LOCUS       sequence-435466-        7876 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Mammalian expression of improved GABA sensor (negative change in 
            fluorescence).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7876)
  AUTHORS   Kolb I, Hasseman JP, Matsumoto A, Arthur BJ, Zhang Y, Tsang A, Reep 
            D, Tsegaye G, Zheng J, Patel R, Looger LL, Marvin JS, Korff WL, 
            Yonehara K, Turner GC
  TITLE     iGABASnFR2: Improved genetically encoded protein sensors of GABA
  JOURNAL   bioRxiv 2025.03.25.644953
REFERENCE   2  (bases 1 to 7876)
  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: "bioRxiv 
            2025.03.25.644953"
COMMENT     Sequence Label: pGP-CAG-iGABASnFR2n-WPRE-bGH-polyA
FEATURES             Location/Qualifiers
     source          1..7876
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        5..384
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     intron          662..1678
                     /label=chimeric intron
                     /note="chimera between introns from chicken beta-actin and 
                     rabbit beta-globin"
     primer_bind     1602..1624
                     /label=pCAGGS-5
                     /note="Chimeric intron in CAG promoter, forward primer"
     primer_bind     1686..1705
                     /label=pCAG-F
                     /note="Rabbit beta-globin intron, for pCAG plasmids, 
                     forward primer"
     regulatory      1760..1769
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     sig_peptide     1766..1825
                     /codon_start=1
                     /product="leader sequence from mouse immunoglobulin kappa 
                     light chain"
                     /label=Ig-kappa leader
                     /translation="METDTLLLWVLLLWVPGSTG"
     primer_bind     2872..2893
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     complement(2996..3017)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     CDS             3524..3553
                     /codon_start=1
                     /product="Myc (human c-Myc proto-oncogene) epitope tag"
                     /label=Myc
                     /translation="EQKLISEEDL"
     CDS             3557..3703
                     /codon_start=1
                     /product="transmembrane domain from platelet derived growth
                     factor receptor beta"
                     /label=PDGFR-beta TM domain
                     /translation="AVGQDTQEVIVVPHSLPFKVVVISAILALVVLTIISLIILIMLWQ
                     KKPR"
     intron          3772..3904
                     /label=chimeric intron
                     /note="chimera between introns from human beta-globin and 
                     immunoglobulin heavy chain genes"
     primer_bind     3887..3906
                     /label=pMT2-F
                     /note="Synthetic intron, forward primer"
     misc_feature    3913..4501
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(3966..3986)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(4384..4395)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     primer_bind     complement(4528..4545)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    4534..4645
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(4741..4761)
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        complement(4741..4759)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(4780..4796)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4898..4917)
                     /label=Bglob-pA-R
                     /note="Rabbit beta-globin polyA region, reverse primer"
     polyA_signal    4963..5018
                     /label=beta-globin poly(A) signal
                     /note="rabbit beta-globin polyadenylation signal (Gil and 
