LOCUS       sequence-393402-        6784 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  To study u-crystallin protein interactors, control plasmid.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6784)
  AUTHORS   Ollivier M, Soto JS, Linker KE, Moye SL, Jami-Alahmadi Y, Jones AE, 
            Divakaruni AS, Kawaguchi R, Wohlschlegel JA, Khakh BS
  TITLE     Crym-positive striatal astrocytes gate perseverative behaviour.
  JOURNAL   Nature. 2024 Mar;627(8003):358-366. doi: 10.1038/s41586-024-07138-0.
            Epub 2024 Feb 28.
  PUBMED    38418885
REFERENCE   2  (bases 1 to 6784)
  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: 
            "10.1038/s41586-024-07138-0"; journalName: "Nature"; date: 
            "2024-03"; volume: "627"; issue: "8003"; pages: "358-366"
COMMENT     Sequence Label: GfaABC1D-Crym-eGFP
FEATURES             Location/Qualifiers
     source          1..6784
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     protein_bind    177..198
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(213..218,219..236,237..243)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    251..267
                     /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)."
     primer_bind     256..278
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     275..291
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     275..291
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     repeat_region   329..458
                     /label=AAV2 ITR
     intron          1309..1441
                     /label=chimeric intron
                     /note="chimera between introns from human beta-globin and 
                     immunoglobulin heavy chain genes"
     primer_bind     1424..1443
                     /label=pMT2-F
                     /note="Synthetic intron, forward primer"
     promoter        1486..1504
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     regulatory      2482..2491
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(2488..2490,2491..2493,2494..3207)
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             join(2488..2490,2491..2493,2494..3204)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(2533..2554)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(2794..2813)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     3141..3162
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     misc_feature    3242..3830
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(3295..3315)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(3713..3724)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     polyA_signal    3879..4000
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3890..3909)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     3944..3963
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     repeat_region   4162..4291
                     /label=AAV2 ITR
     primer_bind     complement(4310..4327)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(4310..4326)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4319..4341)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      4468..4923
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4555..4574)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4765..4786
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        4949..5053
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5054..5122,5123..5914)
                     /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"
     rep_origin      6085..6673
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6574..6593
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 gagtgagctg ataccgctcg ccgcagccga acgaccgagc gcagcgagtc agtgagcgag
       61 gaagcggaag agcgcccaat acgcaaaccg cctctccccg cgcgttggcc gattcattaa
      121 tgcagctggc acgacaggtt tcccgactgg aaagcgggca gtgagcgcaa cgcaattaat
      181 gtgagttagc tcactcatta ggcaccccag gctttacact ttatgcttcc ggctcgtatg
      241 ttgtgtggaa ttgtgagcgg ataacaattt cacacaggaa acagctatga ccatgattac
      301 gccagattta attaaggcct taattaggct gcgcgctcgc tcgctcactg aggccgcccg
