LOCUS       sequence-388569-        6739 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Protein production of the split HaloTag based calcium recorder 
            Caprola_11 in bacteria.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6739)
  AUTHORS   Huppertz MC, Wilhelm J, Grenier V, Schneider MW, Falt T, Porzberg N,
            Hausmann D, Hoffmann DC, Hai L, Tarnawski M, Pino G, Slanchev K, 
            Kolb I, Acuna C, Fenk LM, Baier H, Hiblot J, Johnsson K
  TITLE     Recording physiological history of cells with chemical labeling.
  JOURNAL   Science. 2024 Feb 23;383(6685):890-897. doi: 
            10.1126/science.adg0812. Epub 2024 Feb 22.
  PUBMED    38386755
REFERENCE   2  (bases 1 to 6739)
  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: 
            "10.1126/science.adg0812"; journalName: "Science"; date: 
            "2024-02-23- 23"; volume: "383"; issue: "6685"; pages: "890-897"
COMMENT     Sequence Label: pET-51b(+)-Caprola_11
FEATURES             Location/Qualifiers
     source          1..6739
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             1064..1093
                     /codon_start=1
                     /product="10xHis affinity tag"
                     /label=10xHis
                     /translation="HHHHHHHHHH"
     primer_bind     complement(1116..1134)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      1130..1177
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     rep_origin      1214..1669
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1301..1320)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     1511..1532
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     CDS             complement(join(1784..2575,2576..2644))
                     /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     2407..2426
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(2645..2737)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      2818..3406
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3307..3326
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     3560..3577
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    3592..3731
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(3657..3676)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     3817..3839
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(3833..4024)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     CDS             complement(4596..5678)
                     /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
                     EDGTRLGVEHLVALGHQQIALLAGPLSSVSARLRLAGWHKYITRNQIQPIAEREGDWSA
                     MSGFQQTMQMLNEGIVPTAMLVANDQMALGAMRAITESGLRVGADISVVGYDDTEDSSC
                     YIPPLTTIKQDFRLLGQTSVDRLLQLSQGQAVKGNQLLPVSLVKRKTTLAPNTQTASPR
                     ALADSLMQLARQVSRLESGQ"
     primer_bind     5640..5659
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(5679..5756)
                     /gene="lacI"
                     /label=lacI promoter
     primer_bind     5962..5981
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     primer_bind     6069..6088
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        6069..6087