                     Proudfoot, 1987)"
     primer_bind     complement(5017..5036)
                     /label=rbglobpA-R
                     /note="Rabbit beta-globin polyA, reverse primer. Also 
                     called rb-glob-pA-term-R"
     primer_bind     complement(5379..5395)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    5403..5419
                     /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(5427..5433,5434..5451,5452..5457))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5472..5493
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        5552..5747
                     /label=SV40 promoter
                     /note="SV40 early promoter"
     rep_origin      5598..5733
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     polyA_signal    5753..5887
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5790..5809)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     5844..5863
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(5955..5972)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(6126..6714)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(6206..6225)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6885..7676,7677..7745))
                     /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     7508..7527
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7746..7850)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 ggtcgacatt gattattgac tagttattaa tagtaatcaa ttacggggtc attagttcat
       61 agcccatata tggagttccg cgttacataa cttacggtaa atggcccgcc tggctgaccg
      121 cccaacgacc cccgcccatt gacgtcaata atgacgtatg ttcccatagt aacgccaata
      181 gggactttcc attgacgtca atgggtggag tatttacggt aaactgccca cttggcagta
      241 catcaagtgt atcatatgcc aagtacgccc cctattgacg tcaatgacgg taaatggccc
      301 gcctggcatt atgcccagta catgacctta tgggactttc ctacttggca gtacatctac
      361 gtattagtca tcgctattac catggtcgag gtgagcccca cgttctgctt cactctcccc
      421 atctcccccc cctccccacc cccaattttg tatttattta ttttttaatt attttgtgca
      481 gcgatggggg cggggggggg gggggggcgc gccrggsggg gggggggggg gsgggggggg
      541 gggsggggsg rggsggrgrg gkgcggcggc agccaatcag agcggcgcgc tccgaaagtt
      601 tccttttatg gcgaggcggc ggcggcggcg gccctataaa aagcgaagcg cgcggcgggc
      661 gggagtcgct gcgcgctgcc ttcgccccgt gccccgctcc gccgccgcct cgcgccgccc
      721 gccccggctc tgactgaccg cgttactccc acaggtgagc gggcgggacg gcccttctcc
      781 tccgggctgt aattagcgct tggtttaatg acggcttgtt tcttttctgt ggctgcgtga
      841 aagccttgag gggctccggg agggcccttt gtgcgggggg agcggctcgg ggggtgcgtg
      901 cgtgtgtgtg tgcgtgggga gcgccgcgtg cggctccgcg ctgcccggcg gctgtgagcg
      961 ctgcgggcgc ggcgcggggc tttgtgcgct ccgcagtgtg cgcgagggga gcgcggccgg
     1021 gggcggtgcc ccgcggtgcg gggggggctg cgaggggaac aaaggctgcg tgcggggtgt
     1081 gtgcgtgggg gggtgagcag ggggtgtggg cgcgtcggtc gggctgcaac cccccctgca
     1141 cccccctccc cgagttgctg agcacggccc ggcttcgggt gcggggctcc gtacggggcg
     1201 tggcgcgggg ctcgccgtgc cgggcggggg gtggcggcag gkgggggtgc cgggcggggc
     1261 ggggccgcck cgggccgggg rgggckcggg ggrggggcgc ggcggccccc ggagcgccgg
     1321 cggctgtcga ggcgcggcga gccgcagcca ttgcctttta tggtaatcgt gcgagagggc
     1381 gcagggactt cctttgtccc aaatctgtgc ggagccgaaa tctgggaggc gccgccgcac
     1441 cccctctagc gggcgcgggg cgaagcggtg cggcgccggc aggaaggaaa tgggcgggga
     1501 gggccttcgt gcgtcgccgc gccgccgtcc ccttctccct ctccagcctc ggggctgtcc
     1561 gcggggggac ggctgccttc gggggggacg gggcagggcg gggttcggct tctggcgtgt
     1621 gaccggcggc tctagagcct ctgctaacca tgttcatgcc ttcttctttt tcctacagct
     1681 cctgggcaac gtgctggtta ttgtgctgtc tcatcatttt ggcaaagaat tcggtaccgc
     1741 gggcccggga accggtgccg ccaccatgga gacagacaca ctcctgctat gggtactgct
     1801 gctctgggtt ccaggttcca ctggtgacag atctgaaagc atcaatttcg tgagctgggg
