LOCUS       Exported                4999 bp ds-DNA     circular SYN 12-MAY-2021
DEFINITION  ssAAV genome with MCS for inserting MiniP promoter to drive an 
            emerald GFP (EmGFP) reporter. Contains WPRE.
ACCESSION   .
VERSION     .
KEYWORDS    pEMS2131
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4999)
  AUTHORS   Simpson E, Korecki AJ, Fornes O, McGill TJ, Cueva-Vargas JL, 
            Agostinone J, Farkas RA, Hickmott JW, Lam SL, Mathelier A, Renner 
            LM, Stoddard J, Zhou M, Di Polo A, Neuringer M, Wasserman WW
  TITLE     New MiniPromoter Ple345 (NEFL) Drives Strong and Specific Expression
            in Retinal Ganglion Cells of Mouse and Primate Retina.
  JOURNAL   Hum Gene Ther. 2018 Jul 31. doi: 10.1089/hum.2018.118.
  PUBMED    30062914
REFERENCE   2  (bases 1 to 4999)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported May 12, 2021 from SnapGene Server 1.1.58
            http://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..4999
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     repeat_region   1..130
                     /label=AAV2 ITR
     intron          249..381
                     /label=chimeric intron
                     /note="chimera between introns from human beta-globin and 
                     immunoglobulin heavy chain genes"
     primer_bind     364..383
                     /label=pMT2-F
                     /note="Synthetic intron, forward primer"
     regulatory      402..411
                     /regulatory_class="other"
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
     CDS             408..1127
                     /codon_start=1
                     /product="Emerald GFP"
                     /label=EmGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTFTYGVQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHKVYITADKQKNGIK
                     VNFKTRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(453..474)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(714..733)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1061..1082
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     complement(1193..1213)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(1611..1622)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     polyA_signal    1743..1864
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1754..1773)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     1808..1827
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     repeat_region   2060..2189
                     /label=AAV2 ITR
     primer_bind     complement(2208..2225)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(2208..2224)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2217..2239)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      2366..2821
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2453..2472)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2663..2684
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        2847..2951
                     /gene="bla"
                     /label=AmpR promoter
     CDS             2952..3812
                     /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(3170..3189)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      3983..4571
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4472..4491
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     4725..4742
