LOCUS Exported 6289 bp ds-DNA circular SYN 12-MAY-2021 DEFINITION mitochondrial calcium sensor. ACCESSION . VERSION . KEYWORDS pCAG mito-GCaMP5G SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 6289) AUTHORS Kwon SK, Sando R 3rd, Lewis TL, Hirabayashi Y, Maximov A, Polleux F TITLE LKB1 Regulates Mitochondria-Dependent Presynaptic Calcium Clearance and Neurotransmitter Release Properties at Excitatory Synapses along Cortical Axons. JOURNAL PLoS Biol. 2016 Jul 18;14(7):e1002516. doi: 10.1371/journal.pbio.1002516. eCollection 2016 Jul. PUBMED 27429220 REFERENCE 2 (bases 1 to 6289) AUTHORS . TITLE Direct Submission JOURNAL Exported May 12, 2021 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..6289 /organism="synthetic DNA construct" /mol_type="other DNA" rep_origin complement(42..629) /direction=LEFT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" primer_bind complement(122..141) /label=pBR322ori-F /note="pBR322 origin, forward primer" CDS complement(800..1660) /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 1423..1442 /label=Amp-R /note="Ampicillin resistance gene, reverse primer" promoter complement(1661..1765) /gene="bla" /label=AmpR promoter enhancer 1796..2175 /label=CMV enhancer /note="human cytomegalovirus immediate early enhancer" promoter 2177..2454 /label=chicken beta-actin promoter intron 2455..3467 /label=chimeric intron /note="chimera between introns from chicken beta-actin and rabbit beta-globin" primer_bind 3391..3413 /label=pCAGGS-5 /note="Chimeric intron in CAG promoter, forward primer" primer_bind 3475..3494 /label=pCAG-F /note="Rabbit beta-globin intron, for pCAG plasmids, forward primer" CDS 3528..3602 /codon_start=1 /product="mitochondrial presequence of human cytochrome c oxidase subunit VIII (Rizutto et al., 1989)" /label=COX8 presequence /translation="MSVLTPLLLRGLTGSARRLPVPRAK" CDS 3639..3656 /codon_start=1 /product="6xHis affinity tag" /label=6xHis /translation="HHHHHH" CDS 3660..3692 /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 3696..3719 /codon_start=1 /product="Xpress(TM) epitope tag, including an enterokinase recognition and cleavage site" /label=Xpress(TM) tag /translation="DLYDDDDK" regulatory 3726..3735 /regulatory_class="other" /note="vertebrate consensus sequence for strong initiation of translation (Kozak, 1987)" CDS 3747..3803 /codon_start=1 /product="calmodulin-binding peptide from avian smooth muscle myosin light chain kinase" /label=calmodulin-binding peptide /translation="RRKWNKTGHAVRAIGRLSS" primer_bind 4016..4037 /label=EGFP-C /note="EGFP, forward primer" primer_bind complement(4143..4164) /label=EGFP-N /note="EGFP, reverse primer" primer_bind complement(4404..4423) /label=EXFP-R /note="For distinguishing EGFP variants, reverse primer" CDS 4539..4982 /codon_start=1 /product="calcium-binding protein" /label=calmodulin /translation="DQLTEEQIAEFKEAFSLFDKDGDGTITTKELGTVMRSLGQNPTEA ELQDMINEVDADGDGTIDFPEFLTMMARKMKYTDSEEEIREAFRVFDKDGNGYISAAEL RHVMTNLGEKLTDEEVDEMIREADIDGDGQVNYEEFVQMMTAK" primer_bind