaccgcc accggggacg
     2521 gcttcaccct gcgggaggtg ccggtggacc cgcccctgac caaggtgttc cccgacgacg
     2581 aatcggacga cgaatcggac gacggggagg acggcgaccc ggactcccgg acgttcgtcg
     2641 cgtacgggga cgacggcgac ctggcgggct tcgtggtcgt ctcgtactcc ggctggaacc
     2701 gccggctgac cgtcgaggac atcgaggtcg ccccggagca ccgggggcac ggggtcgggc
     2761 gcgcgttgat ggggctcgcg acggagttcg cccgcgagcg gggcgccggg cacctctggc
     2821 tggaggtcac caacgtcaac gcaccggcga tccacgcgta ccggcggatg gggttcaccc
     2881 tctgcggcct ggacaccgcc ctgtacgacg gcaccgcctc ggacggcgag caggcgctct
     2941 acatgagcat gccctgcccc taatcagtac tgacaataaa aagattcttg ttttcaagaa
     3001 cttgtcattt gtatagtttt tttatattgt agttgttcta ttttaatcaa atgttagcgt
     3061 gatttatatt ttttttcgcc tcgacatcat ctgcccagat gcgaagttaa gtgcgcagaa
     3121 agtaatatca tgcgtcaatc gtatgtgaat gctggtcgct atactgctgt cgattcgata
     3181 ctaacgccgc catccagtgt ttaaacgaac tagtgcggcc gccagttcga gtttatcatt
     3241 atcaatactg ccatttcaaa gaatacgtaa ataattaata gtagtgattt tcctaacttt
     3301 atttagtcaa aaaattagcc tatataacat cgtaggtgtc tgggtgaaca gtttattcct
     3361 ggcatccact aattttaatt ctgctgtaac ccgtacatgc ccaaaatagg gggcgggtta
     3421 cacagaatat ataacatcgt aggtgtctgg gtgaacagtt tattcctggc atccactaaa
     3481 tataatggag cccgcttttt aagctggcat ccactaaaaa aaagaatccc agcaccaaaa
     3541 tattgttttc ttcaccaacc atcagttcat aggtccattc tcttagcgca actacagaga
     3601 acaggggcac aaacaggcaa aaaacgggca caacctcaat ggagtgatgc aacctgcctg
     3661 gagtaaatga tgacacaagg caattgaccc acgcatgtat ctatctcatt ttcttacacc
     3721 ttctattacc ttctgctctc tctgatttgg aaaaagctga aaaaaaaggt tgaaaccagt
     3781 tccctgaaat tattaattgt gagcggataa caaagtataa atagattgtg agcggataac
     3841 aattctgtaa atctatttct taaacttctt aaattctact tttatagtta gtcttttttt
     3901 tagttttaaa acaccaattg tgagcggata acaattaatt gtgagcggat aacaattaca
     3961 cacataaaca aaatggatta taaagatgac gatgacaagg attataaaga tgacgatgac
     4021 aaggattata aagatgacga tgacaaggga ggtggtggtg gtggaggcgg tccgacagag
     4081 aattcatcga tgggtaccca gcttttgttc cctttagtga gggttaattg cgcgcttggc
     4141 gtaatcatgg tcatagctgt ttcctgtgtg aaattgttat ccgctcacaa ttccacacaa
     4201 catacgagcc ggaagcataa agtgtaaagc ctggggtgcc taatgagtga gctaactcac
     4261 attaattgcg ttgcgctcac tgcccgcttt ccagtcggga aacctgtcgt gccagctgca
     4321 ttaatgaatc ggccaacgcg cggggagagg cggtttgcgt attgggcgct cttccgcttc
     4381 ctcgctcact gactcgctgc gctcggtcgt tcggctgcgg cgagcggtat cagctcactc
     4441 aaaggcggta atacggttat ccacagaatc aggggataac gcaggaaaga acatgtgagc
     4501 aaaaggccag caaaaggcca ggaaccgtaa aaaggccgcg ttgctggcgt ttttccatag
     4561 gctccgcccc cctgacgagc atcacaaaaa tcgacgctca agtcagaggt ggcgaaaccc
     4621 gacaggacta taaagatacc aggcgtttcc ccctggaagc tccctcgtgc gctctcctgt
     4681 tccgaccctg ccgcttaccg gatacctgtc cgcctttctc ccttcgggaa gcgtggcgct
     4741 ttctcatagc tcacgctgta ggtatctcag ttcggtgtag gtcgttcgct ccaagctggg
     4801 ctgtgtgcac gaaccccccg ttcagcccga ccgctgcgcc ttatccggta actatcgtct
     4861 tgagtccaac ccggtaagac acgacttatc gccactggca gcagccactg gtaacaggat
     4921 tagcagagcg aggtatgtag gcggtgctac agagttcttg aagtggtggc ctaac
//