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    4859..4880
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        4895..4925
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4933..4949
                     /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     4938..4960
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     4957..4973
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     4957..4973
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
ORIGIN
        1 ctgcgcgctc gctcgctcac tgaggccgcc cgggcaaagc ccgggcgtcg ggcgaccttt
       61 ggtcgcccgg cctcagtgag cgagcgagcg cgcagagagg gagtggccaa ctccatcact
      121 aggggttcct tgtagttaat gattaacccg ccatgctact tatctacgta gccatgctct
      181 aggaagatcg gaattcccta ggccggccac gcgtggcgcg ccgaagttgg tcgtgaggca
      241 ctgggcaggt aagtatcaag gttacaagac aggtttaagg agaccaatag aaactgggct
      301 tgtcgagaca gagaagactc ttgcgtttct gataggcacc tattggtctt actgacatcc
      361 actttgcctt tctctccaca ggtgtccact cccaggcggc cgccaccatg gtgagcaagg
      421 gcgaggagct gttcaccggg gtggtgccca tcctggtcga gctggacggc gacgtaaacg
      481 gccacaagtt cagcgtgtcc ggcgagggcg agggcgatgc cacctacggc aagctgaccc
      541 tgaagttcat ctgcaccacc ggcaagctgc ccgtgccctg gcccaccctc gtgaccacct
      601 tcacctacgg cgtgcagtgc ttcgcccgct accccgacca catgaagcag cacgacttct
      661 tcaagtccgc catgcccgaa ggctacgtcc aggagcgcac catcttcttc aaggacgacg
      721 gcaactacaa gacccgcgcc gaggtgaagt tcgagggcga caccctggtg aaccgcatcg
      781 agctgaaggg catcgacttc aaggaggacg gcaacatcct ggggcacaag ctggagtaca
      841 actacaacag ccacaaggtc tatatcaccg ccgacaagca gaagaacggc atcaaggtga
      901 acttcaagac ccgccacaac atcgaggacg gcagcgtgca gctcgccgac cactaccagc
      961 agaacacccc catcggcgac ggccccgtgc tgctgcccga caaccactac ctgagcaccc
     1021 agtccgccct gagcaaagac cccaacgaga agcgcgatca catggtcctg ctggagttcg
     1081 tgaccgccgc cgggatcact ctcggcatgg acgagctgta caagtaagcg gccgcgatcg
     1141 ctcaacctct ggattacaaa atttgtgaaa gattgactgg tattcttaac tatgttgctc
     1201 cttttacgct atgtggatac gctgctttaa tgcctttgta tcatgctatt gcttcccgta
     1261 tggctttcat tttctcctcc ttgtataaat cctggttgct gtctctttat gaggagttgt
     1321 ggcccgttgt caggcaacgt ggcgtggtgt gcactgtgtt tgctgacgca acccccactg
     1381 gttggggcat tgccaccacc tgtcagctcc tttccgggac tttcgctttc cccctcccta
     1441 ttgccacggc ggaactcatc gccgcctgcc ttgcccgctg ctggacaggg gctcggctgt
     1501 tgggcactga caattccgtg gtgttgtcgg ggaaatcatc gtcctttcct tggctgctcg
     1561 cctgtgttgc cacctggatt ctgcgcggga cgtccttctg ctacgtccct tcggccctca
     1621 atccagcgga ccttccttcc cgcggcctgc tgccggctct gcggcctctt ccgcgtcttc
     1681 gccttcgccc tcagacgagt cggatctccc tttgggccgc ctccccgcga tcgcagacat
     1741 gataagatac attgatgagt ttggacaaac cacaactaga atgcagtgaa aaaaatgctt
     1801 tatttgtgaa atttgtgatg ctattgcttt atttgtaacc attataagct gcaataaaca
     1861 agttaacaac aacaattgca ttcattttat gtttcaggtt cagggggaga tgtgggaggt
     1921 tttttaaagc aagtaaaacc tctacaaatg tggtagtcga aattcccgat aaggatcttc
     1981 gtcgacgaat tcccgataag gatcttccta gagcatggct acgtagataa gtagcatggc
     2041 gggttaatca ttaactacaa ggaaccccta gtgatggagt tggccactcc ctctctgcgc
     2101 gctcgctcgc tcactgaggc cgggcgacca aaggtcgccc gacgcccggg ctttgcccgg
     2161 gcggcctcag tgagcgagcg agcgcgcagc cttaattaac ctaattcact ggccgtcgtt
     2221 ttacaacgtc gtgactggga aaaccctggc gttacccaac ttaatcgcct tgcagcacat
     2281 ccccctttcg ccagctggcg taatagcgaa gaggcccgca ccgatcgccc ttcccaacag
