LOCUS       sequence-377241-        7635 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses wild-type myocilin with a C-terminal enhanced Gaussia 
            luciferase and FLAG tag.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7635)
  AUTHORS   Nakahara E, Hulleman JD
  TITLE     A Simple Secretion Assay for Assessing New and Existing Myocilin 
            Variants.
  JOURNAL   Curr Eye Res. 2022 Jun;47(6):918-922. doi: 
            10.1080/02713683.2022.2047205. Epub 2022 Mar 25.
  PUBMED    35196929
REFERENCE   2  (bases 1 to 7635)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Dec 15, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; doi: 
            "10.1080/02713683.2022.2047205"; journalName: "Curr Eye Res"; date: 
            "2022-06"; volume: "47"; issue: "6"; pages: "918-922"
COMMENT     Sequence Label: pcDNA3 WT MYOC eGLuc2 FLAG
FEATURES             Location/Qualifiers
     source          1..7635
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(44..63)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     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"
     primer_bind     769..789
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     863..882
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        863..881
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             3016..3039
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     promoter        complement(3188..3206)
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     complement(3189..3206)
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     primer_bind     complement(3226..3243)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    3232..3456
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      3502..3930
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3589..3608)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3799..3820
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(3939..3959)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        3944..4273
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      4124..4259
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     4186..4205
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             4340..5134
                     /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(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(4394..4413)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     5004..5023
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    5308..5429
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5345..5364)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     5399..5418
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(5478..5494)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5478..5494)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(5491..5513)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5502..5518
                     /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(join(5526..5532,5533..5550,5551..5556))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5571..5592
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5709..5726)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5880..6468)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5960..5979)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6639..7430,7431..7499))
                     /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     7262..7281
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7500..7604)
                     /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 gcttggtacc
      901 gagctcggat ccagagcttt ccagaggaag cctcaccaag cctctgcaat gaggttcttc
      961 tgtgcacgtt gctgcagctt tgggcctgag atgccagctg tccagctgct gcttctggcc
     1021 tgcctggtgt gggatgtggg ggccaggaca gctcagctca ggaaggccaa tgaccagagt
     1081 ggccgatgcc agtatacctt cagtgtggcc agtcccaatg aatccagctg cccagagcag
     1141 agccaggcca tgtcagtcat ccataactta cagagagaca gcagcaccca acgcttagac
     1201 ctggaggcca ccaaagctcg actcagctcc ctggagagcc tcctccacca attgaccttg
