LOCUS       sequence-405003-        5019 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expression of WHaloCaMP-eNOSpep-EGFP fusion in E. coli.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5019)
  AUTHORS   Farrants H, Shuai Y, Lemon WC, Hernandez CM, Yang S, Patel R, Qiao 
            G, Frei MS, Grimm JB, Hanson TL, Tomaska F, Turner GC, Stringer C, 
            Keller PJ, Beyene AG, Chen Y, Liang Y, Lavis LD, Schreiter ER
  TITLE     A modular chemigenetic calcium indicator enables in vivo functional 
            imaging with near-infrared light.
  JOURNAL   bioRxiv. 2023 Jul 19:2023.07.18.549527. doi: 
            10.1101/2023.07.18.549527. Preprint.
  PUBMED    37503182
REFERENCE   2  (bases 1 to 5019)
  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: "bioRxiv. 
            2023 Jul 19:2023.07.18.549527. doi: 10.1101/2023.07.18.549527. 
            Preprint."
COMMENT     Sequence Label: pRSET-WHaloCaMP-eNOSpep-EGFP
FEATURES             Location/Qualifiers
     source          1..5019
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     20..39
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        20..38
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     RBS             70..92
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             163..180
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             820..1260
                     /codon_start=1
                     /product="calcium-binding protein"
                     /label=calmodulin
                     /translation="DQLTEEQIAEFKEAFSLFDKDGDGTITTKELGTVMRSLGQNPTEA
                     ELQDMINEVDADGNGTIDFPEFLTMMARKMKDTDSEEEIREAFRVFDKDGNGYISAAEL
                     RHVMTNLGEKLTDEEVDEMIREADIDGDGQVNYEEFVQMMTAK"
     CDS             join(1627..1629,1630..1632,1633..2343)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(2420..2438)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      2434..2481
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     rep_origin      2578..3033
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2665..2684)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2875..2896
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        3059..3163
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(3164..3232,3233..4024)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIKHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     complement(3382..3401)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      4195..4783
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4684..4703
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     4937..4954
                     /label=L4440
                     /note="L4440 vector, forward primer"
ORIGIN
        1 gatctcgatc ccgcgaaatt aatacgactc actataggga gaccacaacg gtttccctct
       61 agaaataatt ttgtttaact ttaagaagga gatatacata tgctgcagaa cgagcttgct
      121 cttaagttgg ctggacttga tattaacaag actggaggtt ctcatcatca ccaccaccat
      181 ggatccatgg cagaaatcgg tactggcttt ccattcgacc cccattatgt ggaagtcctg
