LOCUS       Exported                6275 bp ds-DNA     circular SYN 26-FEB-2024
DEFINITION  synthetic circular DNA
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6275)
  AUTHORS   Manuel Iburg
  TITLE     Direct Submission
  JOURNAL   Exported Thursday, Jul 18, 2024 from SnapGene 5.0.8
            https://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..6275
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     rep_origin      13..468
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(478..497)
                     /label=148 SpecRpCDF rev
     CDS             complement(561..1352)
                     /codon_start=1
                     /gene="aadA"
                     /product="aminoglycoside adenylyltransferase (Murphy, 
                     1985)"
                     /label=SmR
                     /note="confers resistance to spectinomycin and 
                     streptomycin"
                     /translation="MREAVIAEVSTQLSEVVGVIERHLEPTLLAVHLYGSAVDGGLKPH
                     SDIDLLVTVTVRLDETTRRALINDLLETSASPGESEILRAVEVTIVVHDDIIPWRYPAK
                     RELQFGEWQRNDILAGIFEPATIDIDLAILLTKAREHSVALVGPAAEELFDPVPEQDLF
                     EALNETLTLWNSPPDWAGDERNVVLTLSRIWYSAVTGKIAPKDVAADWAMERLPAQYQP
                     VILEARQAYLGQEEDRLASRADQLEEFVHYVKGEITKVVGK"
     rep_origin      1459..2276
                     /direction=RIGHT
                     /label=CloDF13 ori
                     /note="Plasmids containing the CloDF13 (CDF) origin of 
                     replication can be propagated in E. coli cells that contain
                     additional plasmids with compatible origins."
     rep_origin      1474..1488
                     /direction=RIGHT
                     /label=CloDF13 ori
                     /note="Plasmids containing the CloDF13 (CDF) origin of 
                     replication can be propagated in E. coli cells that contain
                     additional plasmids with compatible origins."
     rep_origin      1538..1601
                     /direction=RIGHT
                     /label=CloDF13 ori
                     /note="Plasmids containing the CloDF13 (CDF) origin of 
                     replication can be propagated in E. coli cells that contain
                     additional plasmids with compatible origins."
     primer_bind     2381..2402
                     /label=148 SpecRpCDF for
     misc_feature    2462..2601
                     /label=bom
                     /note="basis of mobility region from pBR322"
     CDS             complement(2703..2894)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     CDS             complement(3703..4785)
                     /codon_start=1
                     /gene="lacI"
                     /product="lac repressor"
                     /label=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)."
                     /translation="MKPVTLYDVAEYAGVSYQTVSRVVNQASHVSAKTREKVEAAMAEL
                     NYIPNRVAQQLAGKQSLLIGVATSSLALHAPSQIVAAIKSRADQLGASVVVSMVERSGV