     1261 gaccaggctg ccaggcccca ggagacccag gaggggctgc agagggagct gggcaccctg
     1321 aggcgggagc gggaccagct ggaaacccaa accagagagt tggagactgc ctacagcaac
     1381 ctcctccgag acaagtcagt tctggaggaa gagaagaagc gactaaggca agaaaatgag
     1441 aatctggcca ggaggttgga aagcagcagc caggaggtag caaggctgag aaggggccag
     1501 tgtccccaga cccgagacac tgctcgggct gtgccaccag gctccagaga agtttctacg
     1561 tggaatttgg acactttggc cttccaggaa ctgaagtccg agctaactga agttcctgct
     1621 tcccgaattt tgaaggagag cccatctggc tatctcagga gtggagaggg agacaccgga
     1681 tgtggagaac tagtttgggt aggagagcct ctcacgctga gaacagcaga aacaattact
     1741 ggcaagtatg gtgtgtggat gcgagacccc aagcccacct acccctacac ccaggagacc
     1801 acgtggagaa tcgacacagt tggcacggat gtccgccagg tttttgagta tgacctcatc
     1861 agccagttta tgcagggcta cccttctaag gttcacatac tgcctaggcc actggaaagc
     1921 acgggtgctg tggtgtactc ggggagcctc tatttccagg gcgctgagtc cagaactgtc
     1981 ataagatatg agctgaatac cgagacagtg aaggctgaga aggaaatccc tggagctggc
     2041 taccacggac agttcccgta ttcttggggt ggctacacgg acattgactt ggctgtggat
     2101 gaagcaggcc tctgggtcat ttacagcacc gatgaggcca aaggtgccat tgtcctctcc
     2161 aaactgaacc cagagaatct ggaactcgaa caaacctggg agacaaacat ccgtaagcag
     2221 tcagtcgcca atgccttcat catctgtggc accttgtaca ccgtcagcag ctacacctca
     2281 gcagatgcta ccgtcaactt tgcttatgac acaggcacag gtatcagcaa gaccctgacc
     2341 atcccattca agaaccgcta taagtacagc agcatgattg actacaaccc cctggagaag
     2401 aagctctttg cctgggacaa cttgaacatg gtcacttatg acatcaagct ctccaagatg
     2461 atgttcgaag gctccgctgg ctccgctgct ggttctggcg aatttgaggc caagcccacc
     2521 gagaacaacg aagacttcaa catcgtggcc gtggccagca acttcgcgac cacggatctc
     2581 gatgctgacc gcgggaagtt gcccggcaag aagctgccgc tggaggtgct caaagagata
     2641 gaagccaatg cccggaaagc tggctgcacc aggggctgtc tgatctgcct gtcccacatc
     2701 aagtgcacgc ccaagatgaa gaagttcatc ccaggacgct gccacaccta cgaaggcgac
     2761 aaagagtccg cacagggcgg cataggcgag gcgatcgtcg acattcctga gattcctggg
     2821 ttcaaggact tggagcccat agagcagttc atcgcacagg tcgatctgtg tgtggactgc
     2881 acaactggct gcctcaaagg gcttgccaac gtgcagtgtt ctgacctgct caagaagtgg
     2941 ctgccgcaac gctgtgcgac ctttgccagc aagatccagg gccaggtgga caagatcaag
     3001 ggggccggtg gtgacgatta caaggacgac gatgacaagt gaaaagcctc caagctgtac
     3061 aggcaatggc agaaggagat gctaaattca ctaatcggat ccactagtaa cggccgccag
     3121 tgtgctggaa ttctgcagat atccatcaca ctggcggccg ctcgagcatg catctagagg
     3181 gccctattct atagtgtcac ctaaatgcta gagctcgctg atcagcctcg actgtgcctt
     3241 ctagttgcca gccatctgtt gtttgcccct cccccgtgcc ttccttgacc ctggaaggtg
     3301 ccactcccac tgtcctttcc taataaaatg aggaaattgc atcgcattgt ctgagtaggt
     3361 gtcattctat tctggggggt ggggtggggc aggacagcaa gggggaggat tgggaagaca
     3421 atagcaggca tgctggggat gcggtgggct ctatggcttc tgaggcggaa agaaccagct
     3481 ggggctctag ggggtatccc cacgcgccct gtagcggcgc attaagcgcg gcgggtgtgg
     3541 tggttacgcg cagcgtgacc gctacacttg ccagcgccct agcgcccgct cctttcgctt
     3601 tcttcccttc ctttctcgcc acgttcgccg gctttccccg tcaagctcta aatcgggggc
     3661 tccctttagg gttccgattt agtgctttac ggcacctcga ccccaaaaaa cttgattagg
     3721 gtgatggttc acgtagtggg ccatcgccct gatagacggt ttttcgccct ttgacgttgg
     3781 agtccacgtt ctttaatagt ggactcttgt tccaaactgg aacaacactc aaccctatct
     3841 cggtctattc ttttgattta taagggattt tgccgatttc ggcctattgg ttaaaaaatg
     3901 agctgattta acaaaaattt aacgcgaatt aattctgtgg aatgtgtgtc agttagggtg
     3961 tggaaagtcc ccaggctccc cagcaggcag aagtatgcaa agcatgcatc tcaattagtc
     4021 agcaaccagg tgtggaaagt ccccaggctc cccagcaggc agaagtatgc aaagcatgca
     4081 tctcaattag tcagcaacca tagtcccgcc cctaactccg cccatcccgc ccctaactcc
     4141 gcccagttcc gcccattctc cgccccatgg ctgactaatt ttttttattt atgcagaggc
     4201 cgaggccgcc tctgcctctg agctattcca gaagtagtga ggaggctttt ttggaggcct
     4261 aggcttttgc aaaaagctcc cgggagcttg tatatccatt ttcggatctg atcaagagac
     4321 aggatgagga tcgtttcgca tgattgaaca agatggattg cacgcaggtt ctccggccgc
     4381 ttgggtggag aggctattcg gctatgactg ggcacaacag acaatcggct gctctgatgc