complement(5073..5092) /label=Bglob-pA-R /note="Rabbit beta-globin polyA region, reverse primer" polyA_signal 5138..5193 /label=-beta-globin poly(A) signal /note="rabbit beta-globin polyadenylation signal (Gil and Proudfoot, 1987)" primer_bind complement(5192..5211) /label=rbglobpA-R /note="Rabbit beta-globin polyA, reverse primer. Also called rb-glob-pA-term-R" primer_bind complement(5584..5600) /label=M13 rev /note="common sequencing primer, one of multiple similar variants" primer_bind complement(5584..5600) /label=M13 Reverse /note="In lacZ gene. Also called M13-rev" primer_bind complement(5597..5619) /label=M13/pUC Reverse /note="In lacZ gene" protein_bind 5608..5624 /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(5632..5662) /label=lac promoter /note="promoter for the E. coli lac operon" protein_bind 5677..5698 /label=CAP binding site /bound_moiety="E. coli catabolite activator protein" /note="CAP binding activates transcription in the presence of cAMP." promoter 5757..5952 /label=SV40 promoter /note="SV40 early promoter" rep_origin 5803..5938 /label=SV40 ori /note="SV40 origin of replication" primer_bind 5865..5884 /label=SV40pro-F /note="SV40 promoter/origin, forward primer" polyA_signal 5958..6092 /label=SV40 poly(A) signal /note="SV40 polyadenylation signal" primer_bind complement(5995..6014) /label=SV40pA-R /note="SV40 polyA, reverse primer" primer_bind 6049..6068 /label=EBV-rev /note="SV40 polyA terminator, reverse primer" primer_bind complement(6160..6177) /label=L4440 /note="L4440 vector, forward primer" ORIGIN 1 caaaaggcca ggaaccgtaa aaaggccgcg ttgctggcgt ttttccatag gctccgcccc 61 cctgacgagc atcacaaaaa tcgacgctca agtcagaggt ggcgaaaccc gacaggacta 121 taaagatacc aggcgtttcc ccctggaagc tccctcgtgc gctctcctgt tccgaccctg 181 ccgcttaccg gatacctgtc cgcctttctc ccttcgggaa gcgtggcgct ttctcatagc 241 tcacgctgta ggtatctcag ttcggtgtag gtcgttcgct ccaagctggg ctgtgtgcac 301 gaaccccccg ttcagcccga ccgctgcgcc ttatccggta actatcgtct tgagtccaac 361 ccggtaagac acgacttatc gccactggca gcagccactg gtaacaggat tagcagagcg 421 aggtatgtag gcggtgctac agagttcttg aagtggtggc ctaactacgg ctacactaga 481 agacagtatt tggtatctgc gctctgctga agccagttac cttcggaaaa agagttggta 541 gctcttgatc cggcaaacaa accaccgctg gtagcggtgg tttttttgtt tgcaagcagc 601 agattacgcg cagaaaaaaa ggatctcaag aagatccttt gatcttttct acggggtctg 661 acgctcagtg gaacgaaaac tcacgttaag ggattttggt catgagatta tcaaaaagga 721 tcttcaccta gatcctttta aattaaaaat gaagttttaa atcaatctaa agtatatatg 781 agtaaacttg gtctgacagt taccaatgct taatcagtga ggcacctatc tcagcgatct 841 gtctatttcg ttcatccata gttgcctgac tccccgtcgt gtagataact acgatacggg 901 agggcttacc atctggcccc agtgctgcaa tgataccgcg agacccacgc tcaccggctc 961 cagatttatc agcaataaac cagccagccg gaagggccga gcgcagaagt ggtcctgcaa 1021 ctttatccgc ctccatccag tctattaatt gttgccggga agctagagta agtagttcgc 1081 cagttaatag tttgcgcaac gttgttgcca ttgctacagg catcgtggtg tcacgctcgt 1141 cgtttggtat ggcttcattc agctccggtt cccaacgatc aaggcgagtt acatgatccc 1201 ccatgttgtg caaaaaagcg