LOCUS Exported 5819 bp ds-DNA circular SYN 12-MAY-2021 DEFINITION Fluorescent reporter for calcium signaling. ACCESSION . VERSION . KEYWORDS pZac2.1 gfaABC1D-cyto-GCaMP6f SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 5819) AUTHORS Haustein MD, Kracun S, Lu XH, Shih T, Jackson-Weaver O, Tong X, Xu J, Yang XW, O'Dell TJ, Marvin JS, Ellisman MH, Bushong EA, Looger LL, Khakh BS TITLE Conditions and constraints for astrocyte calcium signaling in the hippocampal mossy fiber pathway. JOURNAL Neuron. 2014 Apr 16;82(2):413-29. doi: 10.1016/j.neuron.2014.02.041. PUBMED 24742463 REFERENCE 2 (bases 1 to 5819) AUTHORS . TITLE Direct Submission JOURNAL Exported May 12, 2021 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..5819 /organism="synthetic DNA construct" /mol_type="other DNA" protein_bind 107..128 /label=CAP binding site /bound_moiety="E. coli catabolite activator protein" /note="CAP binding activates transcription in the presence of cAMP." promoter 143..173 /label=lac promoter /note="promoter for the E. coli lac operon" protein_bind 181..197 /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)." primer_bind 186..208 /label=M13/pUC Reverse /note="In lacZ gene" primer_bind 205..221 /label=M13 rev /note="common sequencing primer, one of multiple similar variants" primer_bind 205..221 /label=M13 Reverse /note="In lacZ gene. Also called M13-rev" repeat_region 259..388 /label=AAV2 ITR intron 1239..1371 /label=chimeric intron /note="chimera between introns from human beta-globin and immunoglobulin heavy chain genes" primer_bind 1354..1373 /label=pMT2-F /note="Synthetic intron, forward primer" promoter 1416..1434 /label=T7 promoter /note="promoter for bacteriophage T7 RNA polymerase" regulatory 1440..1449 /regulatory_class="other" /note="vertebrate consensus sequence for strong initiation of translation (Kozak, 1987)" CDS 1455..1472 /codon_start=1 /product="6xHis affinity tag" /label=6xHis /translation="HHHHHH" CDS 1476..1508 /codon_start=1 /product="leader peptide from bacteriophage T7 gene 10" /label=T7 tag (gene 10 leader) /note="promotes efficient translation in E. coli" /translation="MASMTGGQQMG" CDS 1512..1535 /codon_start=1 /product="Xpress(TM) epitope tag, including an enterokinase recognition and cleavage site" /label=Xpress(TM) tag /translation="DLYDDDDK" regulatory 1548..1557 /regulatory_class="other" /note="vertebrate consensus sequence for strong initiation of translation (Kozak, 1987)" CDS 1569..1625 /codon_start=1 /product="calmodulin-binding peptide from avian smooth muscle myosin light chain kinase" /label=calmodulin-binding peptide /translation="RRKWNKTGHAVRAIGRLSS" primer_bind 1838..1859 /label=EGFP-C /note="EGFP, forward primer" primer_bind complement(1965..1986) /label=EGFP-N /note="EGFP, reverse