                     EACKAAVHNLLAQRVSGLIINYPLDDQDAIAVEAACTNVPALFLDVSDQTPINSIIFSH
                     EDGTRLGVEHLVALGHQQIALLAGPLSSVSARLRLAGWHKYLTRNQIQPIAEREGDWSA
                     MSGFQQTMQMLNEGIVPTAMLVANDQMALGAMRAITESGLRVGADISVVGYDDTEDSSC
                     YIPPLTTIKQDFRLLGQTSVDRLLQLSQGQAVKGNQLLPVSLVKRKTTLAPNTQTASPR
                     ALADSLMQLARQVSRLESGQ"
     promoter        complement(4786..4863)
                     /gene="lacI"
                     /label=lacI promoter
     promoter        5172..5190
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    5191..5215
                     /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(5216..5298)
                     /label=Vec rev LgNterm
     primer_bind     complement(5222..5259)
                     /label=pET26-Gib2-R
                     /note="Ta = 62.5Tm = 61.5"
     primer_bind     complement(5222..5259)
                     /label=pET26b-OptHis-Gib-R
                     /note="Ta = 63.4Tm = 60.4"
     primer_bind     complement(5233..5265)
                     /label=p26-5HisMoLinker-R
                     /note="Ta = 60.4Tm = 57.4"
     primer_bind     complement(5239..5274)
                     /label=FLAG-LgBit rev
     CDS             5263..5286
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     primer_bind     5275..5307
                     /label=FLAG-LgBit for
     CDS             5287..5757
                     /codon_start=1
                     /label=NanoBiT Large Fragment
                     /note="https://www.promega.com/products/protein-purificatio
                     n-and-interactions/protein-interactions/nanobit-complementa
                     tion-assay-for-protein-interactions/nanobit-ppi-starter-sys
                     tems/?activeTab=0"
                     /translation="VFTLEDFVGDWEQTAAYNLDQVLEQGGVSSLLQNLAVSVTPIQRI
                     VRSGENALKIDIHVIIPYEGLSADQMAQIEEVFKVVYPVDDHHFKVILPYGTLVIDGVT
                     PNMLNYFGRPYEGIAVFDGKKITVTGTLWNGNKIIDERLITPDGSMLFRVTIN"
     primer_bind     5287..5309
                     /label=F1 for LgNterm
     primer_bind     5287..5307
                     /label=T-Beast-11S-IF
                     /note="Ta =59.5Tm = 58.5"
     primer_bind     complement(5735..5767)
                     /label=F1 rev LgNterm
     primer_bind     5749..5778
                     /label=Vec for LgNterm
     primer_bind     5758..5776
                     /label=T-NLink-11S-IF
                     /note="Ta = 59.5Tm = 58.5"
     primer_bind     5758..5776
                     /label=11S-Swap-VF
                     /note="Ta = 61.5Tm = 58.7"
     primer_bind     complement(5797..5819)
                     /label=Vec Luc-J r
     primer_bind     5805..5841
                     /label=Luc-J f
     CDS             5812..6111
                     /codon_start=1
                     /label=GvpJ
                     /translation="AVEHNMQSSTIVDVLEKILDKGVVIAGDITVGIADVELLTIKIRL
                     IVASVDKAKEIGMDWWENDPYLSSKGANNKALEEENKMLHERLKTLEEKIETKR"