      241 ggcgagcgca tgcactacgt cgatgttggt ccgcgcgatg gcacccctgt gctgttcctg
      301 cacggtaacc cgacctcctc ctacgtgtgg cgcaacatca tcccgcatgt tgcaccgacc
      361 catcgctgca ttgctccaga cctgatcggt atgggcaaat ccgacaaacc agacctgggt
      421 tatttcttcg acgaccacgt ccgcttcatg gatgccttca tcgaagccct gggtctggaa
      481 gaggtcgtcc tggtcattca cgactggggc tccgctctgg gtttccactg ggccaagcgc
      541 aatccagagc gcgtcaaagg tattgcattt atggagttca tccgccctat cccgacctgg
      601 gacgaatggc cagaatttgc acgcgagacc ttccaggcct tccgcaccac cgacgtcggc
      661 cgcaagctga tcatcgatca gaacgttttt atcgagtgga cgctgccgat ggcagtcatt
      721 cgccgcaaga agaccttcaa ggaggtggcc aacgccgtga agatctctgc ctctctgatg
      781 ggcggaggct ctggcggttc cggcggttca ggtggatcag atcaattaac agaggaacag
      841 attgcggagt ttaaggaagc gttctcttta tttgataagg atggcgacgg tacaatcact
      901 actaaagaat tgggaacagt catgcgctca ttggggcaaa atccgacaga ggctgaattg
      961 caggacatga ttaacgaggt agacgccgat gggaacggga ctatcgactt tccggaattt
     1021 cttactatga tggcacgcaa aatgaaagat accgattctg aagaagagat ccgtgaagcc
     1081 ttccgtgttt ttgataaaga tgggaacggc tacatcagtg ctgctgagtt acgccatgtg
     1141 atgacaaatc tgggggaaaa acttaccgac gaagaagtag acgaaatgat tcgcgaggcg
     1201 gatattgacg gggatggaca agtaaactac gaggaatttg tgcagatgat gaccgccaag
     1261 acgctgactg aagtcgagat ggaccattac cgcgagccgt tcctgaatcc tgttgaccgc
     1321 gagccactgt ggcgcttccc aaacgagctg ccaatcgccg gtgagccagc gaacatcgtc
     1381 gcgctggtcg aagaatacat ggactggctg caccagtccc ctgtcccgaa gctgctgttc
     1441 tggggcaccc caggcgttct gatcccaccg gccgaagccg ctcgcctggc caaaagcctg
     1501 cctaactgca aggctgtgga catcggcccg ggtctgaatc tgctgcaaga agacaacccg
     1561 gacctgatcg gcagcgagat cgcgcgctgg ctgtcgacgc tcgagatttc cggcggcgga
     1621 agcggcatgg tctctaaggg ggaggaactc tttactggtg tggtgccaat acttgtagag
     1681 ctcgatgggg acgttaatgg acacaaattt tctgtctcag gtgagggtga gggagatgca
     1741 acttatggaa aactgacatt gaaattcatc tgcacaacag gcaaattgcc tgtcccctgg
     1801 cctaccctgg taaccaccct cacttatggc gttcagtgct tctcccggta tcctgatcac
     1861 atgaaacagc atgatttctt caaatcagca atgcccgaag gttatgttca agagcggacc
     1921 atctttttca aggacgatgg aaactataaa accagagctg aggttaagtt cgagggagat
     1981 actttggtga atcggatcga actgaagggg atagacttca aagaggatgg gaacattttg
     2041 gggcataaac tcgaatataa ctacaactcc cacaacgtgt atatcatggc tgacaaacag
     2101 aagaacggga tcaaagtaaa ttttaagata cgacacaata tagaggacgg tagtgtccaa
     2161 cttgctgatc actaccagca gaatacaccc atcggagacg gacccgttct cttgcccgat
     2221 aatcactatc tctctacaca atccgctctg tctaaagatc caaacgaaaa gcgggaccac
     2281 atggtactgc ttgagttcgt tacagccgct ggtatcacct tgggcatgga tgagttgtat
     2341 aaataagaat tcgaagcttg atccggctgc taacggtctc gactatacgc ccgaaaggaa
     2401 gctgagttgg ctgctgccac cgctgagcaa taactagcat aaccccttgg ggcctctaaa
     2461 cgggtcttga ggggtttttt gctgaaagga ggaactatat ccggatctgg cgtaatagcg
     2521 aagaggcccg caccgatcgc ccttcccaac agttgcgcag cctgaatggc gaatgggacg
     2581 cgccctgtag cggcgcatta agcgcggcgg gtgtggtggt tacgcgcagc gtgaccgcta
