LOCUS Exported 6559 bp ds-DNA circular SYN 13-MAY-2021 DEFINITION Independent expression of mOrange and human Kvbeta 1 under human synapsin 1 promoter. ACCESSION . VERSION . KEYWORDS mOrange-P2A-hKvb1 SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 6559) TITLE The potassium channel subunit Kv-beta-1 serves as a major control point for syn-aptic facilitation JOURNAL Unpublished REFERENCE 2 (bases 1 to 6559) AUTHORS . TITLE Direct Submission JOURNAL Exported May 13, 2021 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..6559 /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 promoter 470..917 /label=hSyn promoter /note="human synapsin I promoter; confers neuron-specific expression (Kugler et al., 2003)" regulatory 948..957 /regulatory_class="other" /note="vertebrate consensus sequence for strong initiation of translation (Kozak, 1987)" CDS 954..1661 /codon_start=1 /product="photostable monomeric orange derivative of DsRed fluorescent protein (Shaner et al., 2008)" /label=mOrange2 /note="mammalian codon-optimized" /translation="MVSKGEEYNMAIIKEFMRFKVRMEGSVNGHEFEIEGEGEGRPYEG FQTAKLKVTKGGPLPFAWDILSPHFTYGSKAYVKHPADIPDYFKLSFPEGFKWERVMNY EDGGVVTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASSERMYPEDGALK GKIKMRLKLKDGGHYTSEVKTTYKAKKPVQLPGAYIVDIKLDITSHNEDYTIVEQYERA EGRHSTGGMDELYK" CDS 1671..1727 /codon_start=1 /product="2A peptide from porcine teschovirus-1 polyprotein" /label=P2A /note="Eukaryotic ribosomes fail to insert a peptide bond between the Gly and Pro residues, yielding separate polypeptides." /translation="ATNFSLLKQAGDVEENPGP" misc_feature 2947..3535 /label=WPRE /note="woodchuck hepatitis virus posttranscriptional regulatory element" primer_bind complement(3000..3020) /label=WPRE-R /note="WPRE, reverse primer" CDS complement(3418..3429) /codon_start=1 /product="Factor Xa recognition and cleavage site" /label=Factor Xa site /translation="IEGR" primer_bind complement(3538..3554) /label=KS primer /note="common sequencing primer, one of multiple similar variants" primer_bind complement(3539..3555) /label=pBluescriptKS /note="For pBluescript vector" polyA_signal 3584..3705 /label=SV40 poly(A) signal /note="SV40 polyadenylation signal" primer_bind complement(3595..3614) /label=EBV-rev /note="SV40 polyA terminator, reverse primer" primer_bind 3649..3668 /label=SV40pA-R /note="SV40 polyA, reverse primer" repeat_region 3867..3996 /label=AAV2 ITR primer_bind complement(4015..4032) /label=M13 Forward /note="In lacZ gene. Also called M13-F20 or M13 (-21) Forward" primer_bind complement(4015..4031) /label=M13 fwd /note="common sequencing primer, one of multiple similar variants" primer_bind complement(4024..4046) /label=M13/pUC