LOCUS       Exported                6335 bp ds-DNA     circular SYN 01-JUL-2024
DEFINITION  synthetic circular DNA
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6335)
  AUTHORS   eliasa55
  TITLE     Direct Submission
  JOURNAL   Exported Monday, Jul 8, 2024 from SnapGene 5.0.8
            https://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..6335
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     protein_bind    3..27
                     /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)."
     misc_binding    3..25
                     /locus_tag="LacO"
                     /label=LacO
     RBS             42..64
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     misc_feature    71..73
                     /label=ATG
     CDS             83..100
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             113..133
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQG"
     CDS             134..838
                     /codon_start=1
                     /locus_tag="mCherry"
                     /label=mCherry
                     /translation="VSKGEEDNMAIIKEFMRFKVHMEGSVNGHEFEIEGEGEGRPYEGT
                     QTAKLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNFE
                     DGGVVTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASSERMYPEDGALKG
                     EIKQRLKLKDGGHYDAEVKTTYKAKKPVQLPGAYNVNIKLDITSHNEDYTIVEQYERAE
                     GRHSTGGMDELYK"
     primer_bind     854..897
                     /label=Fragment.FOR
     primer_bind     complement(854..873)
                     /label=IDR_Gibson_vector_Rv
     CDS             875..1060
                     /codon_start=1
                     /label=WIPI2d 364-425aa (IDR)
                     /label=WIPI2d or ßeta
                     /translation="TNEILDSASHDCPLVTQTYGAAAGKAYTDDLGAVGGACLEDEASA
                     LRLDEDSEHPPMILRTD"
     primer_bind     complement(1042..1089)
                     /label=Fragment.REV
     primer_bind     1064..1084
                     /label=IDR_vector_Gibson_Fw
     primer_bind     1129..1148
                     /locus_tag="T7"
                     /label=T7
     promoter        1129..1147
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    1148..1172
                     /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)."
     misc_binding    1148..1170
                     /locus_tag="LacO(1)"
                     /label=LacO
                     /label=LacO(1)
     CDS             1281..1325
                     /codon_start=1
                     /product="affinity and epitope tag derived from pancreatic 
                     ribonuclease A"
                     /label=S-Tag
                     /translation="KETAAAKFERQHMDS"
     terminator      1377..1424
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     rep_origin      1461..1916
                     /locus_tag="M13 origin"
                     /label=M13 origin
     rep_origin      1478..1784
                     /locus_tag="F1 ori"
                     /label=F1 ori
