LOCUS       sequence-410974-        7197 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  synthetic circular DNA
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7197)
  AUTHORS   Liu J, Huang C, Shin DH, Yokota H, Jancarik J, Kim JS, Adams PD, Kim
            R, Kim SH
  TITLE     Crystal structure of a heat-inducible transcriptional repressor HrcA
            from Thermotoga maritima: structural insight into DNA binding and 
            dimerization.
  JOURNAL   J Mol Biol. 2005 Jul 29. 350(5):987-96.
  PUBMED    15979091
REFERENCE   2  (bases 1 to 7197)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Dec 15, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "J Mol Biol.
            2005 Jul 29. 350(5):987-96."
COMMENT     Sequence Label: pLIC.B3.TM0851
FEATURES             Location/Qualifiers
     source          1..7197
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     1..20
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        1..19
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    20..44
                     /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)."
     RBS             59..81
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             101..118
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             140..160
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQG"
     CDS             1199..1909
                     /codon_start=1
                     /product="GFP variant optimized for excitation by UV light"
                     /label=GFPuv
                     /translation="KGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLKFI
                     CTTGKLPVPWPTLVTTFSYGVQCFSRYPDHMKRHDFFKSAMPEGYVQERTISFKDDGNY
                     KTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYITADKQKNGIKANF
                     KIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLLEFV
                     TAAGITHGMDELYK"
     primer_bind     complement(1223..1251)
                     /label=GFP-R
                     /note="GFP, reverse primer. Does NOT anneal to EGFP"
     primer_bind     1846..1866
                     /label=GFP-F
                     /note="GFP, forward primer. Does NOT anneal to EGFP"
     CDS             1929..1946
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(1999..2017)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      2013..2060
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     rep_origin      2097..2552
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2184..2203)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2394..2415
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        2578..2682
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(2683..2751,2752..3543)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPAAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     complement(2901..2920)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      3714..4302
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4203..4222
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     4456..4473
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    4488..4627
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(4553..4572)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     4713..4735
