LOCUS       sequence-428954-        5294 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses HaloTag-PILATeS-RGD with 700 pM BiT tag in bacteria.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5294)
  AUTHORS   Zhong BL, Elliot JM, Wang P, Li H, Hall RN, Wang B, Prakash M, Dunn 
            AR
  TITLE     Split Luciferase Molecular Tension Sensors for Bioluminescent 
            Readout of Mechanical Forces in Biological Systems.
  JOURNAL   ACS Sens. 2024 Jul 26;9(7):3489-3495. doi: 
            10.1021/acssensors.3c02664. Epub 2024 Jul 7.
  PUBMED    38973210
REFERENCE   2  (bases 1 to 5294)
  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"; doi: 
            "10.1021/acssensors.3c02664"; journalName: "ACS Sens"; date: 
            "2024-07-26- 26"; volume: "9"; issue: "7"; pages: "3489-3495"
COMMENT     Sequence Label: T7-LacO-Halo-PILATeS-RGD (700pM)
FEATURES             Location/Qualifiers
     source          1..5294
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      578..1166
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     1067..1086
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     1320..1337
                     /label=L4440
                     /note="L4440 vector, forward primer"
     primer_bind     1435..1452
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     1436..1452
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     1915..1934
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        1915..1933
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    1934..1958
                     /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)."
     CDS             3288..3305
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     terminator      3313..3360
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     primer_bind     complement(3583..3602)
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     primer_bind     complement(3608..3627)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     promoter        3669..3760
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(3761..3829,3830..4621)
                     /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(3979..3998)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     CDS             join(4704..5294,1..492)
                     /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     complement(4723..4742)
                     /label=LacI-R
                     /note="LacI, reverse primer"
ORIGIN
        1 ctggctggct ggcataaata tctcactcgc aatcaaattc agccgatagc ggaacgggaa
       61 ggcgactgga gtgccatgtc cggttttcaa caaaccatgc aaatgctgaa tgagggcatc
      121 gttccaactg cgatgctggt tgccaacgat cagatggcgc tgggcgcaat gcgcgccatt
      181 accgagtccg ggctgcgcgt tggtgcggat atctcggtag tgggatacga cgataccgaa
      241 gatagctcat gttatatccc gccgttaacc accatcaaac aggattttcg cctgctgggg
      301 caaaccagcg tggaccgctt gctgcaactc tctcagggcc aggcggtgaa gggcaatcag
      361 ctgttgccag tctcactggt gaaaagaaaa accaccctgg cgcccaatac gcaaaccgcc
      421 tctccccgcg cgttggccga ttcattaatg cagctggcac gacaggtttc ccgactggaa
