LOCUS       Exported                6969 bp ds-DNA     circular SYN 26-APR-2022
DEFINITION  synthetic circular DNA
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6969)
  AUTHORS   Felix Horns
  TITLE     Direct Submission
  JOURNAL   Exported Jul 5, 2023 from SnapGene 4.0.8
            http://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..6969
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     enhancer        235..614
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        615..818
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     promoter        863..881
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     misc_feature    895..921
                     /label=MCS
                     /note="multiple cloning site"
     regulatory      923..932
                     /regulatory_class="other"
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
     CDS             929..1639
                     /codon_start=1
                     /product="monomeric derivative of DsRed fluorescent protein
                     (Shaner et al., 2004)"
                     /label=mCherry
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEEDNMAIIKEFMRFKVHMEGSVNGHEFEIEGEGEGRPYEG
                     TQTAKLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNF
                     EDGGVVTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASSERMYPEDGALK
                     GEIKQRLKLKDGGHYDAEVKTTYKAKKPVQLPGAYNVNIKLDITSHNEDYTIVEQYERA
                     EGRHSTGGMDELYK"
     primer_bind     1640..1662
                     /label=ISPCR
                     /note="ISPCR primer site for amplification using ISPCR 
                     primers"
     primer_bind     1663..1676
                     /label=s5
                     /note="Nextera S5 primer site for amplification using S5xx 
                     index primers"
     misc_feature    1677..1684
                     /label=SgsI
                     /note="Cut site for SgsI"
     misc_feature    1693..1700
                     /label=NotI
                     /note="Cut site for NotI"
     primer_bind     complement(1701..1715)
                     /label=s7
                     /note="Nextera s7 primer site for amplification using N7xx 
                     index primers"
     primer_bind     complement(1716..1738)
                     /label=ISPCR
                     /note="ISPCR primer site for amplification using ISPCR 
                     primers"
     primer_bind     complement(1739..1759)
                     /label=RT
                     /note="RT primer binding site"
     misc_feature    1766..2528
                     /label=PP7 x12
                     /note="PP7 12x stem loop array. Created from Halstead et al
                     Science 2015 (https://www.addgene.org/64542/) by taking 
                     region between SalI/XhoI, then removing 5' and 3' ends that
                     are not participating in RNA structure (as predicted by 
                     Mfold)."
     misc_feature    1776..1781
                     /label=PP7 stem-loop
     misc_feature    1842..1847
                     /label=PP7 stem-loop
     misc_feature    1908..1913
                     /label=PP7 stem-loop
     misc_feature    1974..1979
                     /label=PP7 stem-loop
     misc_feature    2040..2045
                     /label=PP7 stem-loop
     misc_feature    2106..2111
                     /label=PP7 stem-loop
     misc_feature    2175..2180