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(4729..4920)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     CDS             complement(5729..6811)
                     /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"
     primer_bind     6773..6792
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(6812..6889)
                     /gene="lacI"
                     /label=lacI promoter
     primer_bind     7095..7114
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
ORIGIN
        1 taatacgact cactataggg gaattgtgag cggataacaa ttcccctcta gaaataattt
       61 tgtttaactt taagaaggag atatacatat gggcagcaga catcatcatc atcatcacga
      121 ttacgatatc ccaacgaccg aaaaccttta cttccagggc ggcggtggtg gcggcatgag
      181 aaggctcaac aggaagaaca acgaagcctt gaagaaactg aacgatcgtc agagaaaagt
      241 gctttactgc atcgtgagag agtacataga aaataaaaaa cctgtgagct cacagagagt
      301 tctggaagtg agcaatatag agttcagcag tgccactata agaaatgata tgaaaaaatt
      361 ggagtacctt ggatacatat accagcccca cacatcggct gggaggattc caactgacaa
      421 ggggttgaga ttttactacg aagaaatgct gaaaatctca aaagaaacat cagaagcgga
      481 tctcgcagtg gagactttca aatcgatgcc tctcgctgat ccggaaaagg ttctcttcct
      541 tgcgggaaat cttctagctc gattgacgga aggttacgtc ctcatagaaa gaccaaacac
      601 aagagatttg aaaatactca gagtgatgct cattcctgtt tcggaagact acctcatttt
      661 ttccatattg acggaattcg gtgtttccaa agttactcct atcaaaactc aggaacgact
      721 gaactgggaa gaaatcgaaa gacagctcaa cttcctcctc agaggcagaa cagtaggaga
      781 agtgttgatg ggtaaaatag aaagcctgaa aggaagtgga ttcctcaggt tgattgaatc
      841 tctgataggt gagactgtcg agagatatct ggatgcaggt cttgagaacc ttctaaagga
      901 cgaaacgctc actctggagg acatcagaaa ccttctcgaa gaagtgaagg atcagaaatt
      961 tctcgagagc ttagttggag aaggcataac cgtcaggata ggaagggaaa tcggcagaaa
     1021 aaagctggaa aaattcgccg ttttttcagg gaaatacttt aaaggagaat ctcctatcgg
     1081 gagtgtgtac ctcttcacgt cgaaggttac gaagtacgat agaaatcaca gggtgtttga
     1141 atacattctg aaccgtcttt cagagtattt cacttcaact tccaggaggt aggggcgcaa
     1201 aggagaagaa ctgttcactg gtgttgtccc aattcttgtt gaattagatg gtgatgttaa
     1261 tgggcacaaa ttttctgtca gtggagaggg tgaaggtgat gcaacatacg gaaaacttac
     1321 ccttaaattt atttgcacta ctggaaaact acctgttcca tggccaacac ttgtcactac
     1381 tttctcttat ggtgttcaat gcttttcccg ttatccggat cacatgaaac ggcatgactt
     1441 tttcaagagt gccatgcccg aaggttatgt acaggaacgc actatatctt tcaaagatga
     1501 cgggaactac aagacgcgtg ctgaagtcaa gtttgaaggt gatacccttg ttaatcgtat
     1561 cgagttaaaa ggtattgatt ttaaagaaga tggaaacatt ctcggacaca aactcgagta
     1621 caactataac tcacacaatg tatacatcac ggcagacaaa caaaagaatg gaatcaaagc
     1681 taacttcaaa attcgccaca acattgaaga tggctccgtt caactagcag accattatca
     1741 acaaaatact ccaattggcg atggccctgt ccttttacca gacaaccatt acctgtcgac
     1801 acaatctgcc ctttcgaaag atcccaacga aaagcgtgac cacatggtcc ttcttgagtt
     1861 tgtaactgct gctgggatta cacatggcat ggatgagctc tacaaataag cttgcggccg
     1921 cactcgagca ccaccaccac caccactgag atccggctgc taacaaagcc cgaaaggaag
     1981 ctgagttggc tgctgccacc gctgagcaat aactagcata accccttggg gcctctaaac
     2041 gggtcttgag gggttttttg ctgaaaggag gaactatatc cggattggcg aatgggacgc