     CDS             complement(2031..2891)
                     /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"
     CDS             complement(2034..2693)
                     /locus_tag="AmpR"
                     /label=AmpR
     promoter        complement(2892..2984)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      2989..3671
                     /locus_tag="ColE1 origin"
                     /label=ColE1 origin
     rep_origin      3065..3653
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     misc_feature    3839..3981
                     /label=bom
                     /note="basis of mobility region from pBR322"
     CDS             complement(4083..4274)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     CDS             complement(4846..5928)
                     /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"
     CDS             complement(4849..5805)
                     /locus_tag="LacI"
                     /label=LacI
     promoter        complement(5929..6006)
                     /gene="lacI"
                     /label=lacI promoter
     promoter        join(6319..6335,1..2)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
ORIGIN
        1 ggggaattgt gagcggataa caattcccct ctagaaataa ttttgtttaa ctttaagaag
       61 gagatatacc atgggcagca gccatcacca tcatcaccac agccaggatc cggaaaacct
      121 gtattttcag ggagtgagca agggcgagga ggataacatg gccatcatca aggagttcat
      181 gcgcttcaag gtgcacatgg agggctccgt gaacggccac gagttcgaga tcgagggcga
      241 gggcgagggc cgcccctacg agggcaccca gaccgccaag ctgaaggtga ccaagggtgg
      301 ccccctgccc ttcgcctggg acatcctgtc ccctcagttc atgtacggct ccaaggccta
      361 cgtgaagcac cccgccgaca tccccgacta cttgaagctg tccttccccg agggcttcaa
      421 gtgggagcgc gtgatgaact tcgaggacgg cggcgtggtg accgtgaccc aggactcctc
      481 cctgcaggac ggcgagttca tctacaaggt gaagctgcgc ggcaccaact tcccctccga
      541 cggccccgta atgcagaaga agaccatggg ctgggaggcc tcctccgagc ggatgtaccc
      601 cgaggacggc gccctgaagg gcgagatcaa gcagaggctg aagctgaagg acggcggcca
      661 ctacgacgct gaggtcaaga ccacctacaa ggccaagaag cccgtgcagc tgcccggcgc
      721 ctacaacgtc aacatcaagt tggacatcac ctcccacaac gaggactaca ccatcgtgga
      781 acagtacgaa cgcgccgagg gccgccactc caccggcggc atggacgagc tgtacaagtc
      841 cggactcaga tctcgagctc aagcttcgaa ttcgaccaat gagatcttgg actctgcctc
      901 tcacgactgc cccttagtca ctcagacata cggcgcagct gcaggaaaag cctacacaga
      961 cgacctgggt gctgtgggtg gcgcctgcct ggaggacgag gccagcgccc tgcgcctgga
     1021 tgaggacagc gagcacccgc ccatgattct tcggactgac tgagcggccg cataatgctt
     1081 aagtcgaaca gaaagtaatc gtattgtaca cggccgcata atcgaaatta atacgactca
     1141 ctatagggga attgtgagcg gataacaatt ccccatctta gtatattagt taagtataag
     1201 aaggagatat acatatggca gatctcaatt ggatatcggc cggccacgcg atcgctgacg
     1261 tcggtaccct cgagtctggt aaagaaaccg ctgctgcgaa atttgaacgc cagcacatgg
     1321 actcgtctac tagcgcagct taattaacct aggctgctgc caccgctgag caataactag
     1381 cataacccct tggggcctct aaacgggtct tgaggggttt tttgctgaaa ggaggaacta
     1441 tatccggatt ggcgaatggg acgcgccctg tagcggcgca ttaagcgcgg cgggtgtggt