      361 ggcaaagccc gggcgtcggg cgacctttgg tcgcccggcc tcagtgagcg agcgagcgcg
      421 cagagaggga gtggccaact ccatcactag gggttccttg tagttaatga ttaacccgcc
      481 atgctactta tctacgtagc catgctctag gaagatctaa catatcctgg tgtggagtag
      541 gggacgctgc tctgacagag gctcgggggc ctgagctggc tctgtgagct ggggaggagg
      601 cagacagcca ggccttgtct gcaagcagac ctggcagcat tgggctggcc gccccccagg
      661 gcctcctctt catgcccagt gaatgactca ccttggcaca gacacaatgt tcggggtggg
      721 cacagtgcct gcttcccgcc gcaccccagc ccccctcaaa tgccttccga gaagcccatt
      781 gagcaggggg cttgcattgc accccagcct gacagcctgg catcttggga taaaagcagc
      841 acagccccct aggggctgcc cttgctgtgt ggcgccaccg gcggtggaga acaaggctct
      901 attcagcctg tgcccaggaa aggggatcag gggatgccca ggcatggaca gtgggtggca
      961 gggggggaga ggagggctgt ctgcttccca gaagtccaag gacacaaatg ggtgagggga
     1021 gagctctccc catagctggg ctgcggccca accccacccc ctcaggctat gccagggggt
     1081 gttgccaggg gcacccgggc atcgccagtc tagcccactc cttcataaag ccctcgcatc
     1141 ccaggagcga gcagagccag agcaggttgg agaggagacg catcacctcc gctgctcgca
     1201 agctttattg cggtagttta tcacagttaa attgctaacg cagtcagtgc ttctgacaca
     1261 acagtctcga acttaagctg cagaagttgg tcgtgaggca ctgggcaggt aagtatcaag
     1321 gttacaagac aggtttaagg agaccaatag aaactgggct tgtcgagaca gagaagactc
     1381 ttgcgtttct gataggcacc tattggtctt actgacatcc actttgcctt tctctccaca
     1441 ggtgtccact cccagttcaa ttacagctct taaggctaga gtacttaata cgactcacta
     1501 taggctagcc tcgagctcaa gcttcgaatt atgaagcggg cgccagcgtt cctgagcgca
     1561 gaggaggtgc aggatcacct tcgcagctcc agccttctca tcccacccct ggaggccgca
     1621 ctggccaact tctccaaagg tcccgacgga ggggtcatgc agccagtgcg caccgtggtg
     1681 cctgtagcca agcaccgagg cttcctggga gtcatgcctg cctacagtgc tgctgaggat
     1741 gcgctcacca ccaagttagt caccttctat gagggccaca gcaacacagc ggtcccctcc
     1801 catcaggcat cggtgcttct ctttgatccc agcaatggct ccctgctggc ggtcatggat
     1861 ggaaatgtca taactgcaaa gagaacagca gcggtgtctg ccattgccac aaagctgttg
     1921 aagcccccag gcagtgatgt gctgtgcatc cttggagcgg gggtccaggc gtacagtcac
     1981 tatgagatct tcacagagca gttctccttc aaggaggtga gaatgtggaa ccgcaccagg
     2041 gaaaatgctg agaagtttgc aagcacagtg caaggagatg ttcgggtctg ttcatcagtg
     2101 caggaggctg tgacaggtgc tgatgtcatc atcacagtca ccatggcaac agagcccatt
     2161 ttatttggtg aatgggtaaa gccgggggct cacatcaatg ctgttggagc cagcaggcct
     2221 gactggcgag aactggatga cgagctcatg aggcaagcgg tgctgtatgt ggactcccgg
     2281 gaggctgccc tgaaggagtc aggagacgtt ctgttgtcag gggctgacat ctttgctgag
     2341 cttggagaag tgatttcagg agcgaagcct gcacactgtg agaagaccac agtgttcaaa
     2401 tctttgggga tggcagtgga agacctggtt gcagccaaat tagtatatga ttcttggtca
     2461 tctggcaagg atccaccggt cgccaccatg gtgagcaagg gagaggagct gttcaccggg
     2521 gtggtgccca tcctggtcga gctggacggc gacgtaaacg gccacaagtt cagcgtgtcc
     2581 ggcgagggcg agggcgatgc cacctacggc aagctgaccc tgaagttcat ctgcaccacc
     2641 ggcaagctgc ccgtgccctg gcccaccctc gtgaccaccc tgacctacgg cgtgcagtgc
     2701 ttcagccgct accccgacca catgaagcag cacgacttct tcaagtccgc catgcccgaa
     2761 ggctacgtcc aggagcgcac catcttcttc aaggacgacg gcaactacaa gacccgcgcc
     2821 gaggtgaagt tcgagggcga caccctggtg aaccgcatcg agctgaaggg catcgacttc
     2881 aaggaggacg gcaacatcct ggggcacaag ctggagtaca actacaacag ccacaacgtc
     2941 tatatcatgg ccgacaagca gaagaacggc atcaaggtga acttcaagat ccgccacaac
     3001 atcgaggacg gcagcgtgca gctcgccgac cactaccagc agaacacccc catcggcgac
     3061 ggccccgtgc tgctgcccga caaccactac ctgagcaccc agtccgccct gagcaaagac
     3121 cccaacgaga agcgcgatca catggtcctg ctggagttcg tgaccgccgc cgggatcact
     3181 ctcggcatgg acgagctgta caagtaaagc ggccgcgacc ggtatcatca agcttatcga
     3241 taatcaacct ctggattaca aaatttgtga aagattgact ggtattctta actatgttgc
     3301 tccttttacg ctatgtggat acgctgcttt aatgcctttg tatcatgcta ttgcttcccg
     3361 tatggctttc attttctcct ccttgtataa atcctggttg ctgtctcttt atgaggagtt
     3421 gtggcccgtt gtcaggcaac gtggcgtggt gtgcactgtg tttgctgacg caacccccac
     3481 tggttggggc attgccacca cctgtcagct cctttccggg actttcgctt tccccctccc