Forward /note="In lacZ gene" rep_origin 4173..4628 /direction=RIGHT /label=f1 ori /note="f1 bacteriophage origin of replication; arrow indicates direction of (+) strand synthesis" primer_bind complement(4260..4279) /label=F1ori-R /note="F1 origin, reverse primer" primer_bind 4470..4491 /label=F1ori-F /note="F1 origin, forward primer" promoter 4654..4758 /gene="bla" /label=AmpR promoter CDS 4759..5619 /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(4977..4996) /label=Amp-R /note="Ampicillin resistance gene, reverse primer" rep_origin 5790..6378 /direction=RIGHT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" primer_bind 6279..6298 /label=pBR322ori-F /note="pBR322 origin, forward primer" primer_bind 6532..6549 /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 gaagatctct gcagagggcc ctgcgtatga gtgcaagtgg 481 gttttaggac caggatgagg cggggtgggg gtgcctacct gacgaccgac cccgacccac 541 tggacaagca cccaaccccc attccccaaa ttgcgcatcc cctatcagag agggggaggg 601 gaaacaggat gcggcgaggc gcgtgcgcac tgccagcttc agcaccgcgg acagtgcctt 661 cgcccccgcc tggcggcgcg cgccaccgcc gcctcagcac tgaaggcgcg ctgacgtcac 721 tcgccggtcc cccgcaaact ccccttcccg gccaccttgg tcgcgtccgc gccgccgccg 781 gcccagccgg accgcaccac gcgaggcgcg agataggggg gcacgggcgc gaccatctgc 841 gctgcggcgc cggcgactca gcgctgcctc agtctgcggt gggcagcgga ggagtcgtgt 901 cgtgcctgag agcgcagtcg agaattcaag ctgctagcaa ggatcccgcc accatggtga 961 gcaagggcga ggagtataac atggccatca tcaaggagtt catgcgcttc aaggtgcgca 1021 tggagggctc cgtgaacggc cacgagttcg agatcgaggg cgagggcgag ggccgcccct 1081 acgagggctt tcagaccgct aagctgaagg tgaccaaggg tggccccctg cccttcgcct 1141 gggacatcct gtcccctcat ttcacctacg gctccaaggc ctacgtgaag caccccgccg 1201 acatccccga ctacttcaag ctgtccttcc ccgagggctt caagtgggag cgcgtgatga 1261 actacgagga cggcggcgtg gtgaccgtga cccaggactc ctccctgcag gacggcgagt 1321 tcatctacaa ggtgaagctg cgcggcacca acttcccctc cgacggcccc gtgatgcaga 1381 agaagaccat gggctgggag gcctcctccg agcggatgta ccccgaggac ggtgccctga 1441 agggcaagat caagatgagg ctgaagctga aggacggcgg ccactacacc tccgaggtca 1501 agaccaccta caaggccaag aagcccgtgc agctgcccgg cgcctacatc gtcgacatca 1561 agttggacat cacctcccac aacgaggact acaccatcgt ggaacagtac gaacgcgccg 1621 agggccgcca ctccaccggc ggcatggacg agctgtacaa gggaagcgga gctactaact 1681 tcagcctgct gaagcaggct ggagacgtgg aggagaaccc tggacctatg caagtctcca 1741 tagcctgcac agagcacaat ttgaagagtc ggaatggtga ggaccgactt ctgagcaagc 1801 agagctccac cgcccccaat gtggtgaacg cagcccgggc caaattccgc acggtcgcta 1861 tcatcgcgcg cagcctgggg acgttcacgc ctcagcatca catttctctc aaagagtcca 1921 ccgcaaagca gactggcatg aaatatagga atcttggaaa atcaggactc agagtttctt 1981 gcttgggtct tggaacatgg gtgacatttg gaggtcaaat ttcagatgag gttgctgaac 