LOCUS       sequence-413578-        6700 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Mammalian expression of human NKX6-2 cDNA SPAX8-related truncated 
            mutation containing the amino acids 1 to 196; fused with the green 
            fluorescent protein Venus allowing the use of fluorescent based 
            protocols..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6700)
  AUTHORS   Ferreira-Peralta P, Franca B, Murtinheira F, Rodrigues MS, Herrera F
  TITLE     Is spastic ataxia 8 a protein misfolding disorder?
  JOURNAL   Biochim Biophys Acta Mol Basis Dis. 2023 Sep 9;1870(1):166882. doi: 
            10.1016/j.bbadis.2023.166882.
  PUBMED    37696463
REFERENCE   2  (bases 1 to 6700)
  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"; journalName: "Biochim 
            Biophys Acta Mol Basis Dis."; date: "2023-09-9"; volume: 
            "1870(1):166882. doi"; pages: " 10.1016/j.bbadis.2023.166882"
COMMENT     Sequence Label: pcDNA3.1 NKX6-2(1-196)-Venus
FEATURES             Location/Qualifiers
     source          1..6700
                     /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"
     primer_bind     complement(1843..1862)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     complement(2291..2308)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    2297..2521
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      2567..2995
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2654..2673)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2864..2885
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(3004..3024)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        3009..3338
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      3189..3324
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     3251..3270
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             3405..4199
                     /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(3459..3478)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4069..4088
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    4373..4494
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4410..4429)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4464..4483
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(4543..4559)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4543..4559)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4556..4578)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4567..4583
                     /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(4591..4597,4598..4615,4616..4621))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4636..4657
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4774..4791)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4945..5533)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5025..5044)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5704..6495,6496..6564))
                     /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     6327..6346
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6565..6669)
                     /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 gacaccatgg acacaaacag acctggcgcc tttgtgctgt ctagcgctcc tctggccgct
      961 ctgcacaata tggccgagat gaagaccagc ctgtttccat acgctctgca gggacctgcc
     1021 ggctttaagg ctcctgctct tggaggactg ggagcacaac tgcctctggg aacacctcac
     1081 ggcatcagcg atattctggg cagacctgtt ggagctgctg gcggaggatt gctcggagga
     1141 ctgcctagac tgaatggact ggcttcttct gccggcgtgt actttggacc tgctgctgcc
