LOCUS       sequence-202607-        5787 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Generates Nonsense mutant clone.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5787)
  AUTHORS   Pattnaik BR, Shahi PK, Marino MJ, Liu X, York N, Brar S, Chiang J, 
            Pillers DA, Traboulsi EI
  TITLE     A Novel KCNJ13 Nonsense Mutation and Loss of Kir7.1 Channel Function
            Causes Leber Congenital Amaurosis (LCA16).
  JOURNAL   Hum Mutat. 2015 Jul;36(7):720-7. doi: 10.1002/humu.22807. Epub 2015 
            May 20.
  PUBMED    25921210
REFERENCE   2  (bases 1 to 5787)
  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: 
            "10.1002/humu.22807"; journalName: "Hum Mutat"; date: "2015-07"; 
            volume: "36"; issue: "7"; pages: "720-7"
COMMENT     Sequence Label: c1GFP-W53XKir7.1
FEATURES             Location/Qualifiers
     source          1..5787
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        61..364
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        365..568
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     519..539
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     regulatory      607..616
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(613..615,616..618,619..1329)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             join(613..615,616..618,619..1329)
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(658..679)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(919..938)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1266..1287
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     polyA_signal    2575..2696
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2612..2631)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     2666..2685
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(2703..3158)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2840..2861)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     3052..3071
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        3185..3289
                     /gene="bla"
                     /label=AmpR promoter
     promoter        3291..3648
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        3291..3482
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(3314..3334)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      3499..3634
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             3683..4477
                     /codon_start=1
                     /product="aminoglycoside phosphotransferase"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     CDS             3683..4477
                     /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
                     GLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(3737..3756)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4347..4366
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(4665..4684)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    4709..4756
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      5085..5673
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5574..5593
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata tggagttccg
       61 cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc cccgcccatt
      121 gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc attgacgtca
      181 atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt atcatatgcc
      241 aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt atgcccagta