                     /label=PP7 stem-loop
     misc_feature    2241..2246
                     /label=PP7 stem-loop
     misc_feature    2307..2312
                     /label=PP7 stem-loop
     misc_feature    2373..2378
                     /label=PP7 stem-loop
     misc_feature    2439..2444
                     /label=PP7 stem-loop
     misc_feature    2505..2510
                     /label=PP7 stem-loop
     misc_feature    2530..2540
                     /label=MCS
                     /note="multiple cloning site"
     polyA_signal    2566..2790
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      2836..3264
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     promoter        3278..3607
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      3458..3593
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             3674..4468
                     /codon_start=1
                     /gene="aph(3')-II (or nptII)"
                     /product="aminoglycoside phosphotransferase from Tn5"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin)"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     polyA_signal    4642..4763
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4812..4828)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     rep_origin      complement(5214..5802)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     CDS             complement(5973..6833)
                     /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"
     promoter        complement(6834..6938)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 gacggatcgg gagatctccc gatcccctat ggtgcactct cagtacaatc tgctctgatg
       61 ccgcatagtt aagccagtat ctgctccctg cttgtgtgtt ggaggtcgct gagtagtgcg
      121 cgagcaaaat ttaagctaca acaaggcaag gcttgaccga caattgcatg aagaatctgc
      181 ttagggttag gcgttttgcg ctgcttcgcg atgtacgggc cagatatacg cgttgacatt
      241 gattattgac tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata
      301 tggagttccg cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc
      361 cccgcccatt gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc
      421 attgacgtca atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt
      481 atcatatgcc aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt
      541 atgcccagta catgacctta tgggactttc ctacttggca gtacatctac gtattagtca
      601 tcgctattac catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg
      661 actcacgggg atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc
      721 aaaatcaacg ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg
      781 gtaggcgtgt acggtgggag gtctatataa gcagagctct ctggctaact agagaaccca
      841 ctgcttactg gcttatcgaa attaatacga ctcactatag ggagacccaa gctggctagc
      901 gtttaaactt aagcttggta ccgccaccat ggtgagcaag ggcgaggagg ataacatggc
      961 catcatcaag gagttcatgc gcttcaaggt gcacatggag ggctccgtga acggccacga
     1021 gttcgagatc gagggcgagg gcgagggccg cccctacgag ggcacccaga ccgccaagct
     1081 gaaggtgacc aagggtggcc ccctgccctt cgcctgggac atcctgtccc ctcagttcat
     1141 gtacggctcc aaggcctacg tgaagcaccc cgccgacatc cccgactact tgaagctgtc
     1201 cttccccgag ggcttcaagt gggagcgcgt gatgaacttc gaggacggcg gcgtggtgac
     1261 cgtgacccag gactcctccc tgcaggacgg cgagttcatc tacaaggtga agctgcgcgg
     1321 caccaacttc ccctccgacg gccccgtaat gcagaagaag accatgggct gggaggcctc
     1381 ctccgagcgg atgtaccccg aggacggcgc cctgaagggc gagatcaagc agaggctgaa
     1441 gctgaaggac ggcggccact acgacgctga ggtcaagacc acctacaagg ccaagaagcc
