LOCUS       sequence-241917-        5654 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expressing Nuclear Localized GCaMP6s.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5654)
  TITLE     Leifer Lab worm expression plasmids
REFERENCE   2  (bases 1 to 5654)
  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"
COMMENT     Sequence Label: pAS1-his-24::GCaMP6s
FEATURES             Location/Qualifiers
     source          1..5654
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     58..80
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     72..89
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     73..89
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     99..118
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        99..117
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             join(821..829,830..847,848..850,851..883,884..886,887..910,
                     911..937,938..994,995..1000,1001..1270,1271..1288,
                     1289..1723,1724..1729,1730..2173)
                     /codon_start=1
                     /product="improved fluorescent protein-based calcium sensor
                     (Chen et al., 2013)"
                     /label=GCaMP6s
                     /translation="MGSHHHHHHGMASMTGGQQMGRDLYDDDDKDLATMVDSSRRKWNK
                     TGHAVRAIGRLSSLENVYIKADKQKNGIKANFHIRHNIEDGGVQLAYHYQQNTPIGDGP
                     VLLPDNHYLSVQSKLSKDPNEKRDHMVLLEFVTAAGITLGMDELYKGGTGGSMVSKGEE
                     LFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLKFICTTGKLPVPWPTLVTTLT
                     YGVQCFSRYPDHMKQHDFFKSAMPEGYIQERTIFFKDDGNYKTRAEVKFEGDTLVNRIE
                     LKGIDFKEDGNILGHKLEYNLPDQLTEEQIAEFKEAFSLFDKDGDGTITTKELGTVMRS
                     LGQNPTEAELQDMINEVDADGDGTIDFPEFLTMMARKMKYRDTEEEIREAFGVFDKDGN
                     GYISAAELRHVMTNLGEKLTDEEVDEMIREADIDGDGQVNYEEFVQMMTAK"
     regulatory      917..926
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     primer_bind     1207..1228
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     complement(1334..1355)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(1595..1614)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     complement(2939..2959)
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        complement(2939..2957)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(2978..2994)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2978..2994)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(2991..3013)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    3002..3018
                     /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)."
     promoter        complement(join(3026..3032,3033..3050,3051..3056))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3071..3092
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(3209..3226)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3380..3968)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3460..3479)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4139..4930,4931..4999))
                     /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     4762..4781
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5000..5104)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      complement(5130..5585)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(5267..5288)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     5479..5498
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
ORIGIN
        1 agctggcgaa agggggatgt gctgcaaggc gattaagttg ggtaacgcca gggttttccc
