LOCUS       sequence-127688-        6953 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  mammalian expression of rat GAT with N terminal YFP fusion.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6953)
  AUTHORS   Scholze P, Freissmuth M, Sitte HH
  TITLE     Mutations within an intramembrane leucine heptad repeat disrupt 
            oligomer formation of the rat GABA transporter 1.
  JOURNAL   J Biol Chem. 2002 Nov 15;277(46):43682-90. Epub 2002 Sep 9.
  PUBMED    12223478
REFERENCE   2  (bases 1 to 6953)
  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: "J Biol 
            Chem."; date: "2002-11-15"; volume: "277(46)"; pages: "43682-90. 
            Epub 2002 Sep 9"
COMMENT     Sequence Label: pEYFP-C1-rGAT
FEATURES             Location/Qualifiers
     source          1..6953
                     /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 YFP"
                     /label=EYFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTFGYGLQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSYQSALSKDPNEKRDHMVLL
                     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    3403..3537
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     polyA_signal    3741..3862
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     rep_origin      complement(3869..4324)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4006..4027)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     4218..4237
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        4351..4455
                     /gene="bla"
                     /label=AmpR promoter
     promoter        4457..4814
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        4457..4648
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(4480..4500)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      4665..4800
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             4849..5643
                     /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             4849..5643
                     /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(4903..4922)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     5513..5532
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(5831..5850)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    5875..5922
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      6251..6839
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6740..6759
                     /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 caccttcggc tacggcctgc agtgcttcgc 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 ctaccagtcc gccctgagca aagaccccaa cgagaagcgc
     1261 gatcacatgg tcctgctgga gttcgtgacc gccgccggga tcactctcgg catggacgag
     1321 ctgtacaagt ccggactcag atctcgagct caagcttcga attctgcagt cgagtggact
     1381 ggagctgaca tcgcacgcct gcccgccagg atccctgctg ccaagttcgt gctccgagac
     1441 atggcgactg acaacagcaa ggtggctgat gggcagatct ctaccgaggt cagcgaggct
     1501 cctgtggcca gtgacaagcc caaaaccctt gtagtcaagg tgcagaagaa ggctggagac
     1561 ctccctgacc gggacacatg gaagggacgc ttcgacttcc tcatgtcctg tgtgggctac
     1621 gccatcggcc tgggcaatgt atggaggttc ccttatctct gcggaaagaa cggtggtggg
     1681 gccttcctaa ttccatattt cctgacgctc atctttgcgg gtgttcctct cttccttttg