     misc_feature    5934
                     /label=SNP
                     /note="Bacillus megaterium QM B1551 SNP"
     primer_bind     5940..5961
                     /label=Bm6seqF
     primer_bind     complement(6079..6121)
                     /label=Luc-J r
     primer_bind     6103..6128
                     /label=Vec Luc-J f
     primer_bind     6112..6129
                     /label=pBAD-ddXFP-VF
                     /note="Ta = 62.6Tm = 61.6"
     primer_bind     complement(6116..6138)
                     /label=rrnB-F1060_Seq-R
     primer_bind     complement(6118..6157)
                     /label=pBAD2pETv2
                     /note="Ta = 64Tm = 63"
     primer_bind     6138..6159
                     /label=pET26b-ChangeHis-F
                     /note="Ta = 68Tm = 65"
     primer_bind     6138..6155
                     /label=pET26-Gib2-F
                     /note="Ta = 62.5Tm = 64"
     primer_bind     6138..6155
                     /label=p26-5HisMoLinker-F
                     /note="Ta = 60.4Tm = 62"
     terminator      6204..6251
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
ORIGIN
        1 ttggcgaatg ggacgcgccc tgtagcggcg cattaagcgc ggcgggtgtg gtggttacgc
       61 gcagcgtgac cgctacactt gccagcgccc tagcgcccgc tcctttcgct ttcttccctt
      121 cctttctcgc cacgttcgcc ggctttcccc gtcaagctct aaatcggggg ctccctttag
      181 ggttccgatt tagtgcttta cggcacctcg accccaaaaa acttgattag ggtgatggtt
      241 cacgtagtgg gccatcgccc tgatagacgg tttttcgccc tttgacgttg gagtccacgt
      301 tctttaatag tggactcttg ttccaaactg gaacaacact caaccctatc tcggtctatt
      361 cttttgattt ataagggatt ttgccgattt cggcctattg gttaaaaaat gagctgattt
      421 aacaaaaatt taacgcgaat tttaacaaaa tattaacgtt tacaatttca ggtggcactt
      481 ttcggggaaa tgtgcgcgga acccctattt gtttattttt ctaaatacat tcaaatatgt
      541 atccgctcat gaattaattc ttatttgccg actaccttgg tgatctcgcc tttcacgtag
      601 tggacaaatt cttccaactg atctgcgcgc gaggccaagc gatcttcttc ttgtccaaga
      661 taagcctgtc tagcttcaag tatgacgggc tgatactggg ccggcaggcg ctccattgcc
      721 cagtcggcag cgacatcctt cggcgcgatt ttgccggtta ctgcgctgta ccaaatgcgg
      781 gacaacgtaa gcactacatt tcgctcatcg ccagcccagt cgggcggcga gttccatagc
      841 gttaaggttt catttagcgc ctcaaataga tcctgttcag gaaccggatc aaagagttcc
      901 tccgccgctg gacctaccaa ggcaacgcta tgttctcttg cttttgtcag caagatagcc
      961 agatcaatgt cgatcgtggc tggctcgaag atacctgcaa gaatgtcatt gcgctgccat
     1021 tctccaaatt gcagttcgcg cttagctgga taacgccacg gaatgatgtc gtcgtgcaca
     1081 acaatggtga cttctacagc gcggagaatc tcgctctctc caggggaagc cgaagtttcc
     1141 aaaaggtcgt tgatcaaagc tcgccgcgtt gtttcatcaa gccttacggt caccgtaacc
     1201 agcaaatcaa tatcactgtg tggcttcagg ccgccatcca ctgcggagcc gtacaaatgt
     1261 acggccagca acgtcggttc gagatggcgc tcgatgacgc caactacctc tgatagttga
     1321 gtcgatactt cggcgatcac cgcttccctc ataacacccc ttgtattact gtttatgtaa
     1381 gcagacagtt ttattgttca tgaccaaaat cccttaacgt gagttttcgt tccactgagc
     1441 gtcagacccc gtagaaaaga tcaaaggatc ttcgatcaaa ggatcttctt gagatccttt
