LOCUS Exported 6224 bp ds-DNA circular SYN 28-APR-2022 DEFINITION . ACCESSION . VERSION . KEYWORDS . SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 6224) AUTHORS . TITLE Direct Submission JOURNAL Exported Thursday, Apr 28, 2022 from SnapGene 4.3.11 https://www.snapgene.com FEATURES Location/Qualifiers source 1..6224 /organism="synthetic DNA construct" /mol_type="other DNA" source 664..665 /organism="synthetic DNA construct" /plasmid="NEBuilder_Assembly" /note="color: #ffffff" source 664..665 /organism="synthetic DNA construct /mol_type=""other DNA /note=""pCAG" /note="color: #ffffff" source 672..2320 /organism="synthetic DNA construct" /plasmid="NEBuilder_Assembly" /note="color: #ffffff" source 672..1401 /organism="synthetic DNA construct /mol_type=""other DNA /note=""pCAG" /note="color: #ffffff" source 1402..2026 /organism="synthetic DNA construct /mol_type=""other DNA /note=""IRES" /note="color: #ffffff" source 2027..2320 /organism="synthetic DNA construct /mol_type=""other DNA /note=""mSnca" /note="color: #ffffff" misc_feature 1..141 /label=L-ITR /note="color: #a6acb3" primer_bind complement(23..34) /label=L4440 /note="L4440 vector, forward primer" /note="color: black; sequence: AGCGAGTCAGTGAGCGAG" primer_bind 84..95 /label=L4440 /note="L4440 vector, forward primer" /note="color: black; sequence: AGCGAGTCAGTGAGCGAG" primer_bind 150..173 /label=MluI-Syn-F /note="color: black; sequence: ATTACGCGTtaactgcagagggccctg" promoter 156..626 /label=Syn /note="color: #ffffff" primer_bind 509..527 /label=Syn-Seq /note="color: black; sequence: acgcgaggcgcgagatagg" regulatory 673..682 /regulatory_class="other" /note="vertebrate consensus sequence for strong initiation of translation (Kozak, 1987)" /note="color: #a6acb3" CDS 679..1398 /codon_start=1 /product="enhanced GFP" /label=EGFP /note="mammalian codon-optimized" /note="This feature has 3 segments: 1: 679 .. 681 / #05fd14 2: 682 .. 684 / #05fd14 / 1a 3: 685 .. 1398 / #05fd14" /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL EFVTAAGITLGMDELYK" primer_bind complement(1376..1404) /label=GFP-rev /note="color: black; sequence: ctggctttacttgtacagctcgtccatgc" primer_bind 1385..1433 /label=IRES_fwd /note="3'Tm=76.1 3'Ta(annealing temp)=72.0" /note="color: black; sequence: agctgtacaagtaaagcCAGTGTGGTGGTACGTAGGAATTCTCCCTCCC" misc_feature 1426..1992 /label=IRES /note="internal ribosome entry site (IRES) of the encephalomyocarditis virus (EMCV)" /note="color: #a6acb3" primer_bind complement(1515..1534) /label=IRES-R /note="color: black; sequence: cctcacattgccaaaagacg" primer_bind complement(1997..2030) /label=IRES_rev /note="3'Tm=80.3 3'Ta(annealing temp)=72.0" /note="color: black; sequence: tggcgtcgactctagaGGATCCCGGGCCCGCGGT" primer_bind 2007..2044 /label=mSnca_fwd /note="3'Tm=63.8 3'Ta(annealing temp)=61.7" /note="color: black; sequence: cgggatcctctagagtcgacGCCACCATGGATGTGTTC" regulatory 2027..2036 /regulatory_class="other" /note="vertebrate consensus sequence for strong initiation of translation (Kozak, 1987)" /note="color: #a6acb3" CDS 2033..2320 /codon_start=1 /label=mSnca(1-95) /note="This feature has 2 segments: 1: 2033 .. 2317 / #cc99ff 2: 2318 .. 