     1741 gagtgctccc taggccagta cacctccatt gggggcctgg gcgtatggaa gctggctccc
     1801 atgttcaagg gtgtgggcct cgcggcagct gtgctgtcct tctggctgaa catttattac
     1861 attgtcatca tctcctgggc catctactac ctgtacaact ccttcaccac gactctgccg
     1921 tggaaacagt gtgacaaccc ctggaacact gaccgctgct tctccaacta tagcctggtc
     1981 aacaccacca acatgaccag tgccgtggtg gaattctggg agcgcaacat gcaccaaatg
     2041 acagatgggc tagacaagcc aggacagatc cgctggccgc tagccatcac actggccatt
     2101 gcctgggtgc ttgtgtattt ctgcatctgg aagggtgttg gatggactgg aaaggtggtc
     2161 tacttctcag ccacataccc ctacatcatg ctcatcatcc tgttcttccg tggagtgaca
     2221 cttcctgggg ccaaggaggg gatcctcttc tacatcacgc ccaacttccg aaagctgtct
     2281 gattctgagg tgtggcttga cgccgccacc cagatcttct tctcctacgg actgggcctg
     2341 gggtccctga ttgctctggg aagctacaac tctttccaca acaatgtgta cagggactcc
     2401 atcattgttt gctgcatcaa ctcctgcacc agcatgtttg ccggcttcgt catcttctcc
     2461 atcgtgggct tcatggctca tgtcaccaag aggtccatag ccgatgtggc ggcctcaggc
     2521 cctggactgg cattcttggc ataccccgag gctgtgacac agctacccat ctctcccctc
     2581 tgggctatcc tcttcttctc catgctgctg atgctgggca ttgacagcca gttctgtacc
     2641 gtggagggct tcatcactgc cctggtggat gagtacccca ggcttctccg caatcgccgt
     2701 gaactcttca ttgctgccgt gtgcatcgtg tcctacctga ttggcctctc taacatcacc
     2761 cagggtggca tttatgtctt caaactgttc gattattact ctgccagtgg catgagcctg
     2821 ctgttcctgg ttttctttga gtgtgtctcc atttcctggt tttatggtgt caaccggttc
     2881 tatgacaaca tccaggagat ggttggctcc aggccctgca tctggtggaa gctgtgctgg
     2941 tcctttttca cacccatcat tgtggcgggc gtgtttctct tcagtgctgt gcagatgaca
     3001 ccactcacca tgggaagcta cgttttcccc aagtggggcc agggcgtggg ctggctcatg
     3061 gctctgtcct ccatggtgct catccccggg tacatggctt acatgttcct caccctaaag
     3121 ggctctctga agcagcgtct ccaggtcatg attcagccca gtgaagatat cgtgcgccct
     3181 gagaatggtc ctgagcagcc gcaggctggc agctcagcca gcaaggaggc ctacatctag
     3241 gggtacagcc ccctcatcac ccctacactg gcaccctgga ctggctgtac ccacacccct
     3301 tgaagactga agatactctc tgtctccaac tacctcaagg ggcgggtcca gacaccatca
     3361 ccatgcagag aggggagatg ggggacagtc tgaccctagg tgaacttgtt tattgcagct
     3421 tataatggtt acaaataaag caatagcatc acaaatttca caaataaagc atttttttca
     3481 ctgcattcta gttgtggttt gtccaaactc atcaatgtat cttatcatgt ctggatcccc
     3541 gggaattcgg cgcagcacca tggcctgaaa taacctctga aagaggaact tggttaggta
     3601 ccgcgggccc gggatccacc ggatctagat aactgatcat aatcagccat accacatttg
     3661 tagaggtttt acttgcttta aaaaacctcc cacacctccc cctgaacctg aaacataaaa
     3721 tgaatgcaat tgttgttgtt aacttgttta ttgcagctta taatggttac aaataaagca
     3781 atagcatcac aaatttcaca aataaagcat ttttttcact gcattctagt tgtggtttgt
     3841 ccaaactcat caatgtatct taaggcgtaa attgtaagcg ttaatatttt gttaaaattc
     3901 gcgttaaatt tttgttaaat cagctcattt tttaaccaat aggccgaaat cggcaaaatc
     3961 ccttataaat caaaagaata gaccgagata gggttgagtg ttgttccagt ttggaacaag
     4021 agtccactat taaagaacgt ggactccaac gtcaaagggc gaaaaaccgt ctatcagggc
     4081 gatggcccac tacgtgaacc atcaccctaa tcaagttttt tggggtcgag gtgccgtaaa
     4141 gcactaaatc ggaaccctaa agggagcccc cgatttagag cttgacgggg aaagccggcg
     4201 aacgtggcga gaaaggaagg gaagaaagcg aaaggagcgg gcgctagggc gctggcaagt
     4261 gtagcggtca cgctgcgcgt aaccaccaca cccgccgcgc ttaatgcgcc gctacagggc
     4321 gcgtcaggtg gcacttttcg gggaaatgtg cgcggaaccc ctatttgttt atttttctaa