     1861 cggtagcacc caggatgcgc agaagcaggc ctgggccgac ccgttcagca aggccagcgg
     1921 cattaccgtg gtccaggatg ggcccaccga ctacggcaaa ctcaaggcca tggtcgaaag
     1981 cggcaacgtg cagtgggacg tggtggatgt agaggccgac ttcgccttgc gcgccgccgc
     2041 cgaaggcctg ctcgaacccc tggatttctc ggtgatccag cgcgacaaga tcgatccgcg
     2101 gttcgtgagt gaccatggag tgggggcctt cctgttctct cacgtgctcg gctacaacga
     2161 aggcaagctc ggagccagca agccccagga ctggaccgcg ctgttcgaca ccaagaccta
     2221 ccccggcaaa cgcgccctct acaaatggcc gagccctggc gtgctcgaac tggccctgct
     2281 cgccgacggc gtaccggccg acaagctcta cccgctggac ctggaccgcg ccttcaagaa
     2341 actcgacacc atcaagaaag acatcgtctg gtggggcggc ggtgcacagt cgcagcagct
     2401 gctggcctcc ggcgaagtca gcatgggcca gttctggaac ggtcgcatcc acgccctgca
     2461 agaagacggc gcgccagtgg gcgtgagctg gaagcagaac ctggtgatgg ccgacatcct
     2521 cgtcgtgcct aaaggcacga agaacaaagc cgccgcgatg aagttcctgg ccagtgccag
     2581 cagcgccaaa ggccaggacg acttttccgc cctgaccgcc tatgctccag tgaacatcga
     2641 cagtgtgcag cgcctcgacc ttgcgcaggt ccggatcacc gccgacaagc agaagaacgg
     2701 catcatggcg aacttcaaga tcccgcacaa cgtggaggac ggcagcgtgc agctcgccga
     2761 ccactaccag cagaacaccc ccatcggcga cggccccgtg ctgctgcccg acaaccacta
     2821 cctgagcacc cagtccgtgc tgagcaaaga ccccaacgag aagcgcgatc acatggtcct
     2881 gctggagttc gtgaccgccg ccgggatcac tctcggcatg gacgagctgt acaagggcgg
     2941 taccggaggg agcatgagca agggcgagga gctgttcacc ggggtggtgc ccatcctggt
     3001 cgagctggac ggcgacgtaa acggccacaa gttcagcgtg cgcggcgagg gcgagggcga
     3061 tgccaccaac ggcaagctga ccctgaagtt catctgcacc accggcaagc tgcccgtgcc
     3121 ctggcccacc ctcgtgacca ccctgaccta cggcgtgcag tgcttcagcc gctaccccga
     3181 ccacatgaag cagcacgact tcttcaagtc cgccatgccc gaaggctacg tccaggagcg
     3241 caccatcagc ttcaaggacg acggcaccta caagacccgc gccgaggtga agttcgaggg
     3301 cgacaccctg gtgaaccgca tcgagctgaa gggcatcgac ttcaaggagg acggcaacat
     3361 cctggggcac aagctggagt acaactggaa cgccaacctg gcaccgaacc tgccgaccgc
     3421 ctacgtcaag gatcagatca ccctcgattt cgcctactgg gccaagaacg gtccggccat
     3481 tgcgacacgg tggaatgaat ggctagtcaa actgcaggtc gacgaacaaa aactcatctc
     3541 agaagaggat ctgaatgctg tgggccagga cacgcaggag gtcatcgtgg tgccacactc
     3601 cttgcccttt aaggtggtgg tgatctcagc catcctggcc ctggtggtgc tcaccatcat
     3661 ctcccttatc atcctcatca tgctttggca gaagaagcca cgttaggaat tcgatatcgg
     3721 taccgatgaa tagctcgagg taccgatgaa tagctaaggt cgaggccgca ggtaagtatc
     3781 aaggttacaa gacaggttta aggagaccaa tagaaactgg gcttgtcgag acagagaaga
     3841 ctcttgcgtt tctgataggc acctattggt cttactgaca tccactttgc ctttctctcc
     3901 acaggtgtcg acaatcaacc tctggattac aaaatttgtg aaagattgac tggtattctt
     3961 aactatgttg ctccttttac gctatgtgga tacgctgctt taatgccttt gtatcatgct
     4021 attgcttccc gtatggcttt cattttctcc tccttgtata aatcctggtt gctgtctctt
     4081 tatgaggagt tgtggcccgt tgtcaggcaa cgtggcgtgg tgtgcactgt gtttgctgac
     4141 gcaaccccca ctggttgggg cattgccacc acctgtcagc tcctttccgg gactttcgct
     4201 ttccccctcc ctattgccac ggcggaactc atcgccgcct gccttgcccg ctgctggaca
     4261 ggggctcggc tgttgggcac tgacaattcc gtggtgttgt cggggaagct gacgtccttt
     4321 ccatggctgc tcgcctgtgt tgccacctgg attctgcgcg ggacgtcctt ctgctacgtc
     4381 ccttcggccc tcaatccagc ggaccttcct tcccgcggcc tgctgccggc tctgcggcct
     4441 cttccgcgtc ttcgccttcg ccctcagacg agtcggatct ccctttgggc cgcctccccg
     4501 cctggaattc gagctcggta cgatcagcct cgactgtgcc ttctagttgc cagccatctg
     4561 ttgtttgccc ctcccccgtg ccttccttga ccctggaagg tgccactccc actgtccttt
     4621 cctaataaaa tgaggaaatt gcatcgcatt gtctgagtag gtgtcattct attctggggg
     4681 gtggggtggg gcaggacagc aagggggagg attgggaaga caatagccca gcttttgttc
     4741 cctttagtga gggttaattg cgcgcttggc gtaatcatgg tcatagctgt ttcctgtgtg
     4801 aaattgttat ccgctaattc actcctcagg tgcaggctgc ctatcagaag gtggtggctg