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    6088..6112
                     /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             6127..6149
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             6165..6188
                     /codon_start=1
                     /product="peptide that binds Strep-Tactin(R), an engineered
                     form of streptavidin"
                     /label=Strep-Tag II
                     /translation="WSHPQFEK"
     CDS             6195..6209
                     /codon_start=1
                     /product="enterokinase recognition and cleavage site"
                     /label=enterokinase site
                     /translation="DDDDK"
     CDS             6264..6704
                     /codon_start=1
                     /product="calcium-binding protein"
                     /label=calmodulin
                     /translation="DQLTEEQIAEFKEAFSLFDKDGDGTITTKELGTVMRSLGQNPTEA
                     ELQDMINEVDADGDGTIDFPEFLTMMARKMKDTDSEEEIREAFRVFDKDGNGYISAAEL
                     RHVMTNLGEKLTDEEVDEMIREADIDGDGQVNYEEFVVMMTAK"
ORIGIN
        1 gccgccgcca ccaccacctc ctccgcctcc tccaccacca ccgccaccgc ctccaccacc
       61 tggcggcagc agggtcgact catcacgtcg taagtggaat aagacagatc acgcactgag
      121 agctataggt cggctgagct cactcgaggg cggttctgac gtcggccgca agctgatcat
      181 cgatcagaac gtttttatcg agggtacgct gccgatgggt gtcgtccgcc cgctgactga
      241 agtcgagatg gaccattacc gcgagccgtt cctgaatcct gttgaccgcg agccactgtg
      301 gcgcttccca aacgagctgc caatcgccgg tgagccagcg aacatcgtcg cgctggtcga
      361 agaatacatg gactggctgc accagtcccc tgtcccgaag ctgctgttct ggggcacccc
      421 aggcgttctg atcccaccgg ccgaagccgc tcgcctggcc aaaagcctgc ctaactgcaa
      481 ggctgtggac atcggcccgg gtctgaatct gctgcaagaa gacaacccgg acctgatcgg
      541 cagcgagatc gcgcgctggc tgtctactct ggagattggt ggcaccggtg gctctggcgg
      601 taccggtggc tctggcggtt ctatcggtac tggctttcca ttcgaccccc attatgtgga
      661 agtcctgggc gagcgcatgc actacgtcga tgttggtccg cgcgatggca cccctgtgct
      721 gttcctgcac ggtaacccga cctcctccta cgtgtggcgc aacatcatcc cacatgttgc
      781 accgacccat cgctgcattg ctccagacct gatcggtatg ggcaaatccg acaaaccaga
      841 cctgggttat ttcttcgacg accacgtccg cttcatggat gccttcatcg aagccctggg
      901 tctggaagag gtcgtcctgg tcattcacga ctggggctcc gctctgggtt tccactgggc
      961 caagcgcaat ccagagcgcg tcaaaggtat tgcatttatg gagttcatcc gccctatccc
     1021 gacctgggac gaatgggctc cgggcttctc ctcaatttcc gctcatcacc accatcatca
     1081 ccatcaccac cactaattaa cctaggctgc tgccaccgct gagcaataac tagcataacc
     1141 ccttggggcc tctaaacggg tcttgagggg ttttttgctg aaaggaggaa ctatatccgg
     1201 attggcgaat gggacgcgcc ctgtagcggc gcattaagcg cggcgggtgt ggtggttacg
     1261 cgcagcgtga ccgctacact tgccagcgcc ctagcgcccg ctcctttcgc tttcttccct
     1321 tcctttctcg ccacgttcgc cggctttccc cgtcaagctc taaatcgggg gctcccttta
     1381 gggttccgat ttagtgcttt acggcacctc gaccccaaaa aacttgatta gggtgatggt
     1441 tcacgtagtg ggccatcgcc ctgatagacg gtttttcgcc ctttgacgtt ggagtccacg
     1501 ttctttaata gtggactctt gttccaaact ggaacaacac tcaaccctat ctcggtctat
     1561 tcttttgatt tataagggat tttgccgatt tcggcctatt ggttaaaaaa tgagctgatt
     1621 taacaaaaat ttaacgcgaa ttttaacaaa atattaacgt ttacaatttc tggcggcacg