     2101 gccctgtagc ggcgcattaa gcgcggcggg tgtggtggtt acgcgcagcg tgaccgctac
     2161 acttgccagc gccctagcgc ccgctccttt cgctttcttc ccttcctttc tcgccacgtt
     2221 cgccggcttt ccccgtcaag ctctaaatcg ggggctccct ttagggttcc gatttagtgc
     2281 tttacggcac ctcgacccca aaaaacttga ttagggtgat ggttcacgta gtgggccatc
     2341 gccctgatag acggtttttc gccctttgac gttggagtcc acgttcttta atagtggact
     2401 cttgttccaa actggaacaa cactcaaccc tatctcggtc tattcttttg atttataagg
     2461 gattttgccg atttcggcct attggttaaa aaatgagctg atttaacaaa aatttaacgc
     2521 gaattttaac aaaatattaa cgcttacaat ttaggtggca cttttcgggg aaatgtgcgc
     2581 ggaaccccta tttgtttatt tttctaaata cattcaaata tgtatccgct catgagacaa
     2641 taaccctgat aaatgcttca ataatattga aaaaggaaga gtatgagtat tcaacatttc
     2701 cgtgtcgccc ttattccctt ttttgcggca ttttgccttc ctgtttttgc tcacccagaa
     2761 acgctggtga aagtaaaaga tgctgaagat cagttgggtg cacgagtggg ttacatcgaa
     2821 ctggatctca acagcggtaa gatccttgag agttttcgcc ccgaagaacg ttttccaatg
     2881 atgagcactt ttaaagttct gctatgtggc gcggtattat cccgtattga cgccgggcaa
     2941 gagcaactcg gtcgccgcat acactattct cagaatgact tggttgagta ctcaccagtc
     3001 acagaaaagc atcttacgga tggcatgaca gtaagagaat tatgcagtgc tgccataacc
     3061 atgagtgata acactgcggc caacttactt ctgacaacga tcggaggacc gaaggagcta
     3121 accgcttttt tgcacaacat gggggatcat gtaactcgcc ttgatcgttg ggaaccggag
     3181 ctgaatgaag ccataccaaa cgacgagcgt gacaccacga tgcctgcagc aatggcaaca
     3241 acgttgcgca aactattaac tggcgaacta cttactctag cttcccggca acaattaata
     3301 gactggatgg aggcggataa agttgcagga ccacttctgc gctcggccct tccggctggc
     3361 tggtttattg ctgataaatc tggagccggt gagcgtgggt ctcgcggtat cattgcagca
     3421 ctggggccag atggtaagcc ctcccgtatc gtagttatct acacgacggg gagtcaggca
     3481 actatggatg aacgaaatag acagatcgct gagataggtg cctcactgat taagcattgg
     3541 taactgtcag accaagttta ctcatatata ctttagattg atttaaaact tcatttttaa
     3601 tttaaaagga tctaggtgaa gatccttttt gataatctca tgaccaaaat cccttaacgt
     3661 gagttttcgt tccactgagc gtcagacccc gtagaaaaga tcaaaggatc ttcttgagat
     3721 cctttttttc tgcgcgtaat ctgctgcttg caaacaaaaa aaccaccgct accagcggtg
     3781 gtttgtttgc cggatcaaga gctaccaact ctttttccga aggtaactgg cttcagcaga
     3841 gcgcagatac caaatactgt ccttctagtg tagccgtagt taggccacca cttcaagaac
     3901 tctgtagcac cgcctacata cctcgctctg ctaatcctgt taccagtggc tgctgccagt
     3961 ggcgataagt cgtgtcttac cgggttggac tcaagacgat agttaccgga taaggcgcag
     4021 cggtcgggct gaacgggggg ttcgtgcaca cagcccagct tggagcgaac gacctacacc
     4081 gaactgagat acctacagcg tgagctatga gaaagcgcca cgcttcccga agggagaaag
     4141 gcggacaggt atccggtaag cggcagggtc ggaacaggag agcgcacgag ggagcttcca
     4201 gggggaaacg cctggtatct ttatagtcct gtcgggtttc gccacctctg acttgagcgt
     4261 cgatttttgt gatgctcgtc aggggggcgg agcctatgga aaaacgccag caacgcggcc
     4321 tttttacggt tcctggcctt ttgctggcct tttgctcaca tgttctttcc tgcgttatcc
     4381 cctgattctg tggataaccg tattaccgcc tttgagtgag ctgataccgc tcgccgcagc
     4441 cgaacgaccg agcgcagcga gtcagtgagc gaggaagcgg aagagcgcct gatgcggtat
     4501 tttctcctta cgcatctgtg cggtatttca caccgcaatg gtgcactctc agtacaatct
     4561 gctctgatgc cgcatagtta agccagtata cactccgcta tcgctacgtg actgggtcat
     4621 ggctgcgccc cgacacccgc caacacccgc tgacgcgccc tgacgggctt gtctgctccc