     4441 cgccgtgttc cggctgtcag cgcaggggcg cccggttctt tttgtcaaga ccgacctgtc
     4501 cggtgccctg aatgaactgc aggacgaggc agcgcggcta tcgtggctgg ccacgacggg
     4561 cgttccttgc gcagctgtgc tcgacgttgt cactgaagcg ggaagggact ggctgctatt
     4621 gggcgaagtg ccggggcagg atctcctgtc atctcacctt gctcctgccg agaaagtatc
     4681 catcatggct gatgcaatgc ggcggctgca tacgcttgat ccggctacct gcccattcga
     4741 ccaccaagcg aaacatcgca tcgagcgagc acgtactcgg atggaagccg gtcttgtcga
     4801 tcaggatgat ctggacgaag agcatcaggg gctcgcgcca gccgaactgt tcgccaggct
     4861 caaggcgcgc atgcccgacg gcgaggatct cgtcgtgacc catggcgatg cctgcttgcc
     4921 gaatatcatg gtggaaaatg gccgcttttc tggattcatc gactgtggcc ggctgggtgt
     4981 ggcggaccgc tatcaggaca tagcgttggc tacccgtgat attgctgaag agcttggcgg
     5041 cgaatgggct gaccgcttcc tcgtgcttta cggtatcgcc gctcccgatt cgcagcgcat
     5101 cgccttctat cgccttcttg acgagttctt ctgagcggga ctctggggtt cgaaatgacc
     5161 gaccaagcga cgcccaacct gccatcacga gatttcgatt ccaccgccgc cttctatgaa
     5221 aggttgggct tcggaatcgt tttccgggac gccggctgga tgatcctcca gcgcggggat
     5281 ctcatgctgg agttcttcgc ccaccccaac ttgtttattg cagcttataa tggttacaaa
     5341 taaagcaata gcatcacaaa tttcacaaat aaagcatttt tttcactgca ttctagttgt
     5401 ggtttgtcca aactcatcaa tgtatcttat catgtctgta taccgtcgac ctctagctag
     5461 agcttggcgt aatcatggtc atagctgttt cctgtgtgaa attgttatcc gctcacaatt
     5521 ccacacaaca tacgagccgg aagcataaag tgtaaagcct ggggtgccta atgagtgagc
     5581 taactcacat taattgcgtt gcgctcactg cccgctttcc agtcgggaaa cctgtcgtgc
     5641 cagctgcatt aatgaatcgg ccaacgcgcg gggagaggcg gtttgcgtat tgggcgctct
     5701 tccgcttcct cgctcactga ctcgctgcgc tcggtcgttc ggctgcggcg agcggtatca
     5761 gctcactcaa aggcggtaat acggttatcc acagaatcag gggataacgc aggaaagaac
     5821 atgtgagcaa aaggccagca aaaggccagg aaccgtaaaa aggccgcgtt gctggcgttt
     5881 ttccataggc tccgcccccc tgacgagcat cacaaaaatc gacgctcaag tcagaggtgg
     5941 cgaaacccga caggactata aagataccag gcgtttcccc ctggaagctc cctcgtgcgc
     6001 tctcctgttc cgaccctgcc gcttaccgga tacctgtccg cctttctccc ttcgggaagc
     6061 gtggcgcttt ctcatagctc acgctgtagg tatctcagtt cggtgtaggt cgttcgctcc
     6121 aagctgggct gtgtgcacga accccccgtt cagcccgacc gctgcgcctt atccggtaac
     6181 tatcgtcttg agtccaaccc ggtaagacac gacttatcgc cactggcagc agccactggt
     6241 aacaggatta gcagagcgag gtatgtaggc ggtgctacag agttcttgaa gtggtggcct
     6301 aactacggct acactagaag aacagtattt ggtatctgcg ctctgctgaa gccagttacc
     6361 ttcggaaaaa gagttggtag ctcttgatcc ggcaaacaaa ccaccgctgg tagcggtggt
     6421 ttttttgttt gcaagcagca gattacgcgc agaaaaaaag gatctcaaga agatcctttg
     6481 atcttttcta cggggtctga cgctcagtgg aacgaaaact cacgttaagg gattttggtc
     6541 atgagattat caaaaaggat cttcacctag atccttttaa attaaaaatg aagttttaaa
     6601 tcaatctaaa gtatatatga gtaaacttgg tctgacagtt accaatgctt aatcagtgag
     6661 gcacctatct cagcgatctg tctatttcgt tcatccatag ttgcctgact ccccgtcgtg
     6721 tagataacta cgatacggga gggcttacca tctggcccca gtgctgcaat gataccgcga
     6781 gacccacgct caccggctcc agatttatca gcaataaacc agccagccgg aagggccgag
     6841 cgcagaagtg gtcctgcaac tttatccgcc tccatccagt ctattaattg ttgccgggaa
     6901 gctagagtaa gtagttcgcc agttaatagt ttgcgcaacg ttgttgccat tgctacaggc
     6961 atcgtggtgt cacgctcgtc gtttggtatg gcttcattca gctccggttc ccaacgatca
     7021 aggcgagtta catgatcccc catgttgtgc aaaaaagcgg ttagctcctt cggtcctccg
     7081 atcgttgtca gaagtaagtt ggccgcagtg ttatcactca tggttatggc agcactgcat
     7141 aattctctta ctgtcatgcc atccgtaaga tgcttttctg tgactggtga gtactcaacc
     7201 aagtcattct gagaatagtg tatgcggcga ccgagttgct cttgcccggc gtcaatacgg
     7261 gataataccg cgccacatag cagaacttta aaagtgctca tcattggaaa acgttcttcg
     7321 gggcgaaaac tctcaaggat cttaccgctg ttgagatcca gttcgatgta acccactcgt
     7381 gcacccaact gatcttcagc atcttttact ttcaccagcg tttctgggtg agcaaaaaca
     7441 ggaaggcaaa atgccgcaaa aaagggaata agggcgacac ggaaatgttg aatactcata
     7501 ctcttccttt ttcaatatta ttgaagcatt tatcagggtt attgtctcat gagcggatac
     7561 atatttgaat gtatttagaa aaataaacaa ataggggttc cgcgcacatt tccccgaaaa
     7621 gtgccacctg acgtc
//
