LOCUS       sequence-439383-        8919 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses His-tagged NCAN-Full length to produce recombinant 
            protein.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8919)
  AUTHORS   Irala D, Wang S, Sakers K, Nagendren L, Ulloa Severino FP, Bindu DS,
            Savage JT, Eroglu C
  TITLE     Astrocyte-secreted neurocan controls inhibitory synapse formation 
            and function.
  JOURNAL   Neuron. 2024 Mar 27:S0896-6273(24)00161-2. doi: 
            10.1016/j.neuron.2024.03.007.
  PUBMED    38574730
REFERENCE   2  (bases 1 to 8919)
  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: "Neuron. 
            2024 Mar 27:S0896-6273(24)00161-2. doi: 
            10.1016/j.neuron.2024.03.007."
COMMENT     Sequence Label: pAPtag5-IgK-NCAN-Full length
FEATURES             Location/Qualifiers
     source          1..8919
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(48..67)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        239..618
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        619..822
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     773..793
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     867..886
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        867..885
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     regulatory      903..912
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     sig_peptide     909..968
                     /codon_start=1
                     /product="leader sequence from mouse immunoglobulin kappa 
                     light chain"
                     /label=Ig-kappa leader
                     /translation="METDTLLLWVLLLWVPGSTG"
     CDS             4842..4871
                     /codon_start=1
                     /product="Myc (human c-Myc proto-oncogene) epitope tag"
                     /label=Myc
                     /translation="EQKLISEEDL"
     CDS             4887..4904
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(4927..4944)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    4933..5157
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      5203..5631
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(5290..5309)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     5500..5521
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(5640..5660)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        5645..5974
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      5825..5960
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     5887..5906
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     promoter        6022..6069
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             6088..6462
                     /codon_start=1
                     /gene="Sh ble from Streptoalloteichus hindustanus"
                     /product="antibiotic-binding protein"
                     /label=BleoR
                     /note="confers resistance to bleomycin, phleomycin, and 
                     Zeocin(TM)"
                     /translation="MAKLTSAVPVLTARDVAGAVEFWTDRLGFSRDFVEDDFAGVVRDD
                     VTLFISAVQDQVVPDNTLAWVWVRGLDELYAEWSEVVSTNFRDASGPAMTEIGEQPWGR
                     EFALRDPAGNCVHFVAEEQD"
     polyA_signal    6592..6713
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(6629..6648)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     6683..6702
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(6762..6778)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(6762..6778)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(6775..6797)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    6786..6802
                     /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(6810..6816,6817..6834,6835..6840))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    6855..6876