     1201 gttgccagag gctatcctaa acctctggct gagcttcccg gcagacctcc tattttttgg
     1261 cctggtgttg tgcagggcgc tccttggaga gatcctagac ttgctggacc agctcctgca
     1321 ggcggagtgc tggataagga tggcaagaag aagcacagca gacccacctt cagcggccag
     1381 cagatctttg ccctggaaaa gaccttcgag cagaccaagt atctggctgg ccctgagaga
     1441 gccagactgg cctattctct gggcatgacc gagagccaag tgaaagtgtg gttcaagctt
     1501 ggcggaggtg gaagcggcgg aggcggatct ggtaccatgg tttctaaggg cgaagaactg
     1561 ttcaccggcg tggtgcctat cctggtggaa ctggatgggg atgtgaacgg ccacaagttc
     1621 tctgtgtctg gcgaaggcga aggggatgcc acatacggca agctgaccct gaagctgatc
     1681 tgcaccacag gcaaactgcc cgtgccttgg cctacactgg tcacaacact cggctacggc
     1741 ctgcagtgct ttgccagata tcccgaccac atgaagcagc acgatttctt caagagcgcc
     1801 atgcctgagg gctacgtgca agagcggacc atcttcttca aggacgacgg caactacaag
     1861 accagggccg aagtgaagtt cgagggcgac accctggtca acagaatcga gctgaagggc
     1921 atcgacttta aagaggacgg caacatcctg ggccacaagc tcgagtacaa ctacaacagc
     1981 cacaacgtgt acatcaccgc cgacaagcag aagaacggca tcaaggccaa cttcaagatc
     2041 cggcacaaca tcgaggatgg cggcgtgcag ctggccgatc actatcagca gaacacccct
     2101 atcggagatg gccccgtcct gctgcctgat aaccactacc tgagctacca gtctgccctg
     2161 agcaaggacc ccaatgagaa gagggatcac atggtgctgc tggaattcgt gacagccgcc
     2221 ggaatcacac tcggcatgga cgagctgtac aaatgaggat cctaagggcc cgtttaaacc
     2281 cgctgatcag cctcgactgt gccttctagt tgccagccat ctgttgtttg cccctccccc
     2341 gtgccttcct tgaccctgga aggtgccact cccactgtcc tttcctaata aaatgaggaa
     2401 attgcatcgc attgtctgag taggtgtcat tctattctgg ggggtggggt ggggcaggac
     2461 agcaaggggg aggattggga agacaatagc aggcatgctg gggatgcggt gggctctatg
     2521 gcttctgagg cggaaagaac cagctggggc tctagggggt atccccacgc gccctgtagc
     2581 ggcgcattaa gcgcggcggg tgtggtggtt acgcgcagcg tgaccgctac acttgccagc
     2641 gccctagcgc ccgctccttt cgctttcttc ccttcctttc tcgccacgtt cgccggcttt
     2701 ccccgtcaag ctctaaatcg ggggctccct ttagggttcc gatttagtgc tttacggcac
     2761 ctcgacccca aaaaacttga ttagggtgat ggttcacgta gtgggccatc gccctgatag
     2821 acggtttttc gccctttgac gttggagtcc acgttcttta atagtggact cttgttccaa
     2881 actggaacaa cactcaaccc tatctcggtc tattcttttg atttataagg gattttgccg
     2941 atttcggcct attggttaaa aaatgagctg atttaacaaa aatttaacgc gaattaattc
     3001 tgtggaatgt gtgtcagtta gggtgtggaa agtccccagg ctccccagca ggcagaagta
     3061 tgcaaagcat gcatctcaat tagtcagcaa ccaggtgtgg aaagtcccca ggctccccag
     3121 caggcagaag tatgcaaagc atgcatctca attagtcagc aaccatagtc ccgcccctaa
     3181 ctccgcccat cccgccccta actccgccca gttccgccca ttctccgccc catggctgac
     3241 taattttttt tatttatgca gaggccgagg ccgcctctgc ctctgagcta ttccagaagt
     3301 agtgaggagg cttttttgga ggcctaggct tttgcaaaaa gctcccggga gcttgtatat
     3361 ccattttcgg atctgatcaa gagacaggat gaggatcgtt tcgcatgatt gaacaagatg
     3421 gattgcacgc aggttctccg gccgcttggg tggagaggct attcggctat gactgggcac
     3481 aacagacaat cggctgctct gatgccgccg tgttccggct gtcagcgcag gggcgcccgg
     3541 ttctttttgt caagaccgac ctgtccggtg ccctgaatga actgcaggac gaggcagcgc
     3601 ggctatcgtg gctggccacg acgggcgttc cttgcgcagc tgtgctcgac gttgtcactg
     3661 aagcgggaag ggactggctg ctattgggcg aagtgccggg gcaggatctc ctgtcatctc
     3721 accttgctcc tgccgagaaa gtatccatca tggctgatgc aatgcggcgg ctgcatacgc
     3781 ttgatccggc tacctgccca ttcgaccacc aagcgaaaca tcgcatcgag cgagcacgta
     3841 ctcggatgga agccggtctt gtcgatcagg atgatctgga cgaagagcat caggggctcg
     3901 cgccagccga actgttcgcc aggctcaagg cgcgcatgcc cgacggcgag gatctcgtcg
