LOCUS       sequence-326582-        5580 bp    DNA     circular SYN 15-APR-2025
DEFINITION  LacI protein regulates the expression of a tetracycline resistance 
            gene and yellow fluorescent protein.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5580)
  AUTHORS   Tack DS, Tonner PD, Pressman A, Olson ND, Levy SF, Romantseva EF, 
            Alperovich N, Vasilyeva O, Ross D
  TITLE     The genotype-phenotype landscape of an allosteric protein.
  JOURNAL   Mol Syst Biol. 2021 Mar;17(3):e10179. doi: 10.15252/msb.202010179.
  PUBMED    33784029
REFERENCE   2  (bases 1 to 5580)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Apr 15, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Mol Syst 
            Biol."; date: "2021-03"; volume: "17(3):e10179. doi"; pages: " 
            10.15252/msb.202010179"
COMMENT     Sequence Label: pTY1-LacI
FEATURES             Location/Qualifiers
     source          1..5580
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     869..888
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        join(1064..1069,1070..1085,1086..1092)
                     /label=tac promoter
                     /note="strong E. coli promoter; hybrid between the trp and 
                     lac UV5 promoters"
     protein_bind    1100..1116
                     /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_RNA        1121..1170
                     /label=sTRSV HHRz
                     /note="hammerhead ribozyme from the tobacco ringspot virus 
                     satellite RNA (Khvorova et al., 2003)"
     RBS             2431..2439
                     /label=Shine-Dalgarno sequence
                     /note="full consensus sequence for ribosome-binding sites 
                     upstream of start codons in E. coli; complementary to a 
                     region in the 3' end of the 16S rRNA (Chen et al., 1994)"
     primer_bind     complement(3312..3331)
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     CDS             3323..4138
                     /codon_start=1
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     CDS             3323..4138
                     /codon_start=1
                     /gene="aph(3')-Ia"
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     primer_bind     complement(3396..3415)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     rep_origin      4227..4772
                     /direction=RIGHT
                     /label=p15A ori
                     /note="Plasmids containing the medium-copy-number p15A 
                     origin of replication can be propagated in E. coli cells 
                     that contain a second plasmid with the ColE1 origin.
                     "
     primer_bind     4673..4692
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     terminator      complement(5126..5173)
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     primer_bind     5185..5204
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     CDS             complement(join(5405..5580,1..907))
                     /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"
ORIGIN
        1 ggtccacgct ggtttgcccc agcaggcgaa aatcctgttt gatggtggtt aacggcggga
       61 tataacatga gctatcttcg gtatcgtcgt atcccactac cgagatatcc gcaccaacgc
      121 gcagcccgga ctcggtaatg gcgcgcattg cgcccagcgc catctgatcg ttggcaacca