      481 agcgggcagt gactcatgac caaaatccct taacgtgagt tacgcgcgcg tcgttccact
      541 gagcgtcaga ccccgtagaa aagatcaaag gatcttcttg agatcctttt tttctgcgcg
      601 taatctgctg cttgcaaaca aaaaaaccac cgctaccagc ggtggtttgt ttgccggatc
      661 aagagctacc aactcttttt ccgaaggtaa ctggcttcag cagagcgcag ataccaaata
      721 ctgttcttct agtgtagccg tagttagccc accacttcaa gaactctgta gcaccgccta
      781 catacctcgc tctgctaatc ctgttaccag tggctgctgc cagtggcgat aagtcgtgtc
      841 ttaccgggtt ggactcaaga cgatagttac cggataaggc gcagcggtcg ggctgaacgg
      901 ggggttcgtg cacacagccc agcttggagc gaacgaccta caccgaactg agatacctac
      961 agcgtgagct atgagaaagc gccacgcttc ccgaagggag aaaggcggac aggtatccgg
     1021 taagcggcag ggtcggaaca ggagagcgca cgagggagct tccaggggga aacgcctggt
     1081 atctttatag tcctgtcggg tttcgccacc tctgacttga gcgtcgattt ttgtgatgct
     1141 cgtcaggggg gcggagccta tggaaaaacg ccagcaacgc ggccttttta cggttcctgg
     1201 ccttttgctg gccttttgct cacatgttct ttcctgcgtt atcccctgat tctgtggata
     1261 accgtattac cgcctttgag tgagctgata ccgctcgccg cagccgaacg accgagcgca
     1321 gcgagtcagt gagcgaggaa gcggaaggcg agagtaggga actgccaggc atcaaactaa
     1381 gcagaaggcc cctgacggat ggcctttttg cgtttctaca aactctttct gtgttgtaaa
     1441 acgacggcca gtcttaagct cgggccccct gggcggttct gataacgagt aatcgttaat
     1501 ccgcaaataa cgtaaaaacc cgcttcggcg ggttttttta tggggggagt ttagggaaag
     1561 agcatttgtc agaatattta agggcgcctg tcactttgct tgatatatga gaattattta
     1621 accttataaa tgagaaaaaa gcaacgcact ttaaataaga tacgttgctt tttcgattga
     1681 tgaacaccta taattaaact attcatctat tatttatgat tttttgtata tacaatattt
     1741 ctagtttgtt aaagagaatt aagaaaataa atctcgaaaa taataaaggg aaaatcagtt
     1801 tttgatatca aaattataca tgtcaacgat aatacaaaat ataatacaaa ctataagatg
     1861 ttatcagtat ttattatcat ttagaataaa ttttgtgtcg cccttccgcg aaattaatac
     1921 gactcactat aggggaattg tgagcggata acaattcccc tctagaaata attttgttta
     1981 acttttagga ggtaaaacat atgggcagcg aaattggcac tggttttccg tttgatccgc
     2041 attacgttga agttttaggc gaacgcatgc actacgtaga cgtgggtccg cgtgacggca
     2101 ccccggtcct gttcctgcac ggcaatccaa cctctagcta tgtctggcgc aacattatcc
     2161 cgcacgtcgc gccgacgcac cgtagtattg caccggatct gatcggtatg ggtaaaagcg
     2221 ataaaccgga cctgggctat ttctttgacg accacgttcg ttttatggat gcgttcattg
     2281 aggccttggg tctggaagag gtggtgttgg ttatccatga ctggggtagc gctctgggct
     2341 tccactgggc gaagcgcaac ccggaacgtg tgaagggcat cgcatttatg gagttcattc
     2401 gcccgattcc gacctgggac gagtggccgg agttcgcgcg tgaaacgttc caggcctttc
     2461 gcaccaccga tgtcggtcgt aagctgatta ttgaccaaaa tgttttcatc gagggcacgt
     2521 tgccgatggg cgttgttcgt ccgctgaccg aagtcgagat ggaccattac cgtgagccgt
     2581 ttttgaatcc ggtggatcgt gagccgctgt ggcgtttccc gaacgagctg ccgatcgccg
     2641 gtgaaccggc gaacatcgtt gcgctggttg aagagtacat ggactggctg caccagagcc
     2701 cggtgccgaa actgctgttt tggggtacgc cgggtgtgct gatcccgcct gcggaagcag
     2761 cccgcctggc gaaatccctg ccgaacgcga aggcagtgga tattggtcca ggcctgaatc
     2821 tgctgcagga agataacccg gacctgattg gtagcgaaat cgcgcgctgg ctgagcacgt