     3961 tgacccatgg cgatgcctgc ttgccgaata tcatggtgga aaatggccgc ttttctggat
     4021 tcatcgactg tggccggctg ggtgtggcgg accgctatca ggacatagcg ttggctaccc
     4081 gtgatattgc tgaagagctt ggcggcgaat gggctgaccg cttcctcgtg ctttacggta
     4141 tcgccgctcc cgattcgcag cgcatcgcct tctatcgcct tcttgacgag ttcttctgag
     4201 cgggactctg gggttcgaaa tgaccgacca agcgacgccc aacctgccat cacgagattt
     4261 cgattccacc gccgccttct atgaaaggtt gggcttcgga atcgttttcc gggacgccgg
     4321 ctggatgatc ctccagcgcg gggatctcat gctggagttc ttcgcccacc ccaacttgtt
     4381 tattgcagct tataatggtt acaaataaag caatagcatc acaaatttca caaataaagc
     4441 atttttttca ctgcattcta gttgtggttt gtccaaactc atcaatgtat cttatcatgt
     4501 ctgtataccg tcgacctcta gctagagctt ggcgtaatca tggtcatagc tgtttcctgt
     4561 gtgaaattgt tatccgctca caattccaca caacatacga gccggaagca taaagtgtaa
     4621 agcctggggt gcctaatgag tgagctaact cacattaatt gcgttgcgct cactgcccgc
     4681 tttccagtcg ggaaacctgt cgtgccagct gcattaatga atcggccaac gcgcggggag
     4741 aggcggtttg cgtattgggc gctcttccgc ttcctcgctc actgactcgc tgcgctcggt
     4801 cgttcggctg cggcgagcgg tatcagctca ctcaaaggcg gtaatacggt tatccacaga
     4861 atcaggggat aacgcaggaa agaacatgtg agcaaaaggc cagcaaaagg ccaggaaccg
     4921 taaaaaggcc gcgttgctgg cgtttttcca taggctccgc ccccctgacg agcatcacaa
     4981 aaatcgacgc tcaagtcaga ggtggcgaaa cccgacagga ctataaagat accaggcgtt
     5041 tccccctgga agctccctcg tgcgctctcc tgttccgacc ctgccgctta ccggatacct
     5101 gtccgccttt ctcccttcgg gaagcgtggc gctttctcat agctcacgct gtaggtatct
     5161 cagttcggtg taggtcgttc gctccaagct gggctgtgtg cacgaacccc ccgttcagcc
     5221 cgaccgctgc gccttatccg gtaactatcg tcttgagtcc aacccggtaa gacacgactt
     5281 atcgccactg gcagcagcca ctggtaacag gattagcaga gcgaggtatg taggcggtgc
     5341 tacagagttc ttgaagtggt ggcctaacta cggctacact agaagaacag tatttggtat
     5401 ctgcgctctg ctgaagccag ttaccttcgg aaaaagagtt ggtagctctt gatccggcaa
     5461 acaaaccacc gctggtagcg gtggtttttt tgtttgcaag cagcagatta cgcgcagaaa
     5521 aaaaggatct caagaagatc ctttgatctt ttctacgggg tctgacgctc agtggaacga
     5581 aaactcacgt taagggattt tggtcatgag attatcaaaa aggatcttca cctagatcct
     5641 tttaaattaa aaatgaagtt ttaaatcaat ctaaagtata tatgagtaaa cttggtctga
     5701 cagttaccaa tgcttaatca gtgaggcacc tatctcagcg atctgtctat ttcgttcatc
     5761 catagttgcc tgactccccg tcgtgtagat aactacgata cgggagggct taccatctgg
     5821 ccccagtgct gcaatgatac cgcgagaccc acgctcaccg gctccagatt tatcagcaat
     5881 aaaccagcca gccggaaggg ccgagcgcag aagtggtcct gcaactttat ccgcctccat
     5941 ccagtctatt aattgttgcc gggaagctag agtaagtagt tcgccagtta atagtttgcg
     6001 caacgttgtt gccattgcta caggcatcgt ggtgtcacgc tcgtcgtttg gtatggcttc
     6061 attcagctcc ggttcccaac gatcaaggcg agttacatga tcccccatgt tgtgcaaaaa
     6121 agcggttagc tccttcggtc ctccgatcgt tgtcagaagt aagttggccg cagtgttatc
     6181 actcatggtt atggcagcac tgcataattc tcttactgtc atgccatccg taagatgctt
     6241 ttctgtgact ggtgagtact caaccaagtc attctgagaa tagtgtatgc ggcgaccgag
     6301 ttgctcttgc ccggcgtcaa tacgggataa taccgcgcca catagcagaa ctttaaaagt
     6361 gctcatcatt ggaaaacgtt cttcggggcg aaaactctca aggatcttac cgctgttgag
     6421 atccagttcg atgtaaccca ctcgtgcacc caactgatct tcagcatctt ttactttcac
     6481 cagcgtttct gggtgagcaa aaacaggaag gcaaaatgcc gcaaaaaagg gaataagggc
     6541 gacacggaaa tgttgaatac tcatactctt cctttttcaa tattattgaa gcatttatca
     6601 gggttattgt ctcatgagcg gatacatatt tgaatgtatt tagaaaaata aacaaatagg
     6661 ggttccgcgc acatttcccc gaaaagtgcc acctgacgtc
//