      301 catgacctta tgggactttc ctacttggca gtacatctac gtattagtca tcgctattac
      361 catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg actcacgggg
      421 atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc aaaatcaacg
      481 ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg gtaggcgtgt
      541 acggtgggag gtctatataa gcagagctgg tttagtgaac cgtcagatcc gctagcgcta
      601 ccggtcgcca ccatggtgag caagggcgag gagctgttca ccggggtggt gcccatcctg
      661 gtcgagctgg acggcgacgt aaacggccac aagttcagcg tgtccggcga gggcgagggc
      721 gatgccacct acggcaagct gaccctgaag ttcatctgca ccaccggcaa gctgcccgtg
      781 ccctggccca ccctcgtgac caccctgacc tacggcgtgc agtgcttcag ccgctacccc
      841 gaccacatga agcagcacga cttcttcaag tccgccatgc ccgaaggcta cgtccaggag
      901 cgcaccatct tcttcaagga cgacggcaac tacaagaccc gcgccgaggt gaagttcgag
      961 ggcgacaccc tggtgaaccg catcgagctg aagggcatcg acttcaagga ggacggcaac
     1021 atcctggggc acaagctgga gtacaactac aacagccaca acgtctatat catggccgac
     1081 aagcagaaga acggcatcaa ggtgaacttc aagatccgcc acaacatcga ggacggcagc
     1141 gtgcagctcg ccgaccacta ccagcagaac acccccatcg gcgacggccc cgtgctgctg
     1201 cccgacaacc actacctgag cacccagtcc gccctgagca aagaccccaa cgagaagcgc
     1261 gatcacatgg tcctgctgga gttcgtgacc gccgccggga tcactctcgg catggacgag
     1321 ctgtacaagt ccggactcag atctcgagct caagcttcga attctgacag cagtaattgc
     1381 aaagttattg ctcctctcct aagtcaaaga taccggagga tggtcaccaa ggatggccac
     1441 agcacacttc aaatggatgg cgctcaaaga ggtcttgcat atcttcgaga tgcttgggga
     1501 atcctaatgg acatgcgcta gcgttggatg atgttggtct tttctgcttc ttttgttgtc
     1561 cactggcttg tctttgcagt gctctggtat gttctggctg agatgaatgg tgatctggaa
     1621 ctagatcatg atgccccacc tgaaaaccac actatctgtg tcaagtatat caccagtttc
     1681 acagctgcat tctccttctc cctggagaca caactcacaa ttggttatgg taccatgttc
     1741 cccagtggtg actgtccaag tgcaatcgcc ttacttgcca tacaaatgct cctaggcctc
     1801 atgctagagg cttttatcac aggtgctttt gtggcgaaga ttgcccggcc aaaaaatcga
     1861 gctttttcaa ttcgctttac tgacacagca gtagtagctc acatggatgg caaacctaat
     1921 cttatcttcc aagtggccaa cacccgacct agccctctaa ccagtgtccg ggtctcagct
     1981 gtactctatc aggaaagaga aaatggcaaa ctctaccaga ccagtgtgga tttccacctt
     2041 gatggcatca gttctgacga atgtccattc ttcatctttc cactaacgta ctatcactcc
     2101 attacaccat caagtcctct ggctactctg ctccagcatg aaaatccttc tcactttgaa
     2161 ttagttgtat tcctttcagc aatgcaggag ggcactggag aaatatgcca aaggaggaca
     2221 tcctacctac cgtctgaaat catgttacat cactgttttg catctctgtt gacccgaggt
     2281 tccaaaggtg aatatcaaat caagatggag aattttgaca agactgtccc tgaatttcca
     2341 actcctctgg tttctaaaag cccaaacagg actgacctgg atatccacat caatggacaa
     2401 agcattgaca attttcagat ctctgaaaca ggactgacag aataaggatc caccggatct
     2461 agataactga tcataatcag ccataccaca tttgtagagg ttttacttgc tttaaaaaac
     2521 ctcccacacc tccccctgaa cctgaaacat aaaatgaatg caattgttgt tgttaacttg
     2581 tttattgcag cttataatgg ttacaaataa agcaatagca tcacaaattt cacaaataaa
     2641 gcattttttt cactgcattc tagttgtggt ttgtccaaac tcatcaatgt atcttaacgc
     2701 gtaaattgta agcgttaata ttttgttaaa attcgcgtta aatttttgtt aaatcagctc
     2761 attttttaac caataggccg aaatcggcaa aatcccttat aaatcaaaag aatagaccga
     2821 gatagggttg agtgttgttc cagtttggaa caagagtcca ctattaaaga acgtggactc
     2881 caacgtcaaa gggcgaaaaa ccgtctatca gggcgatggc ccactacgtg aaccatcacc
     2941 ctaatcaagt tttttggggt cgaggtgccg taaagcacta aatcggaacc ctaaagggag
     3001 cccccgattt agagcttgac ggggaaagcc ggcgaacgtg gcgagaaagg aagggaagaa