     2341 ttgcgcagcc tgaatggcga atgggacgcg ccctgtagcg gcgcattaag cgcggcgggt
     2401 gtggtggtta cgcgcagcgt gaccgctaca cttgccagcg ccctagcgcc cgctcctttc
     2461 gctttcttcc cttcctttct cgccacgttc gccggctttc cccgtcaagc tctaaatcgg
     2521 gggctccctt tagggttccg atttagtgct ttacggcacc tcgaccccaa aaaacttgat
     2581 tagggtgatg gttcacgtag tgggccatcg ccctgataga cggtttttcg ccctttgacg
     2641 ttggagtcca cgttctttaa tagtggactc ttgttccaaa ctggaacaac actcaaccct
     2701 atctcggtct attcttttga tttataaggg attttgccga tttcggccta ttggttaaaa
     2761 aatgagctga tttaacaaaa atttaacgcg aattttaaca aaatattaac gcttacaatt
     2821 taggtggcac ttttcgggga aatgtgcgcg gaacccctat ttgtttattt ttctaaatac
     2881 attcaaatat gtatccgctc atgagacaat aaccctgata aatgcttcaa taatattgaa
     2941 aaaggaagag tatgagtatt caacatttcc gtgtcgccct tattcccttt tttgcggcat
     3001 tttgccttcc tgtttttgct cacccagaaa cgctggtgaa agtaaaagat gctgaagatc
     3061 agttgggtgc acgagtgggt tacatcgaac tggatctcaa cagcggtaag atccttgaga
     3121 gttttcgccc cgaagaacgt tttccaatga tgagcacttt taaagttctg ctatgtggcg
     3181 cggtattatc ccgtattgac gccgggcaag agcaactcgg tcgccgcata cactattctc
     3241 agaatgactt ggttgagtac tcaccagtca cagaaaagca tcttacggat ggcatgacag
     3301 taagagaatt atgcagtgct gccataacca tgagtgataa cactgcggcc aacttacttc
     3361 tgacaacgat cggaggaccg aaggagctaa ccgctttttt gcacaacatg ggggatcatg
     3421 taactcgcct tgatcgttgg gaaccggagc tgaatgaagc cataccaaac gacgagcgtg
     3481 acaccacgat gcctgtagca atggcaacaa cgttgcgcaa actattaact ggcgaactac
     3541 ttactctagc ttcccggcaa caattaatag actggatgga ggcggataaa gttgcaggac
     3601 cacttctgcg ctcggccctt ccggctggct ggtttattgc tgataaatct ggagccggtg
     3661 agcgtgggtc tcgcggtatc attgcagcac tggggccaga tggtaagccc tcccgtatcg
     3721 tagttatcta cacgacgggg agtcaggcaa ctatggatga acgaaataga cagatcgctg
     3781 agataggtgc ctcactgatt aagcattggt aactgtcaga ccaagtttac tcatatatac
     3841 tttagattga tttaaaactt catttttaat ttaaaaggat ctaggtgaag atcctttttg
     3901 ataatctcat gaccaaaatc ccttaacgtg agttttcgtt ccactgagcg tcagaccccg
     3961 tagaaaagat caaaggatct tcttgagatc ctttttttct gcgcgtaatc tgctgcttgc
     4021 aaacaaaaaa accaccgcta ccagcggtgg tttgtttgcc ggatcaagag ctaccaactc
     4081 tttttccgaa ggtaactggc ttcagcagag cgcagatacc aaatactgtt cttctagtgt
     4141 agccgtagtt aggccaccac ttcaagaact ctgtagcacc gcctacatac ctcgctctgc
     4201 taatcctgtt accagtggct gctgccagtg gcgataagtc gtgtcttacc gggttggact
     4261 caagacgata gttaccggat aaggcgcagc ggtcgggctg aacggggggt tcgtgcacac
     4321 agcccagctt ggagcgaacg acctacaccg aactgagata cctacagcgt gagctatgag
     4381 aaagcgccac gcttcccgaa gggagaaagg cggacaggta tccggtaagc ggcagggtcg
     4441 gaacaggaga gcgcacgagg gagcttccag ggggaaacgc ctggtatctt tatagtcctg
     4501 tcgggtttcg ccacctctga cttgagcgtc gatttttgtg atgctcgtca ggggggcgga
     4561 gcctatggaa aaacgccagc aacgcggcct ttttacggtt cctggccttt tgctggcctt
     4621 ttgctcacat gttctttcct gcgttatccc ctgattctgt ggataaccgt attaccgcct
     4681 ttgagtgagc tgataccgct cgccgcagcc gaacgaccga gcgcagcgag tcagtgagcg
     4741 aggaagcgga agagcgccca atacgcaaac cgcctctccc cgcgcgttgg ccgattcatt
     4801 aatgcagctg gcacgacagg tttcccgact ggaaagcggg cagtgagcgc aacgcaatta
     4861 atgtgagtta gctcactcat taggcacccc aggctttaca ctttatgctt ccggctcgta
     4921 tgttgtgtgg aattgtgagc ggataacaat ttcacacagg aaacagctat gaccatgatt
     4981 acgccagatt taattaagg
//