     1501 ttttctgcgc gtaatctttt gccctgtaaa cgaaaaacca cctggggagg tggtttgatc
     1561 gaaggttaag cagttcccca actgacttaa ccttcgaaaa aaccacctgg ggaggtggtt
     1621 tgatcgaagg ttaagtcagt tggggaactg cttaaccgtg gtaactggct ttcgcagagc
     1681 acagcaacca aatctgtcct tccagtgtag ccggactttg gcgcacactt caagagcaac
     1741 cgcgtgttta gctaaacaaa tcctctgcga actcccagtt accaatggct gctgccagtg
     1801 gcgttttacc gtgcttttcc gggttggact caagtgaaca gttaccggat aaggcgcagc
     1861 agtcgggctg aacggggagt tcttgcttac agcccagctt ggagcgaacg acctacaccg
     1921 agccgagata ccagtgtgtg agctatgaga aagcgccaca cttcccgtaa gggagaaagg
     1981 cggaacaggt atccggtaaa cggcagggtc ggaacaggag agcgcaagag ggagcgaccc
     2041 gccggaaacg gtggggatct ttaagtcctg tcgggtttcg cccgtactgt cagattcatg
     2101 gttgagcctc acggctccca cagatgcacc ggaaaagcgt ctgtttatgt gaactctggc
     2161 aggagggcgg agcctatgga aaaacgccac cggcgcggcc ctgctgtttt gcctcacatg
     2221 ttagtcccct gcttatccac ggaatctgtg ggtaactttg tatgtgtccg cagcgcaacg
     2281 ccagcaacgc ggccttttta cggttcctgg ccttttgctg gccttttgct cacatgttct
     2341 ttcctgcgtt atcccctgat tctgtggata accgtattac cgcctttgag tgagctgata
     2401 ccgctcgccg cagccgaacg accgagcgca gcgagtcagt gagcgaggaa gcggaagagc
     2461 gcctgatgcg gtattttctc cttacgcatc tgtgcggtat ttcacaccgc aatggtgcac
     2521 tctcagtaca atctgctctg atgccgcata gttaagccag tatacactcc gctatcgcta
     2581 cgtgactggg tcatggctgc gccccgacac ccgccaacac ccgctgacgc gccctgacgg
     2641 gcttgtctgc tcccggcatc cgcttacaga caagctgtga ccgtctccgg gagctgcatg
     2701 tgtcagaggt tttcaccgtc atcaccgaaa cgcgcgaggc agctgcggta aagctcatca
     2761 gcgtggtcgt gaagcgattc acagatgtct gcctgttcat ccgcgtccag ctcgttgagt
     2821 ttctccagaa gcgttaatgt ctggcttctg ataaagcggg ccatgttaag ggcggttttt
     2881 tcctgtttgg tcactgatgc ctccgtgtaa gggggatttc tgttcatggg ggtaatgata
     2941 ccgatgaaac gagagaggat gctcacgata cgggttactg atgatgaaca tgcccggtta
     3001 ctggaacgtt gtgagggtaa acaactggcg gtatggatgc ggcgggacca gagaaaaatc
     3061 actcagggtc aatgccagcg cttcgttaat acagatgtag gtgttccaca gggtagccag
     3121 cagcatcctg cgatgcagat ccggaacata atggtgcagg gcgctgactt ccgcgtttcc
     3181 agactttacg aaacacggaa accgaagacc attcatgttg ttgctcaggt cgcagacgtt
     3241 ttgcagcagc agtcgcttca cgttcgctcg cgtatcggtg attcattctg ctaaccagta
     3301 aggcaacccc gccagcctag ccgggtcctc aacgacagga gcacgatcat gcgcacccgt
     3361 ggggccgcca tgccggcgat aatggcctgc ttctcgccga aacgtttggt ggcgggacca
     3421 gtgacgaagg cttgagcgag ggcgtgcaag attccgaata ccgcaagcga caggccgatc
     3481 atcgtcgcgc tccagcgaaa gcggtcctcg ccgaaaatga cccagagcgc tgccggcacc
     3541 tgtcctacga gttgcatgat aaagaagaca gtcataagtg cggcgacgat agtcatgccc
     3601 cgcgcccacc ggaaggagct gactgggttg aaggctctca agggcatcgg tcgagatccc
     3661 ggtgcctaat gagtgagcta acttacatta attgcgttgc gctcactgcc cgctttccag
     3721 tcgggaaacc tgtcgtgcca gctgcattaa tgaatcggcc aacgcgcggg gagaggcggt
     3781 ttgcgtattg ggcgccaggg tggtttttct tttcaccagt gagacgggca acagctgatt
     3841 gcccttcacc gcctggccct gagagagttg cagcaagcgg tccacgctgg tttgccccag