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(6993..7010)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(7164..7752)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(7244..7263)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(7923..8714,8715..8783))
                     /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     8546..8565
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(8784..8888)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 gacggatcgg gagatcgatc tcccgatccc ctatggtgca ctctcagtac aatctgctct
       61 gatgccgcat agttaagcca gtatctgctc cctgcttgtg tgttggaggt cgctgagtag
      121 tgcgcgagca aaatttaagc tacaacaagg caaggcttga ccgacaattg catgaagaat
      181 ctgcttaggg ttaggcgttt tgcgctgctt cgcgatgtac gggccagata tacgcgttga
      241 cattgattat tgactagtta ttaatagtaa tcaattacgg ggtcattagt tcatagccca
      301 tatatggagt tccgcgttac ataacttacg gtaaatggcc cgcctggctg accgcccaac
      361 gacccccgcc cattgacgtc aataatgacg tatgttccca tagtaacgcc aatagggact
      421 ttccattgac gtcaatgggt ggagtattta cggtaaactg cccacttggc agtacatcaa
      481 gtgtatcata tgccaagtac gccccctatt gacgtcaatg acggtaaatg gcccgcctgg
      541 cattatgccc agtacatgac cttatgggac tttcctactt ggcagtacat ctacgtatta
      601 gtcatcgcta ttaccatggt gatgcggttt tggcagtaca tcaatgggcg tggatagcgg
      661 tttgactcac ggggatttcc aagtctccac cccattgacg tcaatgggag tttgttttgg
      721 caccaaaatc aacgggactt tccaaaatgt cgtaacaact ccgccccatt gacgcaaatg
      781 ggcggtaggc gtgtacggtg ggaggtctat ataagcagag ctctctggct aactagagaa
      841 cccactgctt actggcttat cgaaattaat acgactcact atagggagac ccaagctggc
      901 tagccaccat ggagacagac acactcctgc tatgggtact gctgctctgg gttccaggtt
      961 ccactggtga cgcggcccag atgggggccc cgtttgtctg ggccttgggc cttttgatgc
     1021 tgcagatgct gctctttgtg gctggggaac agggcacaca ggatatcacc gatgccagcg
     1081 aaagggggct ccacatgcag aagctggggt ctgggtcagt gcaggctgcg ctggcggagc
     1141 tggtggccct gccctgtctc tttaccctgc agccacggcc aagcgcagcc cgagatgccc
     1201 ctcggataaa gtggaccaag gtgcggactg cgtcgggcca gcgacaggac ttgcccatcc
     1261 tggtggccaa ggacaatgtc gtgagggtgg ccaaaagctg gcagggacga gtgtcactgc
     1321 cttcctaccc ccggcgccga gccaacgcca cgctacttct ggggccactg agggccagtg
     1381 actctgggct gtaccgctgc caggtggtga ggggcatcga ggatgagcag gacctggtgc
     1441 ccttggaggt gacaggtgtt gtgttccact accgatcagc ccgggaccgc tatgcactga
     1501 ccttcgctga ggcccaggag gcctgccgtc tcagctcagc catcattgca gcccctcggc
     1561 atctacaggc tgcctttgag gatggctttg acaactgtga tgctggctgg ctctctgacc
     1621 gcactgttcg gtatcctatc acccagtccc gtcctggttg ctatggcgac cgtagcagcc
     1681 ttccaggggt tcggagctat gggaggcgca acccacagga actctacgat gtgtattgct
     1741 ttgcccggga gctggggggc gaggtcttct acgtgggccc ggcccgccgc ctgacactgg
     1801 ccggcgcgcg tgcacagtgc cgccgccagg gtgccgcgct ggcctcggtg ggacagctgc
     1861 acctggcctg gcatgagggc ctggaccagt gcgacccggg ctggctggcc gacggcagcg
     1921 tgcgctaccc gatccagacg ccgcgccggc gctgcggggg cccagccccg ggcgtgcgca
     1981 ccgtctaccg cttcgctaac cggaccggct tcccctcacc cgccgagcgc ttcgacgcct
     2041 actgcttccg agctcatcac cccacgtcac aacatggaga cctagagacc ccatcctctg
     2101 gggatgaggg ggagattctg tcagcagagg ggcccccagt tagagaactg gagcccaccc
     2161 tggaggagga agaggtggtc acccctgact tccaggagcc tctggtgtcc agtggggaag
     2221 aagaaaccct gattttggag gagaagcagg agtctcaaca gaccctcagc cctacccctg
     2281 gggaccccat gctggcctca tggcccactg gggaagtgtg gctaagcacg gtggccccca
     2341 gccctagcga catgggggca ggcactgcag caagttcaca cacggaggtg gccccaactg
     2401 accctatgcc taggagaagg gggcgcttca aagggttgaa tgggcgctac ttccagcagc
     2461 aggaaccgga gccggggctg caagggggga tggaggccag cgcccagccc cccacctcag
     2521 aggctgcagt gaaccaaatg gagcctccgt tggccatggc agtcacagag atgttgggca
     2581 gtggccagag ccggagcccc tgggctgatc tgaccaatga ggtggatatg cctggagctg