       61 agtcacgacg ttgtaaaacg acggccagtg agcgcgcgta atacgactca ctatagggcg
      121 aattggagct ccaccgcgga ctagtggatc ccccgggctg cagcctagga tgtctgattc
      181 cgctgttgtt gccgccgctg tcgagccaaa ggtcccaaag gctaaggccg ccaaggccgc
      241 caagccagcc gccaaggccg ccaagccaac caaagttgcc aaggccaagg ctccagtcgc
      301 tcacccacca tacatcaaca tgatcaagga ggccatcaag cagctcaagg accgcaaagg
      361 agcttccaag caggcaatcc tcaagttcat ctcccaaaac tacaagctcg gagacaatgt
      421 catccagatc aatgctcatc tccgtcaggc tctcaagcgt ggtgtcacca gcaaggccct
      481 tgttcaagcc gccggatccg gagccaacgg acgtttccgt gtgccagaga aggccgccgc
      541 cgccaagaag ccagcagcag ccaagaagcc agcagcagcc aagaaaccag cagccgccaa
      601 gaaagccact ggagagaaga aggcaaagaa gccagccgct gccaagccaa agaaggccgc
      661 caccggagat aagaaggtca agaaggccaa gagcccaaag aaggttgcca agccagctgc
      721 caagaaggtc gccaagtctc cagccaagaa ggccgctcca aagaagatcg ccaagccagc
      781 tgccaagaag gccgccaagc cagccgccaa ggccgcatgc atgggttctc atcatcatca
      841 tcatcatggt atggctagca tgactggtgg acagcaaatg ggtcgggatc tgtacgacga
      901 tgacgataag gatctcgcca ccatggtcga ctcatcacgt cgtaagtgga ataagacagg
      961 tcacgcagtc agagctatag gtcggctgag ctcactcgag aacgtctata tcaaggccga
     1021 caagcagaag aacggcatca aggcgaactt ccacatccgc cacaacatcg aggacggcgg
     1081 cgtgcagctc gcctaccact accagcagaa cacccccatc ggcgacggcc ccgtgctgct
     1141 gcccgacaac cactacctga gcgtgcagtc caaactttcg aaagacccca acgagaagcg
     1201 cgatcacatg gtcctgctgg agttcgtgac cgccgccggg atcactctcg gcatggacga
     1261 gctgtacaag ggcggtaccg gagggagcat ggtgagcaag ggcgaggagc tgttcaccgg
     1321 ggtggtgccc atcctggtcg agctggacgg cgacgtaaac ggccacaagt tcagcgtgtc
     1381 cggcgagggt gagggcgatg ccacctacgg caagctgacc ctgaagttca tctgcaccac
     1441 cggcaagctg cccgtgccct ggcccaccct cgtgaccacc ctgacctacg gcgtgcagtg
     1501 cttcagccgc taccccgacc acatgaagca gcacgacttc ttcaagtccg ccatgcccga
     1561 aggctacatc caggagcgca ccatcttctt caaggacgac ggcaactaca agacccgcgc
     1621 cgaggtgaag ttcgagggcg acaccctggt gaaccgcatc gagctgaagg gcatcgactt
     1681 caaggaggac ggcaacatcc tggggcacaa gctggagtac aacctgccgg accaactgac
     1741 tgaagagcag atcgcagaat ttaaagaggc tttctcccta tttgacaagg acggggatgg
     1801 gacaataaca accaaggagc tggggacggt gatgcggtct ctggggcaga accccacaga
     1861 agcagagctg caggacatga tcaatgaagt agatgccgac ggtgacggca caatcgactt
     1921 ccctgagttc ctgacaatga tggcaagaaa aatgaaatac agggacacgg aagaagaaat
     1981 tagagaagcg ttcggtgtgt ttgataagga tggcaatggc tacatcagtg cagcagagct
     2041 tcgccacgtg atgacaaacc ttggagagaa gttaacagat gaagaggttg atgaaatgat
     2101 cagggaagca gacatcgatg gggatggtca ggtaaactac gaagagtttg tacaaatgat
     2161 gacagcgaag tgaaagcttt ggaattcatc tcgcgcccgt gcctctgact tctaagtcca
     2221 attactcttc aacatcccta catgctcttt ctccctgtgc tcccaccccc tatttttgtt
     2281 attatcaaaa aacttctctt aatttctttg ttttttagct tcttttaagt cacctctaac
     2341 aatgaaattg tgtagattca aaaatagaat taattcgtaa taaaaagtcg aaaaaaattg
     2401 tgctccctcc ccccattaat aataattcta tcccaaaatc tacacaatgt tctgtgtaca
     2461 cttcttatgt tttttacttc tgataaattt ttttgaaaca tcatagaaaa aaccgcacac
     2521 aaaatacctt atcatatgtt acgtttcagt ttatgaccgc aatttttatt tcttcgcacg
     2581 tctgggcctc tcatgacgtc aaatcatgct catcgtgaaa aagttttgga gtatttttgg
     2641 aatttttcaa tcaagtgaaa gtttatgaaa ttaattttcc tgcttttgct ttttggggtt
     2701 tcccctattg tttgtcaaga tttcgaggac ggcgtttttc ttgctaaaat cacaagtatt
     2761 gatgagcacg atgcaagaaa gatcggaaga aggtttgggt ttgaggctca gtggaaggtg
     2821 agtagaagtt gataatttga aagtggagta gtgtctatgg ggtttttgcc ttaaatgaca