2041 ggctgatgac catcgcctat gaaagtggtg ttaacctctt tgatactgcc gaagtctatg 2101 ctgctggaaa ggctgaagtg attctgggga gcatcatcaa gaagaaaggc tggaggaggt 2161 ccagtctggt cataacaacc aaactctact ggggtggaaa agctgaaaca gaaagagggc 2221 tgtcaagaaa gcatattatt gaaggattga agggctccct ccagaggctg cagctcgagt 2281 atgtggatgt ggtctttgca aatcgaccgg acagtaacac tcccatggaa gaaattgtcc 2341 gagccatgac acatgtgata aaccaaggca tggcgatgta ctggggcacc tcgagatgga 2401 gtgctatgga gatcatggaa gcctattctg tagcaagaca gttcaatatg atcccaccgg 2461 tctgtgaaca agctgagtac catcttttcc agagagagaa agtggaggtc cagctgccag 2521 agctctacca caaaataggt gttggcgcaa tgacatggtc tccacttgcc tgtggaatca 2581 tctcaggaaa atacggaaac ggggtgcctg aaagttccag ggcttcactg aagtgctacc 2641 agtggttgaa agaaagaatt gtaagtgaag aagggagaaa acagcaaaac aagctaaaag 2701 acctttcccc aattgcggag cgtctgggat gcacactacc tcagctagct gttgcgtggt 2761 gcctgagaaa tgaaggtgtg agttctgtgc tcctgggatc atccactcct gaacaactca 2821 ttgaaaacct tggtgccatt caggttctcc caaagatgac atcacatgtg gtaaatgaga 2881 ttgataacat actgcgcaac aagccctaca gcaagaagga ctatagatca taagaagctt 2941 atcgataatc aacctctgga ttacaaaatt tgtgaaagat tgactggtat tcttaactat 3001 gttgctcctt ttacgctatg tggatacgct gctttaatgc ctttgtatca tgctattgct 3061 tcccgtatgg ctttcatttt ctcctccttg tataaatcct ggttgctgtc tctttatgag 3121 gagttgtggc ccgttgtcag gcaacgtggc gtggtgtgca ctgtgtttgc tgacgcaacc 3181 cccactggtt ggggcattgc caccacctgt cagctccttt ccgggacttt cgctttcccc 3241 ctccctattg ccacggcgga actcatcgcc gcctgccttg cccgctgctg gacaggggct 3301 cggctgttgg gcactgacaa ttccgtggtg ttgtcgggga aatcatcgtc ctttccttgg 3361 ctgctcgcct gtgttgccac ctggattctg cgcgggacgt ccttctgcta cgtcccttcg 3421 gccctcaatc cagcggacct tccttcccgc ggcctgctgc cggctctgcg gcctcttccg 3481 cgtcttcgcc ttcgccctca gacgagtcgg atctcccttt gggccgcctc cccgcatcga 3541 taccgtcgac ctcgacccgg gcggccgctt cgagcagaca tgataagata cattgatgag 3601 tttggacaaa ccacaactag aatgcagtga aaaaaatgct ttatttgtga aatttgtgat 3661 gctattgctt tatttgtaac cattataagc tgcaataaac aagttaacaa caacaattgc 3721 attcatttta tgtttcaggt tcagggggag atgtgggagg ttttttaaag caagtaaaac 3781 ctctacaaat gtggtaaaat cgataaggat cttcctagag catggctacg tagataagta 3841 gcatggcggg ttaatcatta actacaagga acccctagtg atggagttgg ccactccctc 3901 tctgcgcgct cgctcgctca ctgaggccgg gcgaccaaag gtcgcccgac gcccgggctt 3961 tgcccgggcg gcctcagtga gcgagcgagc gcgcagcctt aattaaccta attcactggc 4021 cgtcgtttta caacgtcgtg actgggaaaa ccctggcgtt acccaactta atcgccttgc 4081 agcacatccc cctttcgcca gctggcgtaa tagcgaagag gcccgcaccg atcgcccttc 4141 ccaacagttg cgcagcctga atggcgaatg ggacgcgccc tgtagcggcg cattaagcgc 4201 ggcgggtgtg gtggttacgc gcagcgtgac cgctacactt gccagcgccc tagcgcccgc 4261 tcctttcgct ttcttccctt cctttctcgc cacgttcgcc