     3061 agcgaaagga gcgggcgcta gggcgctggc aagtgtagcg gtcacgctgc gcgtaaccac
     3121 cacacccgcc gcgcttaatg cgccgctaca gggcgcgtca ggtggcactt ttcggggaaa
     3181 tgtgcgcgga acccctattt gtttattttt ctaaatacat tcaaatatgt atccgctcat
     3241 gagacaataa ccctgataaa tgcttcaata atattgaaaa aggaagagtc ctgaggcgga
     3301 aagaaccagc tgtggaatgt gtgtcagtta gggtgtggaa agtccccagg ctccccagca
     3361 ggcagaagta tgcaaagcat gcatctcaat tagtcagcaa ccaggtgtgg aaagtcccca
     3421 ggctccccag caggcagaag tatgcaaagc atgcatctca attagtcagc aaccatagtc
     3481 ccgcccctaa ctccgcccat cccgccccta actccgccca gttccgccca ttctccgccc
     3541 catggctgac taattttttt tatttatgca gaggccgagg ccgcctcggc ctctgagcta
     3601 ttccagaagt agtgaggagg cttttttgga ggcctaggct tttgcaaaga tcgatcaaga
     3661 gacaggatga ggatcgtttc gcatgattga acaagatgga ttgcacgcag gttctccggc
     3721 cgcttgggtg gagaggctat tcggctatga ctgggcacaa cagacaatcg gctgctctga
     3781 tgccgccgtg ttccggctgt cagcgcaggg gcgcccggtt ctttttgtca agaccgacct
     3841 gtccggtgcc ctgaatgaac tgcaagacga ggcagcgcgg ctatcgtggc tggccacgac
     3901 gggcgttcct tgcgcagctg tgctcgacgt tgtcactgaa gcgggaaggg actggctgct
     3961 attgggcgaa gtgccggggc aggatctcct gtcatctcac cttgctcctg ccgagaaagt
     4021 atccatcatg gctgatgcaa tgcggcggct gcatacgctt gatccggcta cctgcccatt
     4081 cgaccaccaa gcgaaacatc gcatcgagcg agcacgtact cggatggaag ccggtcttgt
     4141 cgatcaggat gatctggacg aagagcatca ggggctcgcg ccagccgaac tgttcgccag
     4201 gctcaaggcg agcatgcccg acggcgagga tctcgtcgtg acccatggcg atgcctgctt
     4261 gccgaatatc atggtggaaa atggccgctt ttctggattc atcgactgtg gccggctggg
     4321 tgtggcggac cgctatcagg acatagcgtt ggctacccgt gatattgctg aagagcttgg
     4381 cggcgaatgg gctgaccgct tcctcgtgct ttacggtatc gccgctcccg attcgcagcg
     4441 catcgccttc tatcgccttc ttgacgagtt cttctgagcg ggactctggg gttcgaaatg
     4501 accgaccaag cgacgcccaa cctgccatca cgagatttcg attccaccgc cgccttctat
     4561 gaaaggttgg gcttcggaat cgttttccgg gacgccggct ggatgatcct ccagcgcggg
     4621 gatctcatgc tggagttctt cgcccaccct agggggaggc taactgaaac acggaaggag
     4681 acaataccgg aaggaacccg cgctatgacg gcaataaaaa gacagaataa aacgcacggt
     4741 gttgggtcgt ttgttcataa acgcggggtt cggtcccagg gctggcactc tgtcgatacc
     4801 ccaccgagac cccattgggg ccaatacgcc cgcgtttctt ccttttcccc accccacccc
     4861 ccaagttcgg gtgaaggccc agggctcgca gccaacgtcg gggcggcagg ccctgccata
     4921 gcctcaggtt actcatatat actttagatt gatttaaaac ttcattttta atttaaaagg
     4981 atctaggtga agatcctttt tgataatctc atgaccaaaa tcccttaacg tgagttttcg
     5041 ttccactgag cgtcagaccc cgtagaaaag atcaaaggat cttcttgaga tccttttttt
     5101 ctgcgcgtaa tctgctgctt gcaaacaaaa aaaccaccgc taccagcggt ggtttgtttg
     5161 ccggatcaag agctaccaac tctttttccg aaggtaactg gcttcagcag agcgcagata
     5221 ccaaatactg tccttctagt gtagccgtag ttaggccacc acttcaagaa ctctgtagca
     5281 ccgcctacat acctcgctct gctaatcctg ttaccagtgg ctgctgccag tggcgataag
     5341 tcgtgtctta ccgggttgga ctcaagacga tagttaccgg ataaggcgca gcggtcgggc
     5401 tgaacggggg gttcgtgcac acagcccagc ttggagcgaa cgacctacac cgaactgaga
     5461 tacctacagc gtgagctatg agaaagcgcc acgcttcccg aagggagaaa ggcggacagg
     5521 tatccggtaa gcggcagggt cggaacagga gagcgcacga gggagcttcc agggggaaac
     5581 gcctggtatc tttatagtcc tgtcgggttt cgccacctct gacttgagcg tcgatttttg
     5641 tgatgctcgt caggggggcg gagcctatgg aaaaacgcca gcaacgcggc ctttttacgg
     5701 ttcctggcct tttgctggcc ttttgctcac atgttctttc ctgcgttatc ccctgattct
     5761 gtggataacc gtattaccgc catgcat
//