2320 / #cc99ff" /translation="MDVFMKGLSKAKEGVVAAAEKTKQGVAEAAGKTKEGVLYVGSKTK EGVVHGVTTVAEKTKEQVTNVGGAVVTGVTAVAQKTVEGAGNIAAATGFV" misc_feature 2341..2949 /label=WPRE /note="color: #a6acb3" primer_bind complement(2404..2424) /label=WPRE-R /note="WPRE, reverse primer" /note="color: black; sequence: CATAGCGTAAAAGGAGCAACA" primer_bind complement(2564..2583) /label=seq-wpre-R (Sui) /note="sw primer box2 #1392" /note="color: black; sequence: gttgcgtcagcaaacacagt" misc_feature 2953..2955 /label=AfeI /note="color: #ff99cc" polyA_signal 2969..3447 /label=hGH polyA /note="color: #a6acb3" primer_bind complement(2984..3001) /label=bGH-R /note="color: black; sequence: gcactggggaggggtcac" misc_feature 3487..3627 /label=R-ITR /note="color: #a6acb3" primer_bind complement(3533..3544) /label=L4440 /note="L4440 vector, forward primer" /note="color: black; sequence: AGCGAGTCAGTGAGCGAG" primer_bind 3594..3605 /label=L4440 /note="L4440 vector, forward primer" /note="color: black; sequence: AGCGAGTCAGTGAGCGAG" rep_origin 3719..4025 /label=f1 Origin /note="color: #ffff00" primer_bind complement(3789..3808) /label=F1ori-R /note="F1 origin, reverse primer" /note="color: black; sequence: AGGGAAGAAAGCGAAAGGAG" primer_bind 3999..4020 /label=F1ori-F /note="F1 origin, forward primer" /note="color: black; sequence: GTGGACTCTTGTTCCAAACTGG" promoter 4439..4543 /gene="bla" /label=AmpR promoter /note="color: #ffffff; direction: RIGHT" CDS 4544..5404 /codon_start=1 /gene="bla" /product="beta-lactamase" /label=AmpR /note="confers resistance to ampicillin, carbenicillin, and related antibiotics" /note="This feature has 2 segments: 1: 4544 .. 4612 / #ccffcc / signal sequence 2: 4613 .. 5404 / #ccffcc Cleavage site after base 4612" /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS LIKHW" misc_feature 4544..5401 /label=AmpR /note="color: #a6acb3" primer_bind complement(4762..4781) /label=Amp-R /note="Ampicillin resistance gene, reverse primer" /note="color: black; sequence: ATAATACCGCGCCACATAGC" rep_origin 5552..6219 /label=pUC ori /note="color: #ffff00" primer_bind 6064..6083 /label=pBR322ori-F /note="pBR322 origin, forward primer" /note="color: black; sequence: GGGAAACGCCTGGTATCTTT" ORIGIN 1 cctgcaggca gctgcgcgct cgctcgctca ctgaggccgc ccgggcaaag cccgggcgtc 61 gggcgacctt tggtcgcccg gcctcagtga gcgagcgagc gcgcagagag ggagtggcca 121 actccatcac taggggttcc tgcggccgca cgcgttaact gcagagggcc ctgcgtatga 181 gtgcaagtgg gttttaggac caggatgagg cggggtgggg gtgcctacct gacgaccgac 241 cccgacccac tggacaagca cccaaccccc attccccaaa ttgcgcatcc cctatcagag 301 agggggaggg gaaacaggat gcggcgaggc gcgtgcgcac tgccagcttc agcaccgcgg 361 acagtgcctt cgcccccgcc tggcggcgcg cgccaccgcc gcctcagcac tgaaggcgcg 421 ctgacgtcac tcgccggtcc cccgcaaact ccccttcccg gccaccttgg tcgcgtccgc 481 gccgccgccg gcccagccgg accgcaccac gcgaggcgcg agataggggg gcacgggcgc 541 gaccatctgc gctgcggcgc cggcgactca gcgctgcctc agtctgcggt gggcagcgga 601 ggagtcgtgt cgtgcctgag agcgcgtccg gagctattag caatatgaaa cctcttaggc 661 gcgccaccgg tcgccaccat ggtgagcaag ggcgaggagc tgttcaccgg ggtggtgccc 721 atcctggtcg agctggacgg cgacgtaaac ggccacaagt tcagcgtgtc cggcgagggc 781 gagggcgatg ccacctacgg caagctgacc ctgaagttca tctgcaccac cggcaagctg 841 cccgtgccct ggcccaccct