     2641 cacttgccag cgccctagcg cccgctcctt tcgctttctt cccttccttt ctcgccacgt
     2701 tcgccggctt tccccgtcaa gctctaaatc gggggctccc tttagggttc cgatttagtg
     2761 ctttacggca cctcgacccc aaaaaacttg attagggtga tggttcacgt agtgggccat
     2821 cgccctgata gacggttttt cgccctttga cgttggagtc cacgttcttt aatagtggac
     2881 tcttgttcca aactggaaca acactcaacc ctatctcggt ctattctttt gatttataag
     2941 ggattttgcc gatttcggcc tattggttaa aaaatgagct gatttaacaa aaatttaacg
     3001 cgaattttaa caaaatatta acgcttacaa tttaggtggc acttttcggg gaaatgtgcg
     3061 cggaacccct atttgtttat ttttctaaat acattcaaat atgtatccgc tcatgagaca
     3121 acaaccctga taaatgcttc aataatattg aaaaaggaag agtatgagta ttaaacattt
     3181 ccgtgtcgcc cttattccct tttttgcggc attttgcctt cctgtttttg ctcacccaga
     3241 aacgctggtg aaagtaaaag atgctgaaga tcagttgggt gcacgagtgg gttacatcga
     3301 actggatctc aacagcggta agatccttga gagttttcgc cccgaagaac gttttccaat
     3361 gatgagcact tttaaagttc tgctatgtgg cgcggtatta tcccgtattg acgccgggca
     3421 agagcaactc ggtcgccgca tacactattc tcagaatgac ttggttgagt actcaccagt
     3481 cacagaaaag catcttacgg atggcatgac agtaagagaa ttatgcagtg ctgccataac
     3541 catgagtgat aacactgcgg ccaacttact tctgacaacg atcggaggac cgaaggagct
     3601 aaccgctttt ttgcacaaca tgggggatca tgtaactcgc cttgatcgtt gggaaccgga
     3661 gctgaatgaa gccataccaa acgacgagcg tgacaccacg atgcctgtag caatggcaac
     3721 aacgttgcgc aaactattaa ctggcgaact acttactcta gcttcccggc aacaattaat
     3781 agactggatg gaggcggata aagttgcagg accacttctg cgctcggccc ttccggctgg
     3841 ctggtttatt gctgataaat ctggagccgg tgagcgtggg tctcgcggta tcattgcagc
     3901 actggggcca gatggtaagc cctcccgtat cgtagttatc tacacgacgg ggagtcaggc
     3961 aactatggat gaacgaaata gacagatcgc tgagataggt gcctcactga ttaagcattg
     4021 gtaactgtca gaccaagttt actcatatat actttagatt gatttaaaac ttcattttta
     4081 atttaaaagg atctaggtga agatcctttt tgataatctc atgaccaaaa tcccttaacg
     4141 tgagttttcg ttccactgag cgtcagaccc cgtagaaaag atcaaaggat cttcttgaga
     4201 tccttttttt ctgcgcgtaa tctgctgctt gcaaacaaaa aaaccaccgc taccagcggt
     4261 ggtttgtttg ccggatcaag agctaccaac tctttttccg aaggtaactg gcttcagcag
     4321 agcgcagata ccaaatactg ttcttctagt gtagccgtag ttaggccacc acttcaagaa
     4381 ctctgtagca ccgcctacat acctcgctct gctaatcctg ttaccagtgg ctgctgccag
     4441 tggcgataag tcgtgtctta ccgggttgga ctcaagacga tagttaccgg ataaggcgca
     4501 gcggtcgggc tgaacggggg gttcgtgcac acagcccagc ttggagcgaa cgacctacac
     4561 cgaactgaga tacctacagc gtgagctatg agaaagcgcc acgcttcccg aagggagaaa
     4621 ggcggacagg tatccggtaa gcggcagggt cggaacagga gagcgcacga gggagcttcc
     4681 agggggaaac gcctggtatc tttatagtcc tgtcgggttt cgccacctct gacttgagcg
     4741 tcgatttttg tgatgctcgt caggggggcg gagcctatgg aaaaacgcca gcaacgcggc
     4801 ctttttacgg ttcctggcct tttgctggcc ttttgctcac atgttctttc ctgcgttatc
     4861 ccctgattct gtggataacc gtattaccgc ctttgagtga gctgataccg ctcgccgcag
     4921 ccgaacgacc gagcgcagcg agtcagtgag cgaggaagcg gaagagcgcc caatacgcaa
     4981 accgcctctc cccgcgcgtt ggccgattca ttaatgcag
//