gttagctcct tcggtcctcc gatcgttgtc agaagtaagt 1261 tggccgcagt gttatcactc atggttatgg cagcactgca taattctctt actgtcatgc 1321 catccgtaag atgcttttct gtgactggtg agtactcaac caagtcattc tgagaatagt 1381 gtatgcggcg accgagttgc tcttgcccgg cgtcaatacg ggataatacc gcgccacata 1441 gcagaacttt aaaagtgctc atcattggaa aacgttcttc ggggcgaaaa ctctcaagga 1501 tcttaccgct gttgagatcc agttcgatgt aacccactcg tgcacccaac tgatcttcag 1561 catcttttac tttcaccagc gtttctgggt gagcaaaaac aggaaggcaa aatgccgcaa 1621 aaaagggaat aagggcgaca cggaaatgtt gaatactcat actcttcctt tttcaatatt 1681 attgaagcat ttatcagggt tattgtctca tgagcggata catatttgaa tgtatttaga 1741 aaaataaaca aataggggtt ccgcgcacat ttccccgaaa agtgccacct gggtcgacat 1801 tgattattga ctagttatta atagtaatca attacggggt cattagttca tagcccatat 1861 atggagttcc gcgttacata acttacggta aatggcccgc ctggctgacc gcccaacgac 1921 ccccgcccat tgacgtcaat aatgacgtat gttcccatag taacgccaat agggactttc 1981 cattgacgtc aatgggtgga gtatttacgg taaactgccc acttggcagt acatcaagtg 2041 tatcatatgc caagtacgcc ccctattgac gtcaatgacg gtaaatggcc cgcctggcat 2101 tatgcccagt acatgacctt atgggacttt cctacttggc agtacatcta cgtattagtc 2161 atcgctatta ccatggtcga ggtgagcccc acgttctgct tcactctccc catctccccc 2221 ccctccccac ccccaatttt gtatttattt attttttaat tattttgtgc agcgatgggg 2281 gcgggggggg ggggggggcg cgcgccaggc ggggcggggc ggggcgaggg gcggggcggg 2341 gcgaggcgga gaggtgcggc ggcagccaat cagagcggcg cgctccgaaa gtttcctttt 2401 atggcgaggc ggcggcggcg gcggccctat aaaaagcgaa gcgcgcggcg ggcgggagtc 2461 gctgcgttgc cttcgccccg tgccccgctc cgcgccgcct cgcgccgccc gccccggctc 2521 tgactgaccg cgttactccc acaggtgagc gggcgggacg gcccttctcc tccgggctgt 2581 aattagcgct tggtttaatg acggctcgtt tcttttctgt ggctgcgtga aagccttaaa 2641 gggctccggg agggcccttt gtgcgggggg gagcggctcg gggggtgcgt gcgtgtgtgt 2701 gtgcgtgggg agcgccgcgt gcggcccgcg ctgcccggcg gctgtgagcg ctgcgggcgc 2761 ggcgcggggc tttgtgcgct ccgcgtgtgc gcgaggggag cgcggccggg ggcggtgccc 2821 cgcggtgcgg gggggctgcg aggggaacaa aggctgcgtg cggggtgtgt gcgtgggggg 2881 gtgagcaggg ggtgtgggcg cggcggtcgg gctgtaaccc ccccctgcac ccccctcccc 2941 gagttgctga gcacggcccg gcttcgggtg cggggctccg tacggggcgt ggcgcggggc 3001 tcgccgtgcc gggcgggggg tggcggcagg tgggggtgcc gggcggggcg gggccgcctc 3061 gggccgggga gggctcgggg gaggggcgcg gcggcccccg gagcgccggc ggctgtcgag 3121 gcgcggcgag ccgcagccat tgccttttat ggtaatcgtg cgagagggcg cagggacttc 3181 ctttgtccca aatctgtgcg gagccgaaat ctgggaggcg ccgccgcacc ccctctagcg 3241 ggcgcggggc gaagcggtgc ggcgccggca ggaaggaaat gggcggggag ggccttcgtg 3301 cgtcgccgcg ccgccgtccc cttctccctc tccagcctcg gggctgtccg cggggggacg 3361 gctgccttcg ggggggacgg ggcagggcgg ggttcggctt ctggcgtgtg accggcggct 3421 ctagagcctc tgctaaccat gttcatgcct tcttcttttt cctacagctc ctgggcaacg 3481 tgctggttat tgtgctgtct catcattttg gcaaagaatt gctcgagatg tccgtcctga 3541 cgccgctgct gctgcggggc ttgacaggct cggcccggcg gctcccagtg ccgcgcgcca 3601 agatccattc gttgggggat ccaccggtca tgggttctca tcatcatcat catcatggta 3661 tggctagcat gactggtgga cagcaaatgg gtcgggatct gtacgacgat gacgataagg 3721 atctcgccac catggtcgac tcatcacgtc