     4381 atacattcaa atatgtatcc gctcatgaga caataaccct gataaatgct tcaataatat
     4441 tgaaaaagga agagtcctga ggcggaaaga accagctgtg gaatgtgtgt cagttagggt
     4501 gtggaaagtc cccaggctcc ccagcaggca gaagtatgca aagcatgcat ctcaattagt
     4561 cagcaaccag gtgtggaaag tccccaggct ccccagcagg cagaagtatg caaagcatgc
     4621 atctcaatta gtcagcaacc atagtcccgc ccctaactcc gcccatcccg cccctaactc
     4681 cgcccagttc cgcccattct ccgccccatg gctgactaat tttttttatt tatgcagagg
     4741 ccgaggccgc ctcggcctct gagctattcc agaagtagtg aggaggcttt tttggaggcc
     4801 taggcttttg caaagatcga tcaagagaca ggatgaggat cgtttcgcat gattgaacaa
     4861 gatggattgc acgcaggttc tccggccgct tgggtggaga ggctattcgg ctatgactgg
     4921 gcacaacaga caatcggctg ctctgatgcc gccgtgttcc ggctgtcagc gcaggggcgc
     4981 ccggttcttt ttgtcaagac cgacctgtcc ggtgccctga atgaactgca agacgaggca
     5041 gcgcggctat cgtggctggc cacgacgggc gttccttgcg cagctgtgct cgacgttgtc
     5101 actgaagcgg gaagggactg gctgctattg ggcgaagtgc cggggcagga tctcctgtca
     5161 tctcaccttg ctcctgccga gaaagtatcc atcatggctg atgcaatgcg gcggctgcat
     5221 acgcttgatc cggctacctg cccattcgac caccaagcga aacatcgcat cgagcgagca
     5281 cgtactcgga tggaagccgg tcttgtcgat caggatgatc tggacgaaga gcatcagggg
     5341 ctcgcgccag ccgaactgtt cgccaggctc aaggcgagca tgcccgacgg cgaggatctc
     5401 gtcgtgaccc atggcgatgc ctgcttgccg aatatcatgg tggaaaatgg ccgcttttct
     5461 ggattcatcg actgtggccg gctgggtgtg gcggaccgct atcaggacat agcgttggct
     5521 acccgtgata ttgctgaaga gcttggcggc gaatgggctg accgcttcct cgtgctttac
     5581 ggtatcgccg ctcccgattc gcagcgcatc gccttctatc gccttcttga cgagttcttc
     5641 tgagcgggac tctggggttc gaaatgaccg accaagcgac gcccaacctg ccatcacgag
     5701 atttcgattc caccgccgcc ttctatgaaa ggttgggctt cggaatcgtt ttccgggacg
     5761 ccggctggat gatcctccag cgcggggatc tcatgctgga gttcttcgcc caccctaggg
     5821 ggaggctaac tgaaacacgg aaggagacaa taccggaagg aacccgcgct atgacggcaa
     5881 taaaaagaca gaataaaacg cacggtgttg ggtcgtttgt tcataaacgc ggggttcggt
     5941 cccagggctg gcactctgtc gataccccac cgagacccca ttggggccaa tacgcccgcg
     6001 tttcttcctt ttccccaccc caccccccaa gttcgggtga aggcccaggg ctcgcagcca
     6061 acgtcggggc ggcaggccct gccatagcct caggttactc atatatactt tagattgatt
     6121 taaaacttca tttttaattt aaaaggatct aggtgaagat cctttttgat aatctcatga
     6181 ccaaaatccc ttaacgtgag ttttcgttcc actgagcgtc agaccccgta gaaaagatca
     6241 aaggatcttc ttgagatcct ttttttctgc gcgtaatctg ctgcttgcaa acaaaaaaac
     6301 caccgctacc agcggtggtt tgtttgccgg atcaagagct accaactctt tttccgaagg
     6361 taactggctt cagcagagcg cagataccaa atactgtcct tctagtgtag ccgtagttag
     6421 gccaccactt caagaactct gtagcaccgc ctacatacct cgctctgcta atcctgttac
     6481 cagtggctgc tgccagtggc gataagtcgt gtcttaccgg gttggactca agacgatagt
     6541 taccggataa ggcgcagcgg tcgggctgaa cggggggttc gtgcacacag cccagcttgg
     6601 agcgaacgac ctacaccgaa ctgagatacc tacagcgtga gctatgagaa agcgccacgc
     6661 ttcccgaagg gagaaaggcg gacaggtatc cggtaagcgg cagggtcgga acaggagagc
     6721 gcacgaggga gcttccaggg ggaaacgcct ggtatcttta tagtcctgtc gggtttcgcc
     6781 acctctgact tgagcgtcga tttttgtgat gctcgtcagg ggggcggagc ctatggaaaa
     6841 acgccagcaa cgcggccttt ttacggttcc tggccttttg ctggcctttt gctcacatgt
     6901 tctttcctgc gttatcccct gattctgtgg ataaccgtat taccgccatg cat
//