ggctttcccc gtcaagctct 4321 aaatcggggg ctccctttag ggttccgatt tagtgcttta cggcacctcg accccaaaaa 4381 acttgattag ggtgatggtt cacgtagtgg gccatcgccc tgatagacgg tttttcgccc 4441 tttgacgttg gagtccacgt tctttaatag tggactcttg ttccaaactg gaacaacact 4501 caaccctatc tcggtctatt cttttgattt ataagggatt ttgccgattt cggcctattg 4561 gttaaaaaat gagctgattt aacaaaaatt taacgcgaat tttaacaaaa tattaacgct 4621 tacaatttag gtggcacttt tcggggaaat gtgcgcggaa cccctatttg tttatttttc 4681 taaatacatt caaatatgta tccgctcatg agacaataac cctgataaat gcttcaataa 4741 tattgaaaaa ggaagagtat gagtattcaa catttccgtg tcgcccttat tccctttttt 4801 gcggcatttt gccttcctgt ttttgctcac ccagaaacgc tggtgaaagt aaaagatgct 4861 gaagatcagt tgggtgcacg agtgggttac atcgaactgg atctcaacag cggtaagatc 4921 cttgagagtt ttcgccccga agaacgtttt ccaatgatga gcacttttaa agttctgcta 4981 tgtggcgcgg tattatcccg tattgacgcc gggcaagagc aactcggtcg ccgcatacac 5041 tattctcaga atgacttggt tgagtactca ccagtcacag aaaagcatct tacggatggc 5101 atgacagtaa gagaattatg cagtgctgcc ataaccatga gtgataacac tgcggccaac 5161 ttacttctga caacgatcgg aggaccgaag gagctaaccg cttttttgca caacatgggg 5221 gatcatgtaa ctcgccttga tcgttgggaa ccggagctga atgaagccat accaaacgac 5281 gagcgtgaca ccacgatgcc tgtagcaatg gcaacaacgt tgcgcaaact attaactggc 5341 gaactactta ctctagcttc ccggcaacaa ttaatagact ggatggaggc ggataaagtt 5401 gcaggaccac ttctgcgctc ggcccttccg gctggctggt ttattgctga taaatctgga 5461 gccggtgagc gtgggtctcg cggtatcatt gcagcactgg ggccagatgg taagccctcc 5521 cgtatcgtag ttatctacac gacggggagt caggcaacta tggatgaacg aaatagacag 5581 atcgctgaga taggtgcctc actgattaag cattggtaac tgtcagacca agtttactca 5641 tatatacttt agattgattt aaaacttcat ttttaattta aaaggatcta ggtgaagatc 5701 ctttttgata atctcatgac caaaatccct taacgtgagt tttcgttcca ctgagcgtca 5761 gaccccgtag aaaagatcaa aggatcttct tgagatcctt tttttctgcg cgtaatctgc 5821 tgcttgcaaa caaaaaaacc accgctacca gcggtggttt gtttgccgga tcaagagcta 5881 ccaactcttt ttccgaaggt aactggcttc agcagagcgc agataccaaa tactgttctt 5941 ctagtgtagc cgtagttagg ccaccacttc aagaactctg tagcaccgcc tacatacctc 6001 gctctgctaa tcctgttacc agtggctgct gccagtggcg ataagtcgtg tcttaccggg 6061 ttggactcaa gacgatagtt accggataag gcgcagcggt cgggctgaac ggggggttcg 6121 tgcacacagc ccagcttgga gcgaacgacc tacaccgaac tgagatacct acagcgtgag 6181 ctatgagaaa gcgccacgct tcccgaaggg agaaaggcgg acaggtatcc ggtaagcggc 6241 agggtcggaa caggagagcg cacgagggag cttccagggg gaaacgcctg gtatctttat 6301 agtcctgtcg ggtttcgcca cctctgactt gagcgtcgat ttttgtgatg ctcgtcaggg 6361 gggcggagcc tatggaaaaa cgccagcaac gcggcctttt tacggttcct ggccttttgc 6421 tggccttttg ctcacatgtt ctttcctgcg ttatcccctg attctgtgga taaccgtatt 6481 accgcctttg agtgagctga taccgctcgc cgcagccgaa cgaccgagcg cagcgagtca 6541 gtgagcgagg aagcggaag //