primer" primer_bind complement(2226..2245) /label=EXFP-R /note="For distinguishing EGFP variants, reverse primer" polyA_signal 2844..2965 /label=SV40 poly(A) signal /note="SV40 polyadenylation signal" primer_bind complement(2855..2874) /label=EBV-rev /note="SV40 polyA terminator, reverse primer" primer_bind 2909..2928 /label=SV40pA-R /note="SV40 polyA, reverse primer" repeat_region 3127..3256 /label=AAV2 ITR primer_bind complement(3275..3292) /label=M13 Forward /note="In lacZ gene. Also called M13-F20 or M13 (-21) Forward" primer_bind complement(3275..3291) /label=M13 fwd /note="common sequencing primer, one of multiple similar variants" primer_bind complement(3284..3306) /label=M13/pUC Forward /note="In lacZ gene" rep_origin 3433..3888 /direction=RIGHT /label=f1 ori /note="f1 bacteriophage origin of replication; arrow indicates direction of (+) strand synthesis" primer_bind complement(3520..3539) /label=F1ori-R /note="F1 origin, reverse primer" primer_bind 3730..3751 /label=F1ori-F /note="F1 origin, forward primer" promoter 3914..4018 /gene="bla" /label=AmpR promoter CDS 4019..4879 /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(4237..4256) /label=Amp-R /note="Ampicillin resistance gene, reverse primer" rep_origin 5050..5638 /direction=RIGHT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" primer_bind 5539..5558 /label=pBR322ori-F /note="pBR322 origin, forward primer" primer_bind 5792..5809 /label=L4440 /note="L4440 vector, forward primer" ORIGIN 1 agcgcccaat acgcaaaccg cctctccccg cgcgttggcc gattcattaa tgcagctggc 61 acgacaggtt tcccgactgg aaagcgggca gtgagcgcaa cgcaattaat gtgagttagc 121 tcactcatta ggcaccccag gctttacact ttatgcttcc ggctcgtatg ttgtgtggaa 181 ttgtgagcgg ataacaattt cacacaggaa acagctatga ccatgattac gccagattta 241 attaaggcct taattaggct gcgcgctcgc tcgctcactg aggccgcccg ggcaaagccc 301 gggcgtcggg cgacctttgg tcgcccggcc tcagtgagcg agcgagcgcg cagagaggga 361 gtggccaact ccatcactag gggttccttg tagttaatga ttaacccgcc atgctactta 421 tctacgtagc catgctctag gaagatctaa catatcctgg tgtggagtag gggacgctgc 481 tctgacagag gctcgggggc ctgagctggc tctgtgagct ggggaggagg cagacagcca 541 ggccttgtct gcaagcagac ctggcagcat tgggctggcc gccccccagg gcctcctctt 601 catgcccagt gaatgactca ccttggcaca gacacaatgt tcggggtggg cacagtgcct 661 gcttcccgcc gcaccccagc ccccctcaaa tgccttccga gaagcccatt gagcaggggg 721 cttgcattgc accccagcct gacagcctgg catcttggga taaaagcagc acagccccct 781 aggggctgcc cttgctgtgt ggcgccaccg gcggtggaga acaaggctct attcagcctg 841 tgcccaggaa aggggatcag gggatgccca ggcatggaca gtgggtggca gggggggaga 901 ggagggctgt ctgcttccca gaagtccaag gacacaaatg ggtgagggga gagctctccc 961 catagctggg ctgcggccca accccacccc ctcaggctat gccagggggt gttgccaggg 1021 gcacccgggc atcgccagtc tagcccactc cttcataaag ccctcgcatc ccaggagcga 1081 gcagagccag agcaggttgg agaggagacg catcacctcc