gtaagtggaa taagacaggt cacgcagtca 3781 gagctatagg tcggctgagc tcactcgaga acgtctatat caaggccgac aagcagaaga 3841 acggcatcaa ggcgaacttc aagatccgcc acaacatcga ggacggcggc gtgcagctcg 3901 cctaccacta ccagcagaac acccccatcg gcgacggccc cgtgctgctg cccgacaacc 3961 actacctgag cgtgcagtcc aaactttcga aagaccccaa cgagaagcgc gatcacatgg 4021 tcctgctgga gttcgtgacc gccgccggga tcactctcgg catggacgag ctgtacaagg 4081 gcggtaccgg agggagcatg gtgagcaagg gcgaggagct gttcaccggg gtggtgccca 4141 tcctggtcga gctggacggc gacgtaaacg gccacaagtt cagcgtgtcc ggcgagggtg 4201 agggcgatgc cacctacggc aagctgaccc tgaagttcat ctgcaccacc ggcaagctgc 4261 ccgtgccctg gcccaccctc gtgaccaccc tgacctacgg cgtgcagtgc ttcagccgct 4321 accccgacca catgaagcag cacgacttct tcaagtccgc catgcccgaa ggctacatcc 4381 aggagcgcac catcttcttc aaggacgacg gcaactacaa gacccgcgcc gaggtgaagt 4441 tcgagggcga caccctggtg aaccgcatcg agctgaaggg catcgacttc aaggaggacg 4501 gcaacatcct ggggcacaag ctggagtaca acctgccgga ccaactgact gaagagcaga 4561 tcgcagaatt taaagaggct ttctccctat ttgacaagga cggggatggg acaataacaa 4621 ccaaggagct ggggacggtg atgcggtctc tggggcagaa ccccacagaa gcagagctgc 4681 aggacatgat caatgaagta gatgccgacg gtgacggcac aatcgacttc cctgagttcc 4741 tgacaatgat ggcaagaaaa atgaaataca cagacagtga agaagaaatt agagaagcgt 4801 tccgtgtgtt tgataaggat ggcaatggct acatcagtgc agcagagctt cgccacgtga 4861 tgacaaacct tggagagaag ttaacagatg aagaggttga tgaaatgatc agggaagcag 4921 acatcgatgg ggatggtcag gtaaactacg aagagtttgt acaaatgatg acagcgaagt 4981 gagcggccgc aattcactcc tcaggtgcag gctgcctatc agaaggtggt ggctggtgtg 5041 gccaatgccc tggctcacaa ataccactga gatctttttc cctctgccaa aaattatggg 5101 gacatcatga agccccttga gcatctgact tctggctaat aaaggaaatt tattttcatt 5161 gcaatagtgt gttggaattt tttgtgtcta tcactcggaa ggacatatgg gagggcaaat 5221 catttaaaac atcagaatga gtatttggtt tagagtttgg caacatatgc ccatatgctg 5281 gctgccatga acaaaggttg gctataaaga ggtcatcagt atatgaaaca gccccctgct 5341 gtccattcct tattccatag aaaagccttg acttgaggtt agattttttt tatattttgt 5401 tttgtgttat ttttttcttt aacatcccta aaattttcct tacatgtttt actagccaga 5461 tttttcctcc tctcctgact actcccagtc atagctgtcc ctcttctctt atggagatcc 5521 ctcgacctgc accgtcgacc agctggtcga cggtgcaccg tcgaccagct tggcgtaatc 5581 atggtcatag ctgtttcctg tgtgaaattg ttatccgctc acaattccac acaacatacg 5641 agccggaagc ataaagtgta aagcctgggg tgcctaatga gtgagctaac tcacattaat 5701 tgcgttgcgc tcactgcccg ctttccagtc gggaaacctg tcgtgccagc ggatccgcat 5761 ctcaattagt cagcaaccat agtcccgccc ctaactccgc ccatcccgcc cctaactccg 5821 cccagttccg cccattctcc gccccatggc tgactaattt tttttattta tgcagaggcc 5881 gaggccgcct cggcctctga gctattccag aagtagtgag gaggcttttt tggaggccta 5941 ggcttttgca aaaagctaac ttgtttattg cagcttataa tggttacaaa taaagcaata 6001 gcatcacaaa tttcacaaat aaagcatttt tttcactgca ttctagttgt ggtttgtcca 6061 aactcatcaa tgtatcttat catgtctgga tccgctgcat taatgaatcg gccaacgcgc 6121 ggggagaggc ggtttgcgta ttgggcgctc ttccgcttcc tcgctcactg actcgctgcg 6181 ctcggtcgtt cggctgcggc gagcggtatc agctcactca aaggcggtaa tacggttatc 6241 cacagaatca ggggataacg caggaaagaa catgtgagca aaaggccag //