     1501 cgtgcagctg cccggcgcct acaacgtcaa catcaagttg gacatcacct cccacaacga
     1561 ggactacacc atcgtggaac agtacgaacg cgccgagggc cgccactcca ccggcggcat
     1621 ggacgagctg tacaagtaga agcagtggta tcaacgcaga gttcgtcggc agcgtcggcg
     1681 cgccatttaa atgcggccgc ccgagcccac gagacactct gcgttgatac cactgcttca
     1741 gagtagttga atgagccacg gatccgcacc agaggatatg gcctcggtgc ctcagcatct
     1801 tcagatgtgc tgtcactgtc caatcttcaa gccagcagag catatgggct cgctggctcc
     1861 actgtcatta cagtggacct gatcttctcc aatcaaagga gcagaggata tggcctcgct
     1921 ccctccgact acatcaggct ggaagtgaca tccatcattc atcccagcag agcatatggg
     1981 ctcgctggga ccacagcatg cctcgaagtg ctgagagact atcatcaaac cgacagagga
     2041 tatggcctcg tcggctccta ctccctcctg gtggaatact tctccactct tcaacccagc
     2101 agagcatatg ggctcgctgg gtccatcaga gatacaagat ctctcgacag atcctacctg
     2161 caaagcacca gaggatatgg cctcggtgcc tcagcatctt cagatgtgct gtcactgtcc
     2221 aatcttcaag ccagcagagc atatgggctc gctggctcca ctgtcattac agtggacctg
     2281 atcttctcca atcaaaggag cagaggatat ggcctcgctc cctccgacta catcaggctg
     2341 gaagtgacat ccatcattca tcccagcaga gcatatgggc tcgctgggac cacagcatgc
     2401 ctcgaagtgc tgagagacta tcatcaaacc gacagaggat atggcctcgt cggctcctac
     2461 tccctcctgg tggaatactt ctccactctt caacccagca gagcatatgg gctcgctggg
     2521 tccatcagtc tagagggccc gtttaaaccc gctgatcagc ctcgactgtg ccttctagtt
     2581 gccagccatc tgttgtttgc ccctcccccg tgccttcctt gaccctggaa ggtgccactc
     2641 ccactgtcct ttcctaataa aatgaggaaa ttgcatcgca ttgtctgagt aggtgtcatt
     2701 ctattctggg gggtggggtg gggcaggaca gcaaggggga ggattgggaa gacaatagca
     2761 ggcatgctgg ggatgcggtg ggctctatgg cttctgaggc ggaaagaacc agctggggct
     2821 ctagggggta tccccacgcg ccctgtagcg gcgcattaag cgcggcgggt gtggtggtta
     2881 cgcgcagcgt gaccgctaca cttgccagcg ccctagcgcc cgctcctttc gctttcttcc
     2941 cttcctttct cgccacgttc gccggctttc cccgtcaagc tctaaatcgg gggctccctt
     3001 tagggttccg atttagtgct ttacggcacc tcgaccccaa aaaacttgat tagggtgatg
     3061 gttcacgtag tgggccatcg ccctgataga cggtttttcg ccctttgacg ttggagtcca
     3121 cgttctttaa tagtggactc ttgttccaaa ctggaacaac actcaaccct atctcggtct
     3181 attcttttga tttataaggg attttgccga tttcggccta ttggttaaaa aatgagctga
     3241 tttaacaaaa atttaacgcg aattaattct gtggaatgtg tgtcagttag ggtgtggaaa
     3301 gtccccaggc tccccagcag gcagaagtat gcaaagcatg catctcaatt agtcagcaac
     3361 caggtgtgga aagtccccag gctccccagc aggcagaagt atgcaaagca tgcatctcaa
     3421 ttagtcagca accatagtcc cgcccctaac tccgcccatc ccgcccctaa ctccgcccag
     3481 ttccgcccat tctccgcccc atggctgact aatttttttt atttatgcag aggccgaggc
     3541 cgcctctgcc tctgagctat tccagaagta gtgaggaggc ttttttggag gcctaggctt
     3601 ttgcaaaaag ctcccgggag cttgtatatc cattttcgga tctgatcaag agacaggatg
     3661 aggatcgttt cgcatgattg aacaagatgg attgcacgca ggttctccgg ccgcttgggt
     3721 ggagaggcta ttcggctatg actgggcaca acagacaatc ggctgctctg atgccgccgt
     3781 gttccggctg tcagcgcagg ggcgcccggt tctttttgtc aagaccgacc tgtccggtgc
     3841 cctgaatgaa ctgcaggacg aggcagcgcg gctatcgtgg ctggccacga cgggcgttcc
     3901 ttgcgcagct gtgctcgacg ttgtcactga agcgggaagg gactggctgc tattgggcga
     3961 agtgccgggg caggatctcc tgtcatctca ccttgctcct gccgagaaag tatccatcat
     4021 ggctgatgca atgcggcggc tgcatacgct tgatccggct acctgcccat tcgaccacca
     4081 agcgaaacat cgcatcgagc gagcacgtac tcggatggaa gccggtcttg tcgatcagga
     4141 tgatctggac gaagagcatc aggggctcgc gccagccgaa ctgttcgcca ggctcaaggc
     4201 gcgcatgccc gacggcgagg atctcgtcgt gacccatggc gatgcctgct tgccgaatat