     1681 atggcatgag attatcaaaa aggatcttca cctagatcct tttaaattaa aaatgaagtt
     1741 ttaaatcaat ctaaagtata tatgagtaaa cttggtctga cagttaccaa tgcttaatca
     1801 gtgaggcacc tatctcagcg atctgtctat ttcgttcatc catagttgcc tgactccccg
     1861 tcgtgtagat aactacgata cgggagggct taccatctgg ccccagtgct gcaatgatac
     1921 cgcgagaccc acgctcaccg gctccagatt tatcagcaat aaaccagcca gccggaaggg
     1981 ccgagcgcag aagtggtcct gcaactttat ccgcctccat ccagtctatt aattgttgcc
     2041 gggaagctag agtaagtagt tcgccagtta atagtttgcg caacgttgtt gccattgcta
     2101 caggcatcgt ggtgtcacgc tcgtcgtttg gtatggcttc attcagctcc ggttcccaac
     2161 gatcaaggcg agttacatga tcccccatgt tgtgcaaaaa agcggttagc tccttcggtc
     2221 ctccgatcgt tgtcagaagt aagttggccg cagtgttatc actcatggtt atggcagcac
     2281 tgcataattc tcttactgtc atgccatccg taagatgctt ttctgtgact ggtgagtact
     2341 caaccaagtc attctgagaa tagtgtatgc ggcgaccgag ttgctcttgc ccggcgtcaa
     2401 tacgtgataa taccgcgcca catagcagaa ctttaaaagt gctcatcatt ggaaaacgtt
     2461 cttcggggcg aaaactctca aggatcttac cgctgttgag atccagttcg atgtaaccca
     2521 ctcgtgcacc caactgatct tcagcatctt ttactttcac cagcgtttct gggtgagcaa
     2581 aaacaggaag gcaaaatgcc gcaaaaaagg gaataagggc gacacggaaa tgttgaatac
     2641 tcatactctt cctttttcaa tcatgattga agcatttatc agggttattg tctcatgagc
     2701 ggatacatat ttgaatgtat ttagaaaaat aaacaaatag gtcatgacca aaatccctta
     2761 acgtgagttt tcgttccact gagcgtcaga ccccgtagaa aagatcaaag gatcttcttg
     2821 agatcctttt tttctgcgcg taatctgctg cttgcaaaca aaaaaaccac cgctaccagc
     2881 ggtggtttgt ttgccggatc aagagctacc aactcttttt ccgaaggtaa ctggcttcag
     2941 cagagcgcag ataccaaata ctgtccttct agtgtagccg tagttaggcc accacttcaa
     3001 gaactctgta gcaccgccta catacctcgc tctgctaatc ctgttaccag tggctgctgc
     3061 cagtggcgat aagtcgtgtc ttaccgggtt ggactcaaga cgatagttac cggataaggc
     3121 gcagcggtcg ggctgaacgg ggggttcgtg cacacagccc agcttggagc gaacgaccta
     3181 caccgaactg agatacctac agcgtgagct atgagaaagc gccacgcttc ccgaagggag
     3241 aaaggcggac aggtatccgg taagcggcag ggtcggaaca ggagagcgca cgagggagct
     3301 tccaggggga aacgcctggt atctttatag tcctgtcggg tttcgccacc tctgacttga
     3361 gcgtcgattt ttgtgatgct cgtcaggggg gcggagccta tggaaaaacg ccagcaacgc
     3421 ggccttttta cggttcctgg ccttttgctg gccttttgct cacatgttct ttcctgcgtt
     3481 atcccctgat tctgtggata accgtattac cgcctttgag tgagctgata ccgctcgccg
     3541 cagccgaacg accgagcgca gcgagtcagt gagcgaggaa gcggaagagc gcctgatgcg
     3601 gtattttctc cttacgcatc tgtgcggtat ttcacaccgc aatggtgcac tctcagtaca
     3661 atctgctctg atgccgcata gttaagccag tatacactcc gctatcgcta cgtgactggg
     3721 tcatggctgc gccccgacac ccgccaacac ccgctgacgc gccctgacgg gcttgtctgc
     3781 tcccggcatc cgcttacaga caagctgtga ccgtctccgg gagctgcatg tgtcagaggt
     3841 tttcaccgtc atcaccgaaa cgcgcgaggc agctgcggta aagctcatca gcgtggtcgt
     3901 gaagcgattc acagatgtct gcctgttcat ccgcgtccag ctcgttgagt ttctccagaa
     3961 gcgttaatgt ctggcttctg ataaagcggg ccatgttaag ggcggttttt tcctgtttgg
     4021 tcactgatgc ctccgtgtaa gggggatttc tgttcatggg ggtaatgata ccgatgaaac
     4081 gagagaggat gctcacgata cgggttactg atgatgaaca tgcccggtta ctggaacgtt
     4141 gtgagggtaa acaactggcg gtatggatgc ggcgggacca gagaaaaatc actcagggtc