     2881 gaatacattc ccaatatacc aaacataact gtttgggccc ggtacccagc ttttgttccc
     2941 tttagtgagg gttaattgcg cgcttggcgt aatcatggtc atagctgttt cctgtgtgaa
     3001 attgttatcc gctcacaatt ccacacaaca tacgagccgg gagcataaag tgtaaagcct
     3061 ggggtgccta atgagtgagc taactcacat taattgcgtt gcgctcactg cccgctttcc
     3121 agtcgggaaa cctgtcgtgc cagctgcatt aatgaatcgg ccaacgcgcg gggagaggcg
     3181 gtttgcgtat tgggcgctct tccgcttcct cgctcactga ctcgctgcgc tcggtcgttc
     3241 ggctgcggcg agcggtatca gctcactcaa aggcggtaat acggttatcc acagaatcag
     3301 gggataacgc aggaaagaac atgtgagcaa aaggccagca aaaggccagg aaccgtaaaa
     3361 aggccgcgtt gctggcgttt ttccataggc tccgcccccc tgacgagcat cacaaaaatc
     3421 gacgctcaag tcagaggtgg cgaaacccga caggactata aagataccag gcgtttcccc
     3481 ctggaagctc cctcgtgcgc tctcctgttc cgaccctgcc gcttaccgga tacctgtccg
     3541 cctttctccc ttcgggaagc gtggcgcttt ctcatagctc acgctgtagg tatctcagtt
     3601 cggtgtaggt cgttcgctcc aagctgggct gtgtgcacga accccccgtt cagcccgacc
     3661 gctgcgcctt atccggtaac tatcgtcttg agtccaaccc ggtaagacac gacttatcgc
     3721 cactggcagc agccactggt aacaggatta gcagagcgag gtatgtaggc ggtgctacag
     3781 agttcttgaa gtggtggcct aactacggct acactagaag aacagtattt ggtatctgcg
     3841 ctctgctgaa gccagttacc ttcggaaaaa gagttggtag ctcttgatcc ggcaaacaaa
     3901 ccaccgctgg tagcggtggt ttttttgttt gcaagcagca gattacgcgc agaaaaaaag
     3961 gatctcaaga agatcctttg atcttttcta cggggtctga cgctcagtgg aacgaaaact
     4021 cacgttaagg gattttggtc atgagattat caaaaaggat cttcacctag atccttttaa
     4081 attaaaaatg aagttttaaa tcaatctaaa gtatatatga gtaaacttgg tctgacagtt
     4141 accaatgctt aatcagtgag gcacctatct cagcgatctg cctatttcgt tcatccatag
     4201 ttgcctgact ccccgtcgtg tagataacta cgatacggga gggcttacca tctggcccca
     4261 gtgctgcaat gataccgcga gacccacgct caccggctcc agatttatca gcaataaacc
     4321 agccagccgg aagggccgag cgcagaagtg gtcctgcaac tttatccgcc tccatccagt
     4381 ctattaattg ttgccgggaa gctagagtaa gtagttcgcc agttaatagt ttgcgcaacg
     4441 ttgttgccat tgctacaggc atcgtggtgt cacgctcgtc gtttggtatg gcttcattca
     4501 gctccggttc ccaacgatca aggcgagtta catgatcccc catgttgtgc aaaaaagcgg
     4561 ttagctcctt cggtcctccg atcgttgtca gaagtaagtt ggccgcagtg ttatcactca
     4621 tggttatggc agcactgcat aattctctta ctgtcatgcc atccgtaaga tgcttttctg
     4681 tgactggtga gtactcaacc aagtcattct gagaatagtg tatgcggcga ccgagttgct
     4741 cttgcccggc gtcaatacgg gataataccg cgccacatag cagaacttta aaagtgctca
     4801 tcattggaaa acgttcttcg gggcgaaaac tctcaaggat cttaccgctg ttgagatcca
     4861 gttcgatgta acccactcgt gcacccaact gatcttcagc atcttttact ttcaccagcg
     4921 tttctgggtg agcaaaaaca ggaaggcaaa atgccgcaaa aaagggaata agggcgacac
     4981 ggaaatgttg aatactcata ctcttccttt ttcaatatta ttgaagcatt tatcagggtt
     5041 attgtctcat gagcggatac atatttgaat gtatttagaa aaataaacaa ataggggttc
     5101 cgcgcacatt tccccgaaaa gtgccaccta aattgtaagc gttaatattt tgttaaaatt
     5161 cgcgttaaat ttttgttaaa tcagctcatt ttttaaccaa taggccgaaa tcggcaaaat
     5221 cccttataaa tcaaaagaat agaccgagat agggttgagt gttgttccag tttggaacaa
     5281 gagtccacta ttaaagaacg tggactccaa cgtcaaaggg cgaaaaaccg tctatcaggg
     5341 cgatggccca ctacgtgaac catcacccta atcaagtttt ttggggtcga ggtgccgtaa
     5401 agcactaaat cggaacccta aagggagccc ccgatttaga gcttgacggg gaaagccggc
     5461 gaacgtggcg agaaaggaag ggaagaaagc gaaaggagcg ggcgctaggg cgctggcaag
     5521 tgtagcggtc acgctgcgcg taaccaccac acccgccgcg cttaatgcgc cgctacaggg
     5581 cgcgtcccat tcgccattca ggctgcgcaa ctgttgggaa gggcgatcgg tgcgggcctc
     5641 ttcgctatta cgcc
//