     4681 ggcatccgct tacagacaag ctgtgaccgt ctccgggagc tgcatgtgtc agaggttttc
     4741 accgtcatca ccgaaacgcg cgaggcagct gcggtaaagc tcatcagcgt ggtcgtgaag
     4801 cgattcacag atgtctgcct gttcatccgc gtccagctcg ttgagtttct ccagaagcgt
     4861 taatgtctgg cttctgataa agcgggccat gttaagggcg gttttttcct gtttggtcac
     4921 tgatgcctcc gtgtaagggg gatttctgtt catgggggta atgataccga tgaaacgaga
     4981 gaggatgctc acgatacggg ttactgatga tgaacatgcc cggttactgg aacgttgtga
     5041 gggtaaacaa ctggcggtat ggatgcggcg ggaccagaga aaaatcactc agggtcaatg
     5101 ccagcgcttc gttaatacag atgtaggtgt tccacagggt agccagcagc atcctgcgat
     5161 gcagatccgg aacataatgg tgcagggcgc tgacttccgc gtttccagac tttacgaaac
     5221 acggaaaccg aagaccattc atgttgttgc tcaggtcgca gacgttttgc agcagcagtc
     5281 gcttcacgtt cgctcgcgta tcggtgattc attctgctaa ccagtaaggc aaccccgcca
     5341 gcctagccgg gtcctcaacg acaggagcac gatcatgcgc acccgtgggg ccgccatgcc
     5401 ggcgataatg gcctgcttct cgccgaaacg tttggtggcg ggaccagtga cgaaggcttg
     5461 agcgagggcg tgcaagattc cgaataccgc aagcgacagg ccgatcatcg tcgcgctcca
     5521 gcgaaagcgg tcctcgccga aaatgaccca gagcgctgcc ggcacctgtc ctacgagttg
     5581 catgataaag aagacagtca taagtgcggc gacgatagtc atgccccgcg cccaccggaa
     5641 ggagctgact gggttgaagg ctctcaaggg catcggtcga gatcccggtg cctaatgagt
     5701 gagctaactt acattaattg cgttgcgctc actgcccgct ttccagtcgg gaaacctgtc
     5761 gtgccagctg cattaatgaa tcggccaacg cgcggggaga ggcggtttgc gtattgggcg
     5821 ccagggtggt ttttcttttc accagtgaga cgggcaacag ctgattgccc ttcaccgcct
     5881 ggccctgaga gagttgcagc aagcggtcca cgctggtttg ccccagcagg cgaaaatcct
     5941 gtttgatggt ggttaacggc gggatataac atgagctgtc ttcggtatcg tcgtatccca
     6001 ctaccgagat atccgcacca acgcgcagcc cggactcggt aatggcgcgc attgcgccca
     6061 gcgccatctg atcgttggca accagcatcg cagtgggaac gatgccctca ttcagcattt
     6121 gcatggtttg ttgaaaaccg gacatggcac tccagtcgcc ttcccgttcc gctatcggct
     6181 gaatttgatt gcgagtgaga tatttatgcc agccagccag acgcagacgc gccgagacag
     6241 aacttaatgg gcccgctaac agcgcgattt gctggtgacc caatgcgacc agatgctcca
     6301 cgcccagtcg cgtaccgtct tcatgggaga aaataatact gttgatgggt gtctggtcag
     6361 agacatcaag aaataacgcc ggaacattag tgcaggcagc ttccacagca atggcatcct
     6421 ggtcatccag cggatagtta atgatcagcc cactgacgcg ttgcgcgaga agattgtgca
     6481 ccgccgcttt acaggcttcg acgccgcttc gttctaccat cgacaccacc acgctggcac
     6541 ccagttgatc ggcgcgagat ttaatcgccg cgacaatttg cgacggcgcg tgcagggcca
     6601 gactggaggt ggcaacgcca atcagcaacg actgtttgcc cgccagttgt tgtgccacgc
     6661 ggttgggaat gtaattcagc tccgccatcg ccgcttccac tttttcccgc gttttcgcag
     6721 aaacgtggct ggcctggttc accacgcggg aaacggtctg ataagagaca ccggcatact
     6781 ctgcgacatc gtataacgtt actggtttca cattcaccac cctgaattga ctctcttccg
     6841 ggcgctatca tgccataccg cgaaaggttt tgcgccattc gatggtgtcc gggatctcga
     6901 cgctctccct tatgcgactc ctgcattagg aagcagccca gtagtaggtt gaggccgttg
     6961 agcaccgccg ccgcaaggaa tggtgcatgc aaggagatgg cgcccaacag tcccccggcc
     7021 acggggcctg ccaccatacc cacgccgaaa caagcgctca tgagcccgaa gtggcgagcc
     7081 cgatcttccc catcggtgat gtcggcgata taggcgccag caaccgcacc tgtggcgccg
     7141 gtgatgccgg ccacgatgcg tccggcgtag aggatcgaga tctcgatccc gcgaaat
//