     4861 gtgtggccaa tgccctggct cacaaatacc actgagatct ttttccctct gccaaaaatt
     4921 atggggacat catgaagccc cttgagcatc tgacttctgg ctaataaagg aaatttattt
     4981 tcattgcaat agtgtgttgg aattttttgt gtctctcact cggaaggaca tatgggaggg
     5041 caaatcattt aaaacatcag aatgagtatt tggtttagag tttggcaaca tatgcccata
     5101 tgctggctgc catgaacaaa ggttggctat aaagaggtca tcagtatatg aaacagcccc
     5161 ctgctgtcca ttccttattc catagaaaag ccttgacttg aggttagatt ttttttatat
     5221 tttgttttgt gttatttttt tctttaacat ccctaaaatt ttccttacat gttttactag
     5281 ccagattttt cctcctctcc tgactactcc cagtcatagc tgtccctctt ctcttatgga
     5341 gatccctcga cctgcagccc aagcttggcg taatcatggt catagctgtt tcctgtgtga
     5401 aattgttatc cgctcacaat tccacacaac atacgagccg gaagcataaa gtgtaaagcc
     5461 tggggtgcct aatgagtgag ctaactcaca ttaattgcgt tgcgctcact gcccgctttc
     5521 cagtcgggaa acctgtcgtg ccagcggatc cgcatctcaa ttagtcagca accatagtcc
     5581 cgcccctaac tccgcccatc ccgcccctaa ctccgcccag ttccgcccat tctccgcccc
     5641 atggctgact aatttttttt atttatgcag aggccgaggc cgcctcggcc tctgagctat
     5701 tccagaagta gtgaggaggc ttttttggag gcctaggctt ttgcaaaaag ctaacttgtt
     5761 tattgcagct tataatggtt acaaataaag caatagcatc acaaatttca caaataaagc
     5821 atttttttca ctgcattcta gttgtggttt gtccaaactc atcaatgtat cttatcatgt
     5881 ctggatccgc tgcattaatg aatcggccaa cgcgcgggga gaggcggttt gcgtattggg
     5941 cgctcttccg cttcctcgct cactgactcg ctgcgctcgg tcgttcggct gcggcgagcg
     6001 gtatcagctc actcaaaggc ggtaatacgg ttatccacag aatcagggga taacgcagga
     6061 aagaacatgt gagcaaaagg ccagcaaaag gccaggaacc gtaaaaaggc cgcgttgctg
     6121 gcgtttttcc ataggctccg cccccctgac gagcatcaca aaaatcgacg ctcaagtcag
     6181 aggtggcgaa acccgacagg actataaaga taccaggcgt ttccccctgg aagctccctc
     6241 gtgcgctctc ctgttccgac cctgccgctt accggatacc tgtccgcctt tctcccttcg
     6301 ggaagcgtgg cgctttctca tagctcacgc tgtaggtatc tcagttcggt gtaggtcgtt
     6361 cgctccaagc tgggctgtgt gcacgaaccc cccgttcagc ccgaccgctg cgccttatcc
     6421 ggtaactatc gtcttgagtc caacccggta agacacgact tatcgccact ggcagcagcc
     6481 actggtaaca ggattagcag agcgaggtat gtaggcggtg ctacagagtt cttgaagtgg
     6541 tggcctaact acggctacac tagaagaaca gtatttggta tctgcgctct gctgaagcca
     6601 gttaccttcg gaaaaagagt tggtagctct tgatccggca aacaaaccac cgctggtagc
     6661 ggtggttttt ttgtttgcaa gcagcagatt acgcgcagaa aaaaaggatc tcaagaagat
     6721 cctttgatct tttctacggg gtctgacgct cagtggaacg aaaactcacg ttaagggatt
     6781 ttggtcatga gattatcaaa aaggatcttc acctagatcc ttttaaatta aaaatgaagt
     6841 tttaaatcaa tctaaagtat atatgagtaa acttggtctg acagttacca atgcttaatc
     6901 agtgaggcac ctatctcagc gatctgtcta tttcgttcat ccatagttgc ctgactcccc
     6961 gtcgtgtaga taactacgat acgggagggc ttaccatctg gccccagtgc tgcaatgata
     7021 ccgcgagacc cacgctcacc ggctccagat ttatcagcaa taaaccagcc agccggaagg
     7081 gccgagcgca gaagtggtcc tgcaacttta tccgcctcca tccagtctat taattgttgc
     7141 cgggaagcta gagtaagtag ttcgccagtt aatagtttgc gcaacgttgt tgccattgct
     7201 acaggcatcg tggtgtcacg ctcgtcgttt ggtatggctt cattcagctc cggttcccaa
     7261 cgatcaaggc gagttacatg atcccccatg ttgtgcaaaa aagcggttag ctccttcggt
     7321 cctccgatcg ttgtcagaag taagttggcc gcagtgttat cactcatggt tatggcagca
     7381 ctgcataatt ctcttactgt catgccatcc gtaagatgct tttctgtgac tggtgagtac
     7441 tcaaccaagt cattctgaga atagtgtatg cggcgaccga gttgctcttg cccggcgtca
     7501 atacgggata ataccgcgcc acatagcaga actttaaaag tgctcatcat tggaaaacgt
     7561 tcttcggggc gaaaactctc aaggatctta ccgctgttga gatccagttc gatgtaaccc
     7621 actcgtgcac ccaactgatc ttcagcatct tttactttca ccagcgtttc tgggtgagca
     7681 aaaacaggaa ggcaaaatgc cgcaaaaaag ggaataaggg cgacacggaa atgttgaata
     7741 ctcatactct tcctttttca atattattga agcatttatc agggttattg tctcatgagc
     7801 ggatacatat ttgaatgtat ttagaaaaat aaacaaatag gggttccgcg cacatttccc
     7861 cgaaaagtgc cacctg
//