gctgctcgca agctttattg 1141 cggtagttta tcacagttaa attgctaacg cagtcagtgc ttctgacaca acagtctcga 1201 acttaagctg cagaagttgg tcgtgaggca ctgggcaggt aagtatcaag gttacaagac 1261 aggtttaagg agaccaatag aaactgggct tgtcgagaca gagaagactc ttgcgtttct 1321 gataggcacc tattggtctt actgacatcc actttgcctt tctctccaca ggtgtccact 1381 cccagttcaa ttacagctct taaggctaga gtacttaata cgactcacta taggctagcg 1441 ccaccatggg ttctcatcat catcatcatc atggtatggc tagcatgact ggtggacagc 1501 aaatgggtcg ggatctgtac gacgatgacg ataaggatct cgctagcgcc accatggtcg 1561 actcatcacg tcgtaagtgg aataagacag gtcacgcagt cagagctata ggtcggctga 1621 gctcactcga gaacgtctat atcaaggccg acaagcagaa gaacggcatc aaggcgaact 1681 tcaagatccg ccacaacatc gaggacggcg gcgtgcagct cgcctaccac taccagcaga 1741 acacccccat cggcgacggc cccgtgctgc tgcccgacaa ccactacctg agcgtgcagt 1801 ccaaactttc gaaagacccc aacgagaagc gcgatcacat ggtcctgctg gagttcgtga 1861 ccgccgccgg gatcactctc ggcatggacg agctgtacaa gggcggtacc ggagggagca 1921 tggtgagcaa gggcgaggag ctgttcaccg gggtggtgcc catcctggtc gagctggacg 1981 gcgacgtaaa cggccacaag ttcagcgtgt ccggcgaggg tgagggcgat gccacctacg 2041 gcaagctgac cctgaagttc atctgcacca ccggcaagct gcccgtgccc tggcccaccc 2101 tcgtgaccac cctgacctac ggcgtgcagt gcttcagccg ctaccccgac cacatgaagc 2161 agcacgactt cttcaagtcc gccatgcccg aaggctacat ccaggagcgc accatcttct 2221 tcaaggacga cggcaactac aagacccgcg ccgaggtgaa gttcgagggc gacaccctgg 2281 tgaaccgcat cgagctgaag ggcatcgact tcaaggagga cggcaacatc ctggggcaca 2341 agctggagta caacctgccg gaccaactga ctgaagagca gatcgcagaa tttaaagagg 2401 aattctccct atttgacaag gacggggatg ggacaataac aaccaaggag ctggggacgg 2461 tgatgcggtc tctggggcag aaccccacag aagcagagct gcaggacatg atcaatgaag 2521 tagatgccga cggtgacggc acaatcgact tccctgagtt cctgacaatg atggcaagaa 2581 aaatgaaata cagggacacg gaagaagaaa ttagagaagc gttcggtgtg tttgataagg 2641 atggcaatgg ctacatcagt gcagcagagc ttcgccacgt gatgacaaac cttggagaga 2701 agttaacaga tgaagaggtt gatgaaatga tcagggaagc agacatcgat ggggatggtc 2761 aggtaaacta cgaagagttt gtacaaatga tgacagcgaa gtgatctaga gtcgacccgg 2821 gcggccgctt cgagcagaca tgataagata cattgatgag tttggacaaa ccacaactag 2881 aatgcagtga aaaaaatgct ttatttgtga aatttgtgat gctattgctt tatttgtaac 2941 cattataagc tgcaataaac aagttaacaa caacaattgc attcatttta tgtttcaggt 3001 tcagggggag atgtgggagg ttttttaaag caagtaaaac ctctacaaat gtggtaaaat 3061 cgataaggat cttcctagag catggctacg tagataagta gcatggcggg ttaatcatta 3121 actacaagga acccctagtg atggagttgg ccactccctc tctgcgcgct cgctcgctca 3181 ctgaggccgg gcgaccaaag gtcgcccgac gcccgggctt tgcccgggcg gcctcagtga 3241 gcgagcgagc gcgcagcctt aattaaccta attcactggc cgtcgtttta caacgtcgtg 3301 actgggaaaa ccctggcgtt acccaactta atcgccttgc agcacatccc cctttcgcca 3361 gctggcgtaa tagcgaagag gcccgcaccg atcgcccttc ccaacagttg cgcagcctga 3421 atggcgaatg ggacgcgccc tgtagcggcg cattaagcgc ggcgggtgtg