     4261 catggtggaa aatggccgct tttctggatt catcgactgt ggccggctgg gtgtggcgga
     4321 ccgctatcag gacatagcgt tggctacccg tgatattgct gaagagcttg gcggcgaatg
     4381 ggctgaccgc ttcctcgtgc tttacggtat cgccgctccc gattcgcagc gcatcgcctt
     4441 ctatcgcctt cttgacgagt tcttctgagc gggactctgg ggttcgaaat gaccgaccaa
     4501 gcgacgccca acctgccatc acgagatttc gattccaccg ccgccttcta tgaaaggttg
     4561 ggcttcggaa tcgttttccg ggacgccggc tggatgatcc tccagcgcgg ggatctcatg
     4621 ctggagttct tcgcccaccc caacttgttt attgcagctt ataatggtta caaataaagc
     4681 aatagcatca caaatttcac aaataaagca tttttttcac tgcattctag ttgtggtttg
     4741 tccaaactca tcaatgtatc ttatcatgtc tgtataccgt cgacctctag ctagagcttg
     4801 gcgtaatcat ggtcatagct gtttcctgtg tgaaattgtt atccgctcac aattccacac
     4861 aacatacgag ccggaagcat aaagtgtaaa gcctggggtg cctaatgagt gagctaactc
     4921 acattaattg cgttgcgctc actgcccgct ttccagtcgg gaaacctgtc gtgccagctg
     4981 cattaatgaa tcggccaacg cgcggggaga ggcggtttgc gtattgggcg ctcttccgct
     5041 tcctcgctca ctgactcgct gcgctcggtc gttcggctgc ggcgagcggt atcagctcac
     5101 tcaaaggcgg taatacggtt atccacagaa tcaggggata acgcaggaaa gaacatgtga
     5161 gcaaaaggcc agcaaaaggc caggaaccgt aaaaaggccg cgttgctggc gtttttccat
     5221 aggctccgcc cccctgacga gcatcacaaa aatcgacgct caagtcagag gtggcgaaac
     5281 ccgacaggac tataaagata ccaggcgttt ccccctggaa gctccctcgt gcgctctcct
     5341 gttccgaccc tgccgcttac cggatacctg tccgcctttc tcccttcggg aagcgtggcg
     5401 ctttctcata gctcacgctg taggtatctc agttcggtgt aggtcgttcg ctccaagctg
     5461 ggctgtgtgc acgaaccccc cgttcagccc gaccgctgcg ccttatccgg taactatcgt
     5521 cttgagtcca acccggtaag acacgactta tcgccactgg cagcagccac tggtaacagg
     5581 attagcagag cgaggtatgt aggcggtgct acagagttct tgaagtggtg gcctaactac
     5641 ggctacacta gaagaacagt atttggtatc tgcgctctgc tgaagccagt taccttcgga
     5701 aaaagagttg gtagctcttg atccggcaaa caaaccaccg ctggtagcgg tggttttttt
     5761 gtttgcaagc agcagattac gcgcagaaaa aaaggatctc aagaagatcc tttgatcttt
     5821 tctacggggt ctgacgctca gtggaacgaa aactcacgtt aagggatttt ggtcatgaga
     5881 ttatcaaaaa ggatcttcac ctagatcctt ttaaattaaa aatgaagttt taaatcaatc
     5941 taaagtatat atgagtaaac ttggtctgac agttaccaat gcttaatcag tgaggcacct
     6001 atctcagcga tctgtctatt tcgttcatcc atagttgcct gactccccgt cgtgtagata
     6061 actacgatac gggagggctt accatctggc cccagtgctg caatgatacc gcgagaccca
     6121 cgctcaccgg ctccagattt atcagcaata aaccagccag ccggaagggc cgagcgcaga
     6181 agtggtcctg caactttatc cgcctccatc cagtctatta attgttgccg ggaagctaga
     6241 gtaagtagtt cgccagttaa tagtttgcgc aacgttgttg ccattgctac aggcatcgtg
     6301 gtgtcacgct cgtcgtttgg tatggcttca ttcagctccg gttcccaacg atcaaggcga
     6361 gttacatgat cccccatgtt gtgcaaaaaa gcggttagct ccttcggtcc tccgatcgtt
     6421 gtcagaagta agttggccgc agtgttatca ctcatggtta tggcagcact gcataattct
     6481 cttactgtca tgccatccgt aagatgcttt tctgtgactg gtgagtactc aaccaagtca
     6541 ttctgagaat agtgtatgcg gcgaccgagt tgctcttgcc cggcgtcaat acgggataat
     6601 accgcgccac atagcagaac tttaaaagtg ctcatcattg gaaaacgttc ttcggggcga
     6661 aaactctcaa ggatcttacc gctgttgaga tccagttcga tgtaacccac tcgtgcaccc
     6721 aactgatctt cagcatcttt tactttcacc agcgtttctg ggtgagcaaa aacaggaagg
     6781 caaaatgccg caaaaaaggg aataagggcg acacggaaat gttgaatact catactcttc
     6841 ctttttcaat attattgaag catttatcag ggttattgtc tcatgagcgg atacatattt
     6901 gaatgtattt agaaaaataa acaaataggg gttccgcgca catttccccg aaaagtgcca
     6961 cctgacgtc
//