     3541 tattgccacg gcggaactca tcgccgcctg ccttgcccgc tgctggacag gggctcggct
     3601 gttgggcact gacaattccg tggtgttgtc ggggaaatca tcgtcctttc cttggctgct
     3661 cgcctatgtt gccacctgga ttctgcgcgg gacgtccttc tgctacgtcc cttcggccct
     3721 caatccagcg gaccttcctt cccgcggcct gctgccggct ctgcggcctc ttccgcgtct
     3781 tcgccttcgc cctcagacga gtcggatctc cctttgggcc gcctccccgc atcgatacct
     3841 ctagagtcga cccgggcggc cgcttcgagc agacatgata agatacattg atgagtttgg
     3901 acaaaccaca actagaatgc agtgaaaaaa atgctttatt tgtgaaattt gtgatgctat
     3961 tgctttattt gtaaccatta taagctgcaa taaacaagtt aacaacaaca attgcattca
     4021 ttttatgttt caggttcagg gggagatgtg ggaggttttt taaagcaagt aaaacctcta
     4081 caaatgtggt aaaatcgata aggatcttcc tagagcatgg ctacgtagat aagtagcatg
     4141 gcgggttaat cattaactac aaggaacccc tagtgatgga gttggccact ccctctctgc
     4201 gcgctcgctc gctcactgag gccgggcgac caaaggtcgc ccgacgcccg ggctttgccc
     4261 gggcggcctc agtgagcgag cgagcgcgca gccttaatta acctaattca ctggccgtcg
     4321 ttttacaacg tcgtgactgg gaaaaccctg gcgttaccca acttaatcgc cttgcagcac
     4381 atcccccttt cgccagctgg cgtaatagcg aagaggcccg caccgatcgc ccttcccaac
     4441 agttgcgcag cctgaatggc gaatgggacg cgccctgtag cggcgcatta agcgcggcgg
     4501 gtgtggtggt tacgcgcagc gtgaccgcta cacttgccag cgccctagcg cccgctcctt
     4561 tcgctttctt cccttccttt ctcgccacgt tcgccggctt tccccgtcaa gctctaaatc
     4621 gggggctccc tttagggttc cgatttagtg ctttacggca cctcgacccc aaaaaacttg
     4681 attagggtga tggttcacgt agtgggccat cgccctgata gacggttttt cgccctttga
     4741 cgttggagtc cacgttcttt aatagtggac tcttgttcca aactggaaca acactcaacc
     4801 ctatctcggt ctattctttt gatttataag ggattttgcc gatttcggcc tattggttaa
     4861 aaaatgagct gatttaacaa aaatttaacg cgaattttaa caaaatatta acgcttacaa
     4921 tttaggtggc acttttcggg gaaatgtgcg cggaacccct atttgtttat ttttctaaat
     4981 acattcaaat atgtatccgc tcatgagaca ataaccctga taaatgcttc aataatattg
     5041 aaaaaggaag agtatgagta ttcaacattt ccgtgtcgcc cttattccct tttttgcggc
     5101 attttgcctt cctgtttttg ctcacccaga aacgctggtg aaagtaaaag atgctgaaga
     5161 tcagttgggt gcacgagtgg gttacatcga actggatctc aacagcggta agatccttga
     5221 gagttttcgc cccgaagaac gttttccaat gatgagcact tttaaagttc tgctatgtgg
     5281 cgcggtatta tcccgtattg acgccgggca agagcaactc ggtcgccgca tacactattc
     5341 tcagaatgac ttggttgagt actcaccagt cacagaaaag catcttacgg atggcatgac
     5401 agtaagagaa ttatgcagtg ctgccataac catgagtgat aacactgcgg ccaacttact
     5461 tctgacaacg atcggaggac cgaaggagct aaccgctttt ttgcacaaca tgggggatca
     5521 tgtaactcgc cttgatcgtt gggaaccgga gctgaatgaa gccataccaa acgacgagcg
     5581 tgacaccacg atgcctgtag caatggcaac aacgttgcgc aaactattaa ctggcgaact
     5641 acttactcta gcttcccggc aacaattaat agactggatg gaggcggata aagttgcagg
     5701 accacttctg cgctcggccc ttccggctgg ctggtttatt gctgataaat ctggagccgg
     5761 tgagcgtggg tctcgcggta tcattgcagc actggggcca gatggtaagc cctcccgtat
     5821 cgtagttatc tacacgacgg ggagtcaggc aactatggat gaacgaaata gacagatcgc
     5881 tgagataggt gcctcactga ttaagcattg gtaactgtca gaccaagttt actcatatat
     5941 actttagatt gatttaaaac ttcattttta atttaaaagg atctaggtga agatcctttt
     6001 tgataatctc atgaccaaaa tcccttaacg tgagttttcg ttccactgag cgtcagaccc
     6061 cgtagaaaag atcaaaggat cttcttgaga tccttttttt ctgcgcgtaa tctgctgctt
     6121 gcaaacaaaa aaaccaccgc taccagcggt ggtttgtttg ccggatcaag agctaccaac
     6181 tctttttccg aaggtaactg gcttcagcag agcgcagata ccaaatactg ttcttctagt
     6241 gtagccgtag ttaggccacc acttcaagaa ctctgtagca ccgcctacat acctcgctct
     6301 gctaatcctg ttaccagtgg ctgctgccag tggcgataag tcgtgtctta ccgggttgga
     6361 ctcaagacga tagttaccgg ataaggcgca gcggtcgggc tgaacggggg gttcgtgcac
     6421 acagcccagc ttggagcgaa cgacctacac cgaactgaga tacctacagc gtgagctatg
     6481 agaaagcgcc acgcttcccg aagggagaaa ggcggacagg tatccggtaa gcggcagggt
     6541 cggaacagga gagcgcacga gggagcttcc agggggaaac gcctggtatc tttatagtcc
     6601 tgtcgggttt cgccacctct gacttgagcg tcgatttttg tgatgctcgt caggggggcg
     6661 gagcctatgg aaaaacgcca gcaacgcggc ctttttacgg ttcctggcct tttgctggcc
     6721 ttttgctcac atgttctttc ctgcgttatc ccctgattct gtggataacc gtattaccgc
     6781 cttt
//