     1501 ggttacgcgc agcgtgaccg ctacacttgc cagcgcccta gcgcccgctc ctttcgcttt
     1561 cttcccttcc tttctcgcca cgttcgccgg ctttccccgt caagctctaa atcgggggct
     1621 ccctttaggg ttccgattta gtgctttacg gcacctcgac cccaaaaaac ttgattaggg
     1681 tgatggttca cgtagtgggc catcgccctg atagacggtt tttcgccctt tgacgttgga
     1741 gtccacgttc tttaatagtg gactcttgtt ccaaactgga acaacactca accctatctc
     1801 ggtctattct tttgatttat aagggatttt gccgatttcg gcctattggt taaaaaatga
     1861 gctgatttaa caaaaattta acgcgaattt taacaaaata ttaacgttta caatttctgg
     1921 cggcacgatg gcatgagatt atcaaaaagg atcttcacct agatcctttt aaattaaaaa
     1981 tgaagtttta aatcaatcta aagtatatat gagtaaactt ggtctgacag ttaccaatgc
     2041 ttaatcagtg aggcacctat ctcagcgatc tgtctatttc gttcatccat agttgcctga
     2101 ctccccgtcg tgtagataac tacgatacgg gagggcttac catctggccc cagtgctgca
     2161 atgataccgc gagacccacg ctcaccggct ccagatttat cagcaataaa ccagccagcc
     2221 ggaagggccg agcgcagaag tggtcctgca actttatccg cctccatcca gtctattaat
     2281 tgttgccggg aagctagagt aagtagttcg ccagttaata gtttgcgcaa cgttgttgcc
     2341 attgctacag gcatcgtggt gtcacgctcg tcgtttggta tggcttcatt cagctccggt
     2401 tcccaacgat caaggcgagt tacatgatcc cccatgttgt gcaaaaaagc ggttagctcc
     2461 ttcggtcctc cgatcgttgt cagaagtaag ttggccgcag tgttatcact catggttatg
     2521 gcagcactgc ataattctct tactgtcatg ccatccgtaa gatgcttttc tgtgactggt
     2581 gagtactcaa ccaagtcatt ctgagaatag tgtatgcggc gaccgagttg ctcttgcccg
     2641 gcgtcaatac gggataatac cgcgccacat agcagaactt taaaagtgct catcattgga
     2701 aaacgttctt cggggcgaaa actctcaagg atcttaccgc tgttgagatc cagttcgatg
     2761 taacccactc gtgcacccaa ctgatcttca gcatctttta ctttcaccag cgtttctggg
     2821 tgagcaaaaa caggaaggca aaatgccgca aaaaagggaa taagggcgac acggaaatgt
     2881 tgaatactca tactcttcct ttttcaatca tgattgaagc atttatcagg gttattgtct
     2941 catgagcgga tacatatttg aatgtattta gaaaaataaa caaataggtc atgaccaaaa
     3001 tcccttaacg tgagttttcg ttccactgag cgtcagaccc cgtagaaaag atcaaaggat
     3061 cttcttgaga tccttttttt ctgcgcgtaa tctgctgctt gcaaacaaaa aaaccaccgc
     3121 taccagcggt ggtttgtttg ccggatcaag agctaccaac tctttttccg aaggtaactg
     3181 gcttcagcag agcgcagata ccaaatactg tccttctagt gtagccgtag ttaggccacc
     3241 acttcaagaa ctctgtagca ccgcctacat acctcgctct gctaatcctg ttaccagtgg
     3301 ctgctgccag tggcgataag tcgtgtctta ccgggttgga ctcaagacga tagttaccgg
     3361 ataaggcgca gcggtcgggc tgaacggggg gttcgtgcac acagcccagc ttggagcgaa
     3421 cgacctacac cgaactgaga tacctacagc gtgagctatg agaaagcgcc acgcttcccg
     3481 aagggagaaa ggcggacagg tatccggtaa gcggcagggt cggaacagga gagcgcacga
     3541 gggagcttcc agggggaaac gcctggtatc tttatagtcc tgtcgggttt cgccacctct
     3601 gacttgagcg tcgatttttg tgatgctcgt caggggggcg gagcctatgg aaaaacgcca
     3661 gcaacgcggc ctttttacgg ttcctggcct tttgctggcc ttttgctcac atgttctttc
     3721 ctgcgttatc ccctgattct gtggataacc gtattaccgc ctttgagtga gctgataccg
     3781 ctcgccgcag ccgaacgacc gagcgcagcg agtcagtgag cgaggaagcg gaagagcgcc
     3841 tgatgcggta ttttctcctt acgcatctgt gcggtatttc acaccgcata tatggtgcac