      181 gcatcgcagt gggaacgatg ccctcattca gcatttgcat ggtttgttga aaaccggaca
      241 tggcactcca gtcgccttcc cgttccgcta tcggctgaat ttgattgcga gtgagatatt
      301 tatgccagcc agccagacgc agacgcgccg agacagaact taatgggccc gctaacagcg
      361 cgatttgctg gtgacccaat gcgaccagat gctccacgcc cagtcgcgta ccgtcctcat
      421 gggagaaaat aatactgttg atgggtgtct ggtcagagac atcaagaaat aacgccggaa
      481 cattagtgca ggcagcttcc acagcaatgg catcctggtc atccagcgga tagttaatga
      541 tcagcccact gacgcgttgc gcgagaagat tgtgcaccgc cgctttacag gcttcgacgc
      601 cgcttcgttc taccatcgac accaccacgc tggcacccag ttgatcggcg cgagatttaa
      661 tcgccgcgac aatttgcgac ggcgcgtgca gggccagact ggaggtggca acgccaatca
      721 gcaacgactg tttgcccgcc agttgttgtg ccacgcggtt gggaatgtaa ttcagctccg
      781 ccatcgccgc ttccactttt tcccgcgttt tcgcagaaac gtggctggcc tggttcacca
      841 cgcgggaaac ggtctgataa gagacaccgg catactctgc gacatcgtat aacgttactg
      901 gtttcatatt caccaccctg aattgactct cttccgggcg ctatcatgcc ataccgcgaa
      961 aggttttgcg ccattcgatg gcgcgccgcc ataaatctcg gtaccaaatt ccagaaaaga
     1021 ggcctcccga aaggggggcc ttttttcgtt ttggtccgag ctgttgacaa ttaatcatcg
     1081 gctcgtataa tgtgtggaat tgtgagcgga taacaattag ctgtcaccgg atgtgctttc
     1141 cggtctgatg agtccgtgag gacgaaacag cctctacaaa taattttgtt taatactagc
     1201 catcaaggag agctgctaca tgagtagcag tacgaaaatt gcgcttgtca tcaccctcct
     1261 ggatgcgatg gggatcggct tgatcatgcc ggtactgcca acccttctgc gcgagttcat
     1321 tgcaagcgaa gatattgcca accatttcgg ggttctgctc gcactgtacg ccttaatgca
     1381 ggtcatcttt gctccctggt taggcaaaat gtcagacagc tttggacgcc gtcctgtttt
     1441 gctgttaagc cttatcggag cgagcctgga ttacctttta ttggccttct cctcggcact
     1501 gtggatgctt tatttgggtc gtttgctgag tgggattaca ggcgcgacgg gtgccgtggc
     1561 ggcgtcggtg attgctgata cgacgtccgc aagtcaacgt gtgaaattgt tcggctggtt
     1621 aggagcctcc tttggcttgg gcttaatcgc tgggccaatt attggcgggt tcgccggcga
     1681 aatctcacca cattcccctt ttttcatcgc ggcattactc aacattgtca cgttcctggt
     1741 ggtgatgttc tggttccgcg aaacgaaaaa cacccgcgat aacacggata cagaggtggg
     1801 ggttgaaacg caatcgaaca gtgtgtacat cacgctcttc aagaccatgc ccatcctgct
     1861 catcatctac ttctccgcac agttgattgg gcaaatcccg gccacagtgt gggttttgtt
     1921 tacggaaaac cgtttcgggt ggaactccat gatggtgggt ttctctctgg ctggattggg
     1981 acttctgcat agtgttttcc aggctttcgt cgctggccgt attgccacaa agtggggaga
     2041 aaaaaccgct gtattgcttg gttttatcgc agatagctct gcgtttgcct tcttggcatt
     2101 tattagcgaa ggctggctcg tgtttccggt attgattctg ttggctgggg gcggtatcgc
     2161 attacccgcg ctgcagggag ttatgtctat tcaaaccaaa tcacaccaac aaggagcgct
     2221 gcaaggctta cttgtgtccc tgaccaacgc aaccggagtc atcgggccac ttctgttcgc
     2281 tgtaatttat aaccactcac tgccaatttg ggatggatgg atctggatca tcggtcttgc
     2341 cttctactgc atcatcattt tgctgtcaat gacattcatg ctgacgcctc aagcccaagg
     2401 atctaaacaa gaaacgagtg cctaacggcg taaggaggta tttttatggt gtcaaagggt
     2461 gaggaactgt ttacggggat cgtcccgatt cttgttgaac ttgacggcga cgtaaatggt
     2521 cacaagtttt ccgtatcggg cgaaggtgag ggcgatgcga cttatgggaa attaacactg
     2581 aaattcattt gcaccaccgg aaaactgccc gttccttggc ctactctggt aaccacgttc
     2641 ggatatggtt tacagtgttt tgctcgctac ccggaccata tgaaactgca cgatttcttc
     2701 aagtccgcca tgccggaggg ctacgtgcag gaacgtacaa tcttcttcaa agacgatggt
     2761 aattacaaga cccgtgctga agttaaattt gagggggata ctttagtcaa tcgtattgaa
     2821 ttgaagggga ttgactttaa ggaagacggt aatatccttg gccacaagct tgaatacaac