     3901 caggcgaaaa tcctgtttga tggtggttaa cggcgggata taacatgagc tgtcttcggt
     3961 atcgtcgtat cccactaccg agatatccgc accaacgcgc agcccggact cggtaatggc
     4021 gcgcattgcg cccagcgcca tctgatcgtt ggcaaccagc atcgcagtgg gaacgatgcc
     4081 ctcattcagc atttgcatgg tttgttgaaa accggacatg gcactccagt cgccttcccg
     4141 ttccgctatc ggctgaattt gattgcgagt gagatattta tgccagccag ccagacgcag
     4201 acgcgccgag acagaactta atgggcccgc taacagcgcg atttgctggt gacccaatgc
     4261 gaccagatgc tccacgccca gtcgcgtacc gtcttcatgg gagaaaataa tactgttgat
     4321 gggtgtctgg tcagagacat caagaaataa cgccggaaca ttagtgcagg cagcttccac
     4381 agcaatggca tcctggtcat ccagcggata gttaatgatc agcccactga cgcgttgcgc
     4441 gagaagattg tgcaccgccg ctttacaggc ttcgacgccg cttcgttcta ccatcgacac
     4501 caccacgctg gcacccagtt gatcggcgcg agatttaatc gccgcgacaa tttgcgacgg
     4561 cgcgtgcagg gccagactgg aggtggcaac gccaatcagc aacgactgtt tgcccgccag
     4621 ttgttgtgcc acgcggttgg gaatgtaatt cagctccgcc atcgccgctt ccactttttc
     4681 ccgcgttttc gcagaaacgt ggctggcctg gttcaccacg cgggaaacgg tctgataaga
     4741 gacaccggca tactctgcga catcgtataa cgttactggt ttcacattca ccaccctgaa
     4801 ttgactctct tccgggcgct atcatgccat accgcgaaag gttttgcgcc attcgatggt
     4861 gtccgggatc tcgacgctct cccttatgcg actcctgcat taggaagcag cccagtagta
     4921 ggttgaggcc gttgagcacc gccgccgcaa ggaatggtgc atgcaaggag atggcgccca
     4981 acagtccccc ggccacgggg cctgccacca tacccacgcc gaaacaagcg ctcatgagcc
     5041 cgaagtggcg agcccgatct tccccatcgg tgatgtcggc gatataggcg ccagcaaccg
     5101 cacctgtggc gccggtgatg ccggccacga tgcgtccggc gtagaggatc gagatctcga
     5161 tcccgcgaaa ttaatacgac tcactatagg ggaattgtga gcggataaca attcccctct
     5221 agaaataatt ttgtttaact ttaagaagga gatatacata tggattataa agatgatgat
     5281 gataaagtct tcacactcga agatttcgtt ggggactggg aacagacagc cgcctacaac
     5341 ctggaccaag tccttgaaca gggaggtgtg tccagtttgc tgcagaatct cgccgtgtcc
     5401 gtaactccga tccaaaggat tgtccggagc ggtgaaaatg ccctgaagat cgacatccat
     5461 gtcatcatcc cgtatgaagg tctgagcgcc gaccaaatgg cccagatcga agaggtgttt
     5521 aaggtggtgt accctgtgga tgatcatcac tttaaggtga tcctgcccta tggcacactg
     5581 gtaatcgacg gggttacgcc gaacatgctg aactatttcg gacggccgta tgaaggcatc
     5641 gccgtgttcg acggcaaaaa gatcactgta acagggaccc tgtggaacgg caacaaaatt
     5701 atcgacgagc gcctgatcac ccccgacggc tccatgctgt tccgagtaac catcaacgtt
     5761 tctcaaggca gttcaggtgg tggcgggagc ggaggtggag gctcgagcgg tgcagtcgaa
     5821 cataatatgc agtcaagtac gattgtagat gtgctcgaaa agattttgga taaaggagtc
     5881 gttatagcgg gggacatcac cgtaggaatt gcagatgtcg agctattaac gataaagatc
     5941 cgcttgattg tggcttcggt tgataaggca aaagaaatcg gcatggactg gtgggaaaat
     6001 gatccgtatc tcagttcaaa aggagccaat aacaaagcgc tcgaagaaga aaataaaatg
     6061 ctgcatgagc ggttaaaaac gcttgaagaa aaaatagaaa cgaaacgtta aggctgtttt
     6121 ggcggatgag agaagattga gatccggctg ctaacaaagc ccgaaaggaa gctgagttgg
     6181 ctgctgccac cgctgagcaa taactagcat aaccccttgg ggcctctaaa cgggtcttga
     6241 ggggtttttt gctgaaagga ggaactatat ccgga
//