     3901 tctcagtaca atctgctctg atgccgcata gttaagccag tatacactcc gctatcgcta
     3961 cgtgactggg tcatggctgc gccccgacac ccgccaacac ccgctgacgc gccctgacgg
     4021 gcttgtctgc tcccggcatc cgcttacaga caagctgtga ccgtctccgg gagctgcatg
     4081 tgtcagaggt tttcaccgtc atcaccgaaa cgcgcgaggc agctgcggta aagctcatca
     4141 gcgtggtcgt gaagcgattc acagatgtct gcctgttcat ccgcgtccag ctcgttgagt
     4201 ttctccagaa gcgttaatgt ctggcttctg ataaagcggg ccatgttaag ggcggttttt
     4261 tcctgtttgg tcactgatgc ctccgtgtaa gggggatttc tgttcatggg ggtaatgata
     4321 ccgatgaaac gagagaggat gctcacgata cgggttactg atgatgaaca tgcccggtta
     4381 ctggaacgtt gtgagggtaa acaactggcg gtatggatgc ggcgggacca gagaaaaatc
     4441 actcagggtc aatgccagcg cttcgttaat acagatgtag gtgttccaca gggtagccag
     4501 cagcatcctg cgatgcagat ccggaacata atggtgcagg gcgctgactt ccgcgtttcc
     4561 agactttacg aaacacggaa accgaagacc attcatgttg ttgctcaggt cgcagacgtt
     4621 ttgcagcagc agtcgcttca cgttcgctcg cgtatcggtg attcattctg ctaaccagta
     4681 aggcaacccc gccagcctag ccgggtcctc aacgacagga gcacgatcat gctagtcatg
     4741 ccccgcgccc accggaagga gctgactggg ttgaaggctc tcaagggcat cggtcgagat
     4801 cccggtgcct aatgagtgag ctaacttaca ttaattgcgt tgcgctcact gcccgctttc
     4861 cagtcgggaa acctgtcgtg ccagctgcat taatgaatcg gccaacgcgc ggggagaggc
     4921 ggtttgcgta ttgggcgcca gggtggtttt tcttttcacc agtgagacgg gcaacagctg
     4981 attgcccttc accgcctggc cctgagagag ttgcagcaag cggtccacgc tggtttgccc
     5041 cagcaggcga aaatcctgtt tgatggtggt taacggcggg atataacatg agctgtcttc
     5101 ggtatcgtcg tatcccacta ccgagatgtc cgcaccaacg cgcagcccgg actcggtaat
     5161 ggcgcgcatt gcgcccagcg ccatctgatc gttggcaacc agcatcgcag tgggaacgat
     5221 gccctcattc agcatttgca tggtttgttg aaaaccggac atggcactcc agtcgccttc
     5281 ccgttccgct atcggctgaa tttgattgcg agtgagatat ttatgccagc cagccagacg
     5341 cagacgcgcc gagacagaac ttaatgggcc cgctaacagc gcgatttgct ggtgacccaa
     5401 tgcgaccaga tgctccacgc ccagtcgcgt accgtcttca tgggagaaaa taatactgtt
     5461 gatgggtgtc tggtcagaga catcaagaaa taacgccgga acattagtgc aggcagcttc
     5521 cacagcaatg gcatcctggt catccagcgg atagttaatg atcagcccac tgacgcgttg
     5581 cgcgagaaga ttgtgcaccg ccgctttaca ggcttcgacg ccgcttcgtt ctaccatcga
     5641 caccaccacg ctggcaccca gttgatcggc gcgagattta atcgccgcga caatttgcga
     5701 cggcgcgtgc agggccagac tggaggtggc aacgccaatc agcaacgact gtttgcccgc
     5761 cagttgttgt gccacgcggt tgggaatgta attcagctcc gccatcgccg cttccacttt
     5821 ttcccgcgtt ttcgcagaaa cgtggctggc ctggttcacc acgcgggaaa cggtctgata
     5881 agagacaccg gcatactctg cgacatcgta taacgttact ggtttcacat tcaccaccct
     5941 gaattgactc tcttccgggc gctatcatgc cataccgcga aaggttttgc gccattcgat
     6001 ggtgtccggg atctcgacgc tctcccttat gcgactcctg cattaggaag cagcccagta
     6061 gtaggttgag gccgttgagc accgccgccg caaggaatgg tgcatgcaag gagatggcgc
     6121 ccaacagtcc cccggccacg gggcctgcca ccatacccac gccgaaacaa gcgctcatga
     6181 gcccgaagtg gcgagcccga tcttccccat cggtgatgtc ggcgatatag gcgccagcaa
     6241 ccgcacctgt ggcgccggtg atgccggcca cgatgcgtcc ggcgtagagg atcgagatcg
     6301 atctcgatcc cgcgaaatta atacgactca ctata
//
