LOCUS       sequence-428948-        5309 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses HaloTag-RGD-PILATeS Force Inert Control with 200 nM BiT 
            tag in bacteria.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5309)
  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 5309)
  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-RGD-PILATeS (200nM, Force Inert)
FEATURES             Location/Qualifiers
     source          1..5309
                     /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             3297..3314
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     terminator      3328..3375
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     primer_bind     complement(3598..3617)
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     primer_bind     complement(3623..3642)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     promoter        3684..3775
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(3776..3844,3845..4636)
                     /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(3994..4013)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     CDS             join(4719..5309,1..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(4738..4757)
                     /label=LacI-R
                     /note="LacI, reverse primer"
ORIGIN
        1 ctggctggct ggcataaata tctcactcgc aatcaaattc agccgatagc ggaacgggaa
       61 ggcgactgga gtgccatgtc cggttttcaa caaaccatgc aaatgctgaa tgagggcatc
      121 gttcccactg 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 ggcgaattca ccgtctatgc cgtcaccggt cgtggcgatt
     2941 ccccggcaag ctcggcggca ggtggcggta tggaaactct gcatattacc aaaaccatga
     3001 aaaacattga agtgccggaa accaaaaccg cgagctttga atgcgaagtg agccatttta
     3061 acgtgccgag catgtggctg aaaaacggcg tggaaattga aatgagcgaa aaatttaaaa
     3121 ttgtggtgca gggcaaactg catcagctga ttattatgaa caccagcacc gaagatagcg
     3181 cggaatatac ctttgtgtgc ggcgttaccg gctatcgtct gtttgagaaa attagcaacg
     3241 atcaggtgag cgcgaccctg accgtgactt ccgagaatct gtacttcgag caaggtcacc
     3301 accatcatca ccattaagga tcccccccta gcataacccc ttggggcctc taaacgggtc
     3361 ttgaggggtt ttttgcccct gagacgcgtc aatcgagttc gtacctaagg gcgacacccc
     3421 ctaattagcc cgggcgaaag gcccagtctt tcgactgagc ctttcgtttt atttgatgcc
     3481 tggcagttcc ctactctcgc atggggagtc cccacactac catcggcgct acggcgtttc
     3541 acttctgagt tcggcatggg gtcaggtggg accaccgcgc tactgccgcc aggcaaacaa
     3601 ggggtgttat gagccatatt caggtataaa tgggctcgcg ataatgttca gaattggtta
     3661 attggttgta acactgaccc ctatttgttt atttttctaa atacattcaa atatgtatcc
     3721 gctcatgaga caataaccct gataaatgct tcaataatat tgaaaaagga agaatatgag
     3781 tattcaacat ttccgtgtcg cccttattcc cttttttgcg gcattttgcc ttcctgtttt
     3841 tgctcaccca gaaacgctgg tgaaagtaaa agatgctgaa gatcagttgg gtgcacgagt
     3901 gggttacatc gaactggatc tcaacagcgg taagatcctt gagagttttc gccccgaaga
     3961 acgttttcca atgatgagca cttttaaagt tctgctatgt ggcgcggtat tatcccgtat
     4021 tgacgccggg caagagcaac tcggtcgccg catacactat tctcagaatg acttggttga
     4081 gtactcacca gtcacagaaa agcatcttac ggatggcatg acagtaagag aattatgcag
     4141 tgctgccata accatgagtg ataacactgc ggccaactta cttctgacaa cgatcggagg
     4201 accgaaggag ctaaccgctt ttttgcacaa catgggggat catgtaactc gccttgatcg
     4261 ttgggaaccg gagctgaatg aagccatacc aaacgacgag cgtgacacca cgatgcctgt
     4321 agcgatggca acaacgttgc gcaaactatt aactggcgaa ctacttactc tagcttcccg
     4381 gcaacaatta atagactgga tggaggcgga taaagttgca ggaccacttc tgcgctcggc
     4441 ccttccggct ggctggttta ttgctgataa atccggagcc ggtgagcgtg gttctcgcgg
     4501 tatcatcgca gcgctggggc cagatggtaa gccctcccgt atcgtagtta tctacacgac
     4561 ggggagtcag gcaactatgg atgaacgaaa tagacagatc gctgagatag gtgcctcact
     4621 gattaagcat tggtaagcgg cgcgccatcg aatggcgcaa aacctttcgc ggtatggcat
     4681 gatagcgccc ggaagagagt caattcaggg tggtgaatat gaaaccagta acgttatacg
     4741 atgtcgcaga gtatgccggt gtctcttatc agaccgtttc ccgcgtggtg aaccaggcca
     4801 gccacgtttc tgcgaaaacg cgggaaaaag tggaagcggc gatggcggag ctgaattaca
     4861 ttcccaaccg cgtggcacaa caactggcgg gcaaacagtc gttgctgatt ggcgttgcca
     4921 cctccagtct ggccctgcac gcgccgtcgc aaattgtcgc ggcgattaaa tctcgcgccg
     4981 atcaactggg tgccagcgtg gtggtgtcga tggtagaacg aagcggcgtc gaagcctgta
     5041 aagcggcggt gcacaatctt ctcgcgcaac gcgtcagtgg gctgatcatt aactatccgc
     5101 tggatgacca ggatgccatt gctgtggaag ctgcctgcac taatgttccg gcgttatttc
     5161 ttgatgtctc tgaccagaca cccatcaaca gtattatttt ctcccatgag gacggtacgc
     5221 gactgggcgt ggagcatctg gtcgcattgg gtcaccagca aatcgcgctg ttagcgggcc
     5281 cattaagttc tgtctcggcg cgtctgcgt
//