     2881 tggagatcag cggcggtgct ggcgagttca tggaaactct gcatattacc aaaaccatga
     2941 aaaacattga agtgccggaa accaaaaccg cgagctttga atgcgaagtg agccatttta
     3001 acgtgccgag catgtggctg aaaaacggcg tggaaattga aatgagcgaa aaatttaaaa
     3061 ttgtggtgca gggcaaactg catcagctga ttattatgaa caccagcacc gaagatagcg
     3121 cggaatatac ctttgtgtgc ggtgttagtg gctggcgtct gtttaagaaa attagcaacg
     3181 atcaggtgag cgcgaccctg accgtgactt ccgagaatct gtacttcgag caaggtaccg
     3241 tctatgccgt caccggtcgt ggcgattccc cggcaagctc ggcggcacac caccatcatc
     3301 accattaacc ccctagcata accccttggg gcctctaaac gggtcttgag gggttttttg
     3361 cccctgagac gcgtcaatcg agttcgtacc taagggcgac accccataat tagcccgggc
     3421 gaaaggccca gtctttcgac tgagcctttc gttttatttg atgcctggca gttccctact
     3481 ctcgcatggg gagtccccac actaccatcg gcgctacggc gtttcacttc tgagttcggc
     3541 atggggtcag gtgggaccac cgcgctactg ccgccaggca aacaaggggt gttatgagcc
     3601 atattcaggt ataaatgggc tcgcgataat gttcagaatt ggttaattgg ttgtaacact
     3661 gacccctatt tgtttatttt tctaaataca ttcaaatatg tatccgctca tgagacaata
     3721 accctgataa atgcttcaat aatattgaaa aaggaagaat atgagtattc aacatttccg
     3781 tgtcgccctt attccctttt ttgcggcatt ttgccttcct gtttttgctc acccagaaac
     3841 gctggtgaaa gtaaaagatg ctgaagatca gttgggtgca cgagtgggtt acatcgaact
     3901 ggatctcaac agcggtaaga tccttgagag ttttcgcccc gaagaacgtt ttccaatgat
     3961 gagcactttt aaagttctgc tatgtggcgc ggtattatcc cgtattgacg ccgggcaaga
     4021 gcaactcggt cgccgcatac actattctca gaatgacttg gttgagtact caccagtcac
     4081 agaaaagcat cttacggatg gcatgacagt aagagaatta tgcagtgctg ccataaccat
     4141 gagtgataac actgcggcca acttacttct gacaacgatc ggaggaccga aggagctaac
     4201 cgcttttttg cacaacatgg gggatcatgt aactcgcctt gatcgttggg aaccggagct
     4261 gaatgaagcc ataccaaacg acgagcgtga caccacgatg cctgtagcga tggcaacaac
     4321 gttgcgcaaa ctattaactg gcgaactact tactctagct tcccggcaac aattaataga
     4381 ctggatggag gcggataaag ttgcaggacc acttctgcgc tcggcccttc cggctggctg
     4441 gtttattgct gataaatccg gagccggtga gcgtggttct cgcggtatca tcgcagcgct
     4501 ggggccagat ggtaagccct cccgtatcgt agttatctac acgacgggga gtcaggcaac
     4561 tatggatgaa cgaaatagac agatcgctga gataggtgcc tcactgatta agcattggta
     4621 agcggcgcgc catcgaatgg cgcaaaacct ttcgcggtat ggcatgatag cgcccggaag
     4681 agagtcaatt cagggtggtg aatatgaaac cagtaacgtt atacgatgtc gcagagtatg
     4741 ccggtgtctc ttatcagacc gtttcccgcg tggtgaacca ggccagccac gtttctgcga
     4801 aaacgcggga aaaagtggaa gcggcgatgg cggagctgaa ttacattccc aaccgcgtgg
     4861 cacaacaact ggcgggcaaa cagtcgttgc tgattggcgt tgccacctcc agtctggccc
     4921 tgcacgcgcc gtcgcaaatt gtcgcggcga ttaaatctcg cgccgatcaa ctgggtgcca
     4981 gcgtggtggt gtcgatggta gaacgaagcg gcgtcgaagc ctgtaaagcg gcggtgcaca
     5041 atcttctcgc gcaacgcgtc agtgggctga tcattaacta tccgctggat gaccaggatg
     5101 ccattgctgt ggaagctgcc tgcactaatg ttccggcgtt atttcttgat gtctctgacc
     5161 agacacccat caacagtatt attttctccc atgaggacgg tacgcgactg ggcgtggagc
     5221 atctggtcgc attgggtcac cagcaaatcg cgctgttagc gggcccatta agttctgtct
     5281 cggcgcgtct gcgt
//