     4201 aatgccagcg cttcgttaat acagatgtag gtgttccaca gggtagccag cagcatcctg
     4261 cgatgcagat ccggaacata atggtgcagg gcgctgactt ccgcgtttcc agactttacg
     4321 aaacacggaa accgaagacc attcatgttg ttgctcaggt cgcagacgtt ttgcagcagc
     4381 agtcgcttca cgttcgctcg cgtatcggtg attcattctg ctaaccagta aggcaacccc
     4441 gccagcctag ccgggtcctc aacgacagga gcacgatcat gctagtcatg ccccgcgccc
     4501 accggaagga gctgactggg ttgaaggctc tcaagggcat cggtcgagat cccggtgcct
     4561 aatgagtgag ctaacttaca ttaattgcgt tgcgctcact gcccgctttc cagtcgggaa
     4621 acctgtcgtg ccagctgcat taatgaatcg gccaacgcgc ggggagaggc ggtttgcgta
     4681 ttgggcgcca gggtggtttt tcttttcacc agtgagacgg gcaacagctg attgcccttc
     4741 accgcctggc cctgagagag ttgcagcaag cggtccacgc tggtttgccc cagcaggcga
     4801 aaatcctgtt tgatggtggt taacggcggg atataacatg agctgtcttc ggtatcgtcg
     4861 tatcccacta ccgagatgtc cgcaccaacg cgcagcccgg actcggtaat ggcgcgcatt
     4921 gcgcccagcg ccatctgatc gttggcaacc agcatcgcag tgggaacgat gccctcattc
     4981 agcatttgca tggtttgttg aaaaccggac atggcactcc agtcgccttc ccgttccgct
     5041 atcggctgaa tttgattgcg agtgatatat ttatgccagc cagccagacg cagacgcgcc
     5101 gagacagaac ttaatgggcc cgctaacagc gcgatttgct ggtgacccaa tgcgaccaga
     5161 tgctccacgc ccagtcgcgt accgtcttca tgggagaaaa taatactgtt gatgggtgtc
     5221 tggtcagaga catcaagaaa taacgccgga acattagtgc aggcagcttc cacagcaatg
     5281 gcatcctggt catccagcgg atagttaatg atcagcccac tgacgcgttg cgcgagaaga
     5341 ttgtgcaccg ccgctttaca ggcttcgacg ccgcttcgtt ctaccatcga caccaccacg
     5401 ctggcaccca gttgatcggc gcgagattta atcgccgcga caatttgcga cggcgcgtgc
     5461 agggccagac tggaggtggc aacgccaatc agcaacgact gtttgcccgc cagttgttgt
     5521 gccacgcggt tgggaatgta attcagctcc gccatcgccg cttccacttt ttcccgcgtt
     5581 ttcgcagaaa cgtggctggc ctggttcacc acgcgggaaa cggtctgata agagacaccg
     5641 gcatactctg cgacatcgta taacgttact ggtttcacat tcaccaccct gaattgactc
     5701 tcttccgggc gctatcatgc cataccgcga aaggttttgc gccattcgat ggtgtccggg
     5761 atctcgacgc tctcccttat gcgactcctg cattaggaag cagcccagta gtaggttgag
     5821 gccgttgagc accgccgccg caaggaatgg tgcatgcaag gagatggcgc ccaacagtcc
     5881 cccggccacg gggcctgcca ccatacccac gccgaaacaa gcgctcatga gcccgaagtg
     5941 gcgagcccga tcttccccat cggtgatgtc ggcgatatag gcgccagcaa ccgcacctgt
     6001 ggcgccggtg atgccggcca cgatgcgtcc ggcgtagagg atcgagatcg atctcgatcc
     6061 cgcgaaatta atacgactca ctatagggga attgtgagcg gataacaatt cccctctaga
     6121 aataattttg tttaacttta agaaggagat ataccatggc aagctggagc cacccgcagt
     6181 tcgaaaaggg tgcagatgac gacgacaagg taccgcatgg cggttctgcc cgcgagacct
     6241 tccaggcctt ccgcaccggt tctgaccaac tgactgaaga gcagatcgca gaatttaaag
     6301 aggctttctc cctatttgac aaggacgggg atgggacaat aacaaccaag gagctgggga
     6361 cggtgatgcg gtctctgggg cagaacccca cagaagcaga gctgcaggac atgatcaatg
     6421 aagtagatgc cgacggtgac ggcacaatcg acttccctga gttcctgaca atgatggcaa
     6481 gaaaaatgaa agacacagac agtgaagaag aaattagaga agcgttccgt gtgtttgata
     6541 aggatggcaa tggctacatc agtgcagcag agcttcgcca cgtgatgaca aaccttggag
     6601 agaagttaac agatgaagag gttgatgaaa tgatcaggga agcagacatc gatggggatg
     6661 gtcaggtaaa ctacgaagag tttgtagtga tgatgacagc gaaggaattc cctcctcctc
     6721 cgccaccgcc accaccgcc
//