     2881 tacaatagtc acaatgtgta tattatggct gataaacaga agaatggcat taaggttaac
     2941 tttaagatcc gtcacaatat cgaagacgga tctgtccagc ttgctgacca ttatcagcaa
     3001 aatacgccga tcggtgacgg tccggttttg ttgccggaca accattacct gtcctatcag
     3061 tcggcgttaa gtaaagatcc gaatgagaaa cgcgaccata tggttttgtt ggagtttgtg
     3121 acggctgctg gcattacgct tgggatggac gagctgtata aataaaagcg ggagaccaga
     3181 aacaaaaaaa ggccccccgt tagggaggcc ttcaataatt ggttgtgtct caaaatctct
     3241 gatgtacatt gcacaagata aaaatatatc atcatgaaca ataaaactgt ctgcttacat
     3301 aaacagtaat acaaggggtg ttatgagcca tattcaacgg gaaacgtctt gctccaggcc
     3361 gcgattaaat tccaacatgg atgctgattt atatgggtat aaatgggctc gcgataatgt
     3421 cgggcaatca ggtgcgacaa tctatcgatt gtatgggaag cccgatgcgc cagagttgtt
     3481 tctgaaacat ggcaaaggta gcgttgccaa tgatgttaca gatgagatgg tcagactaaa
     3541 ctggctgacg gaatttatgc ctcttccgac catcaagcat tttatccgta ctcctgatga
     3601 tgcatggtta ctcaccactg cgatccccgg gaaaacagca ttccaggtat tagaagaata
     3661 tcctgattca ggtgaaaata ttgttgatgc gctggcagtg ttcctgcgcc ggttgcattc
     3721 gattcctgtt tgtaattgtc cttttaacag cgatcgcgta tttcgtctcg ctcaggcgca
     3781 atcacgaatg aataacggtt tggttgatgc gagtgatttt gatgacgagc gtaatggctg
     3841 gcctgttgaa caagtctgga aagaaatgca taagcttttg ccattctcac cggattcagt
     3901 cgtcactcat ggtgatttct cacttgataa ccttattttt gacgagggga aattaatagg
     3961 ttgtattgat gttggacgag tcggaatcgc agaccgatac caggatcttg ccatcctatg
     4021 gaactgcctc ggtgagtttt ctccttcatt acagaaacgg ctttttcaaa aatatggtat
     4081 tgataatcct gatatgaata aattgcagtt tcatttgatg ctcgatgagt ttttctaagg
     4141 taactgtcag accaagttta ctcatatata ctttagattg atttttcgtt ccactgagcg
     4201 tcagacccct taataagatg atcttcttga gatcgttttg gtctgcgcgt aatctcttgc
     4261 tctgaaaacg aaaaaaccgc cttgcagggc ggtttttcga aggttctctg agctaccaac
     4321 tctttgaacc gaggtaactg gcttggagga gcgcagtcac caaaacttgt cctttcagtt
     4381 tagccttaac cggcgcatga cttcaagact aactcctcta aatcaattac cagtggctgc
     4441 tgccagtggt gcttttgcat gtctttccgg gttggactca agacgatagt taccggataa
     4501 ggcgcagcgg tcggactgaa cggggggttc gtgcatacag tccagcttgg agcgaactgc
     4561 ctacccggaa ctgagtgtca ggcgtggaat gagacaaacg cggccataac agcggaatga
     4621 caccggtaaa ccgaaaggca ggaacaggag agcgcacgag ggagccgcca gggggaaacg
     4681 cctggtatct ttatagtcct gtcgggtttc gccaccactg atttgagcgt cagatttcgt
     4741 gatgcttgtc aggggggcgg agcctatgga aaaacggctt tgccgcggcc ctctcacttc
     4801 cctgttaagt atcttcctgg catcttccag gaaatctccg ccccgttcgt aagccatttc
     4861 cgctcgccgc agtcgaacga ccgagcgtag cgagtcagtg agcgaggaag cggaatatat
     4921 cctgtatcac atattctgct gacgcaccgg tgcagccttt tttctcctgc cacatgaagc
     4981 acttcactga caccctcatc agtgccaaca tagtaagcca gtatacactc cgctagcgct
     5041 gaggtctgcc tcgtgcagcg agtcagtgag cgaggaagca cctcagataa aatatttgct
     5101 catgagcccg aagtggcgag cccgacaaaa aacccctcaa gacccgttta gaggccccaa
     5161 ggggttatgc tagtcttccc catcggtgat gtcggcgata taggcgccag caaccgcacc
     5221 tgtggcgccg gtgatgccgg ccacgatgcg tccggcgtag aggatctgct catgtttgac
     5281 agcttatcat cgatgcataa tgtgcctgtc aaatggacga agcagggatt ctgcaaaccc
     5341 tatgctactc cctcgagccg tcaattgtct gattcgttac caattatttt ttcctcctgg
     5401 attatcactg cccgctttcc agtcgggaaa cctgtcgtgc cagctgcatt aatgaatcgg
     5461 ccaacgcgcg gggagaggcg gtttgcgtat tgggcgccag ggtggttttt cttttcacca
     5521 gtgagactgg caacagctga ttgcccttca ccgcctggcc ctgagagagt tgcagcaagc
//