cgtgaccacc ctgacctacg gcgtgcagtg cttcagccgc 901 taccccgacc acatgaagca gcacgacttc ttcaagtccg ccatgcccga aggctacgtc 961 caggagcgca ccatcttctt caaggacgac ggcaactaca agacccgcgc cgaggtgaag 1021 ttcgagggcg acaccctggt gaaccgcatc gagctgaagg gcatcgactt caaggaggac 1081 ggcaacatcc tggggcacaa gctggagtac aactacaaca gccacaacgt ctatatcatg 1141 gccgacaagc agaagaacgg catcaaggtg aacttcaaga tccgccacaa catcgaggac 1201 ggcagcgtgc agctcgccga ccactaccag cagaacaccc ccatcggcga cggccccgtg 1261 ctgctgcccg acaaccacta cctgagcacc cagtccgccc tgagcaaaga ccccaacgag 1321 aagcgcgatc acatggtcct gctggagttc gtgaccgccg ccgggatcac tctcggcatg 1381 gacgagctgt acaagtaaag ccagtgtggt ggtacgtagg aattctccct cccccccccc 1441 taacgttact ggccgaagcc gcttggaata aggccggtgt gcgtttgtct atatgttatt 1501 ttccaccata ttgccgtctt ttggcaatgt gagggcccgg aaacctggcc ctgtcttctt 1561 gacgagcatt cctaggggtc tttcccctct cgccaaagga atgcaaggtc tgttgaatgt 1621 cgtgaaggaa gcagttcctc tggaagcttc ttgaagacaa acaacgtctg tagcgaccct 1681 ttgcaggcag cggaaccccc cacctggcga caggtgcctc tgcggccaaa agccacgtgt 1741 ataagataca cctgcaaagg cggcacaacc ccagtgccac gttgtgagtt ggatagttgt 1801 ggaaagagtc aaatggctct cctcaagcgt attcaacaag gggctgaagg atgcccagaa 1861 ggtaccccat tgtatgggat ctgatctggg gcctcggtgc acatgcttta cgtgtgttta 1921 gtcgaggtta aaaaacgtct aggccccccg aaccacgggg acgtggtttt cctttgaaaa 1981 acacgatgat aatggtaccg cgggcccggg atcctctaga gtcgacgcca ccatggatgt 2041 gttcatgaaa ggactttcaa aggccaagga gggagttgtg gctgctgctg agaaaaccaa 2101 gcagggtgtg gcagaggcag ctggaaagac aaaagaggga gtcctctatg taggttccaa 2161 aactaaggaa ggagtggttc atggagtgac aacagtggct gagaagacca aagagcaagt 2221 gacaaatgtt ggaggagcag tggtgactgg tgtgacagca gtcgctcaga agacagtgga 2281 gggagctggg aatatagctg ctgccactgg ctttgtctaa gctagcgaat tcgatatcaa 2341 gcttatcgat aatcaacctc tggattacaa aatttgtgaa agattgactg gtattcttaa 2401 ctatgttgct ccttttacgc tatgtggata cgctgcttta atgcctttgt atcatgctat 2461 tgcttcccgt atggctttca ttttctcctc cttgtataaa tcctggttgc tgtctcttta 2521 tgaggagttg tggcccgttg tcaggcaacg tggcgtggtg tgcactgtgt ttgctgacgc 2581 aacccccact ggttggggca ttgccaccac ctgtcagctc ctttccggga ctttcgcttt 2641 ccccctccct attgccacgg cggaactcat cgccgcctgc cttgcccgct gctggacagg 2701 ggctcggctg ttgggcactg acaattccgt ggtgttgtcg gggaaatcat cgtcctttcc 2761 ttggctgctc gcctatgttg ccacctggat tctgcgcggg acgtccttct gctacgtccc 2821 ttcggccctc aatccagcgg accttccttc ccgcggcctg ctgccggctc tgcggcctct 2881 tccgcgtctt cgccttcgcc ctcagacgag tcggatctcc ctttgggccg cctccccgca 2941 tcgataccga gcgctgctcg agagatctac gggtggcatc cctgtgaccc ctccccagtg 3001 cctctcctgg ccctggaagt tgccactcca gtgcccacca gccttgtcct aataaaatta 3061 agttgcatca ttttgtctga ctaggtgtcc ttctataata ttatggggtg gaggggggtg 3121 gtatggagca aggggcaagt tgggaagaca acctgtaggg cctgcggggt ctattgggaa 3181 ccaagctgga gtgcagtggc acaatcttgg ctcactgcaa tctccgcctc ctgggttcaa 3241 gcgattctcc tgcctcagcc tcccgagttg ttgggattcc aggcatgcat gaccaggctc 3301 agctaatttt tgtttttttg gtagagacgg ggtttcacca tattggccag gctggtctcc 3361 aactcctaat ctcaggtgat ctacccacct tggcctccca aattgctggg attacaggcg 3421 tgaaccactg ctcccttccc tgtccttctg attttgtagg taaccacgtg cggaccgagc 3481 ggccgcagga acccctagtg atggagttgg ccactccctc tctgcgcgct cgctcgctca 3541 