gtggttacgc 3481 gcagcgtgac cgctacactt gccagcgccc tagcgcccgc tcctttcgct ttcttccctt 3541 cctttctcgc cacgttcgcc ggctttcccc gtcaagctct aaatcggggg ctccctttag 3601 ggttccgatt tagtgcttta cggcacctcg accccaaaaa acttgattag ggtgatggtt 3661 cacgtagtgg gccatcgccc tgatagacgg tttttcgccc tttgacgttg gagtccacgt 3721 tctttaatag tggactcttg ttccaaactg gaacaacact caaccctatc tcggtctatt 3781 cttttgattt ataagggatt ttgccgattt cggcctattg gttaaaaaat gagctgattt 3841 aacaaaaatt taacgcgaat tttaacaaaa tattaacgct tacaatttag gtggcacttt 3901 tcggggaaat gtgcgcggaa cccctatttg tttatttttc taaatacatt caaatatgta 3961 tccgctcatg agacaataac cctgataaat gcttcaataa tattgaaaaa ggaagagtat 4021 gagtattcaa catttccgtg tcgcccttat tccctttttt gcggcatttt gccttcctgt 4081 ttttgctcac ccagaaacgc tggtgaaagt aaaagatgct gaagatcagt tgggtgcacg 4141 agtgggttac atcgaactgg atctcaacag cggtaagatc cttgagagtt ttcgccccga 4201 agaacgtttt ccaatgatga gcacttttaa agttctgcta tgtggcgcgg tattatcccg 4261 tattgacgcc gggcaagagc aactcggtcg ccgcatacac tattctcaga atgacttggt 4321 tgagtactca ccagtcacag aaaagcatct tacggatggc atgacagtaa gagaattatg 4381 cagtgctgcc ataaccatga gtgataacac tgcggccaac ttacttctga caacgatcgg 4441 aggaccgaag gagctaaccg cttttttgca caacatgggg gatcatgtaa ctcgccttga 4501 tcgttgggaa ccggagctga atgaagccat accaaacgac gagcgtgaca ccacgatgcc 4561 tgtagcaatg gcaacaacgt tgcgcaaact attaactggc gaactactta ctctagcttc 4621 ccggcaacaa ttaatagact ggatggaggc ggataaagtt gcaggaccac ttctgcgctc 4681 ggcccttccg gctggctggt ttattgctga taaatctgga gccggtgagc gtgggtctcg 4741 cggtatcatt gcagcactgg ggccagatgg taagccctcc cgtatcgtag ttatctacac 4801 gacggggagt caggcaacta tggatgaacg aaatagacag atcgctgaga taggtgcctc 4861 actgattaag cattggtaac tgtcagacca agtttactca tatatacttt agattgattt 4921 aaaacttcat ttttaattta aaaggatcta ggtgaagatc ctttttgata atctcatgac 4981 caaaatccct taacgtgagt tttcgttcca ctgagcgtca gaccccgtag aaaagatcaa 5041 aggatcttct tgagatcctt tttttctgcg cgtaatctgc tgcttgcaaa caaaaaaacc 5101 accgctacca gcggtggttt gtttgccgga tcaagagcta ccaactcttt ttccgaaggt 5161 aactggcttc agcagagcgc agataccaaa tactgttctt ctagtgtagc cgtagttagg 5221 ccaccacttc aagaactctg tagcaccgcc tacatacctc gctctgctaa tcctgttacc 5281 agtggctgct gccagtggcg ataagtcgtg tcttaccggg ttggactcaa gacgatagtt 5341 accggataag gcgcagcggt cgggctgaac ggggggttcg tgcacacagc ccagcttgga 5401 gcgaacgacc tacaccgaac tgagatacct acagcgtgag ctatgagaaa gcgccacgct 5461 tcccgaaggg agaaaggcgg acaggtatcc ggtaagcggc agggtcggaa caggagagcg 5521 cacgagggag cttccagggg gaaacgcctg gtatctttat agtcctgtcg ggtttcgcca 5581 cctctgactt gagcgtcgat ttttgtgatg ctcgtcaggg gggcggagcc tatggaaaaa 5641 cgccagcaac gcggcctttt tacggttcct ggccttttgc tggccttttg ctcacatgtt 5701 ctttcctgcg ttatcccctg attctgtgga taaccgtatt accgcctttg agtgagctga 5761 taccgctcgc cgcagccgaa cgaccgagcg cagcgagtca gtgagcgagg aagcggaag //