ctgaggccgg gcgaccaaag gtcgcccgac gcccgggctt tgcccgggcg gcctcagtga 3601 gcgagcgagc gcgcagctgc ctgcaggggc gcctgatgcg gtattttctc cttacgcatc 3661 tgtgcggtat ttcacaccgc atacgtcaaa gcaaccatag tacgcgccct gtagcggcgc 3721 attaagcgcg gcgggtgtgg tggttacgcg cagcgtgacc gctacacttg ccagcgccct 3781 agcgcccgct cctttcgctt tcttcccttc ctttctcgcc acgttcgccg gctttccccg 3841 tcaagctcta aatcgggggc tccctttagg gttccgattt agtgctttac ggcacctcga 3901 ccccaaaaaa cttgatttgg gtgatggttc acgtagtggg ccatcgccct gatagacggt 3961 ttttcgccct ttgacgttgg agtccacgtt ctttaatagt ggactcttgt tccaaactgg 4021 aacaacactc aaccctatct cgggctattc ttttgattta taagggattt tgccgatttc 4081 ggcctattgg ttaaaaaatg agctgattta acaaaaattt aacgcgaatt ttaacaaaat 4141 attaacgttt acaattttat ggtgcactct cagtacaatc tgctctgatg ccgcatagtt 4201 aagccagccc cgacacccgc caacacccgc tgacgcgccc tgacgggctt gtctgctccc 4261 ggcatccgct tacagacaag ctgtgaccgt ctccgggagc tgcatgtgtc agaggttttc 4321 accgtcatca ccgaaacgcg cgagacgaaa gggcctcgtg atacgcctat ttttataggt 4381 taatgtcatg ataataatgg tttcttagac gtcaggtggc acttttcggg gaaatgtgcg 4441 cggaacccct atttgtttat ttttctaaat acattcaaat atgtatccgc tcatgagaca 4501 ataaccctga taaatgcttc aataatattg aaaaaggaag agtatgagta ttcaacattt 4561 ccgtgtcgcc cttattccct tttttgcggc attttgcctt cctgtttttg ctcacccaga 4621 aacgctggtg aaagtaaaag atgctgaaga tcagttgggt gcacgagtgg gttacatcga 4681 actggatctc aacagcggta agatccttga gagttttcgc cccgaagaac gttttccaat 4741 gatgagcact tttaaagttc tgctatgtgg cgcggtatta tcccgtattg acgccgggca 4801 agagcaactc ggtcgccgca tacactattc tcagaatgac ttggttgagt actcaccagt 4861 cacagaaaag catcttacgg atggcatgac agtaagagaa ttatgcagtg ctgccataac 4921 catgagtgat aacactgcgg ccaacttact tctgacaacg atcggaggac cgaaggagct 4981 aaccgctttt ttgcacaaca tgggggatca tgtaactcgc cttgatcgtt gggaaccgga 5041 gctgaatgaa gccataccaa acgacgagcg tgacaccacg atgcctgtag caatggcaac 5101 aacgttgcgc aaactattaa ctggcgaact acttactcta gcttcccggc aacaattaat 5161 agactggatg gaggcggata aagttgcagg accacttctg cgctcggccc ttccggctgg 5221 ctggtttatt gctgataaat ctggagccgg tgagcgtggg tctcgcggta tcattgcagc 5281 actggggcca gatggtaagc cctcccgtat cgtagttatc tacacgacgg ggagtcaggc 5341 aactatggat gaacgaaata gacagatcgc tgagataggt gcctcactga ttaagcattg 5401 gtaactgtca gaccaagttt actcatatat actttagatt gatttaaaac ttcattttta 5461 atttaaaagg atctaggtga agatcctttt tgataatctc atgaccaaaa tcccttaacg 5521 tgagttttcg ttccactgag cgtcagaccc cgtagaaaag atcaaaggat cttcttgaga 5581 tccttttttt ctgcgcgtaa tctgctgctt gcaaacaaaa aaaccaccgc taccagcggt 5641 ggtttgtttg ccggatcaag agctaccaac tctttttccg aaggtaactg gcttcagcag 5701 agcgcagata ccaaatactg tccttctagt gtagccgtag ttaggccacc acttcaagaa 5761 ctctgtagca ccgcctacat acctcgctct gctaatcctg ttaccagtgg ctgctgccag 5821 tggcgataag tcgtgtctta ccgggttgga ctcaagacga tagttaccgg ataaggcgca 5881 gcggtcgggc tgaacggggg gttcgtgcac acagcccagc ttggagcgaa cgacctacac 5941 cgaactgaga tacctacagc gtgagctatg agaaagcgcc acgcttcccg aagggagaaa 6001 ggcggacagg tatccggtaa gcggcagggt cggaacagga gagcgcacga gggagcttcc 6061 agggggaaac gcctggtatc tttatagtcc tgtcgggttt cgccacctct gacttgagcg 6121 tcgatttttg tgatgctcgt caggggggcg gagcctatgg aaaaacgcca gcaacgcggc 6181 ctttttacgg ttcctggcct tttgctggcc ttttgctcac atgt //