     2641 gttctgctgg tggcaagagc tccccagagc cctggctgtg gccccctacc atggtcccac
     2701 ccagcatctc aggccacagc agggcccctg tcctggagct agagaaagcc gagggcccca
     2761 gtgccaggcc agccacccca gacctgtttt ggtccccctt ggaggccact gtctcagctc
     2821 ccagccctgc cccctgggag gcattccctg tggccacctc cccagatctc cctatgatgg
     2881 ccatgctgcg tggtcccaaa gagtggatgc taccacaccc cacccccatc tccaccgagg
     2941 ccaatagagt tgaggcacat ggtgaggcca ccgccacggc tccaccctcc cctgctgcag
     3001 agaccaaggt gtattccctg cctctctctt tgaccccaac aggacagggt ggagaggcca
     3061 tgcccacaac acctgagtcc cccagggcag acttcagaga aactggggag accagccctg
     3121 ctcaggtcaa caaagctgag cactccagct ccagcccatg gccttctgta aacaggaatg
     3181 tggctgtagg ttttgtcccc actgagactg ccactgagcc aacgggcctc aggggtatcc
     3241 cggggtctga gtctggggtc ttcgacacag cagaaagccc cacttctggc ttgcaggcca
     3301 ctgtagatga ggtgcaggac ccctggccct cagtgtacag caaagggctg gatgcaagtt
     3361 ccccatctgc ccccctgggg agccctggag tcttcttggt acccaaagtc accccaaatt
     3421 tggagccttg ggttgctaca gatgaaggac ccactgtgaa tcccatggat tccacagtca
     3481 cgccggcccc cagtgatgct agtggaattt gggaacctgg atcccaggtg tttgaagaag
     3541 ccgaaagcac caccttgagc cctcaggtgg ccctggatac aagcattgtg acgcccctca
     3601 cgaccctgga gcagggggac aaggttggag ttccagccat gtctacactg ggctcctcaa
     3661 gctcccaacc ccacccagag ccagaggatc aggtggagac ccagggaaca tcaggagctt
     3721 cagtgcctcc gcatcagagc agtcccctag ggaaaccggc tgttcctcct gggacaccga
     3781 ctgcagccag tgtgggcgag tctgcctcag tttcctcagg ggagcctacg gtaccgtggg
     3841 acccctccag caccctgctg cctgtcaccc tgggcataga ggacttcgaa ctggaggtcc
     3901 tggcagggag cccgggtgta gagagcttct gggaggaggt ggcaagtgga gaggagccag
     3961 ccctgccagg gacccctatg aatgcaggtg cggaggaggt gcactcagat ccctgtgaga
     4021 acaacccttg tcttcatgga gggacatgta atgccaatgg caccatgtat ggctgtagct
     4081 gtgatcaggg cttcgccggg gagaactgtg agattgacat tgatgactgc ctctgcagcc
     4141 cctgtgagaa tggaggcacc tgtattgatg aggtcaatgg ctttgtctgc ctttgcctcc
     4201 ccagctatgg gggcagcttt tgtgagaaag acaccgaggg ctgtgaccgc ggctggcata
     4261 agttccaggg ccactgttac cgctattttg cccaccggag ggcatgggaa gatgccgaga
     4321 aggactgccg ccgccgctcc ggccacctga ccagcgtcca ctcaccggag gaacacagct
     4381 tcattaatag ctttgggcat gaaaacacgt ggatcggcct gaacgacagg atcgtggaga
     4441 gagatttcca gtggacggac aacaccgggc tgcaatttga gaactggcga gagaaccagc
     4501 cggacaattt cttcgcgggt ggcgaggact gtgtggtgat ggtggcgcat gaaagcgggc
     4561 gctggaacga tgtcccctgc aactacaacc taccctatgt ctgcaagaag ggcacagtgc
     4621 tctgtggtcc ccctccggca gtggagaatg cctcactcat cggtgcccgc aaggccaagt
     4681 acaatgtcca tgccactgta aggtaccagt gcaatgaagg atttgcccag caccatgtgg
     4741 ccaccattcg atgccggagc aatggcaagt gggacaggcc ccaaattgtc tgcaccaaac
     4801 ccagacgttc acatcggatg cggcgatctc tagaagggcc cgaacaaaaa ctcatctcag
     4861 aagaggatct gaatagcgcc gtcgaccatc atcatcatca tcattgagtt taaacccgct
     4921 gatcagcctc gactgtgcct tctagttgcc agccatctgt tgtttgcccc tcccccgtgc
     4981 cttccttgac cctggaaggt gccactccca ctgtcctttc ctaataaaat gaggaaattg
     5041 catcgcattg tctgagtagg tgtcattcta ttctgggggg tggggtgggg caggacagca
     5101 agggggagga ttgggaagac aatagcaggc atgctgggga tgcggtgggc tctatggctt
     5161 ctgaggcgga aagaaccagc tggggctcta gggggtatcc ccacgcgccc tgtagcggcg
     5221 cattaagcgc ggcgggtgtg gtggttacgc gcagcgtgac cgctacactt gccagcgccc
     5281 tagcgcccgc tcctttcgct ttcttccctt cctttctcgc cacgttcgcc ggctttcccc
     5341 gtcaagctct aaatcggggg ctccctttag ggttccgatt tagtgcttta cggcacctcg
     5401 accccaaaaa acttgattag ggtgatggtt cacgtagtgg gccatcgccc tgatagacgg
     5461 tttttcgccc tttgacgttg gagtccacgt tctttaatag tggactcttg ttccaaactg
     5521 gaacaacact caaccctatc tcggtctatt cttttgattt ataagggatt ttgccgattt
     5581 cggcctattg gttaaaaaat gagctgattt aacaaaaatt taacgcgaat taattctgtg
     5641 gaatgtgtgt cagttagggt gtggaaagtc cccaggctcc ccagcaggca gaagtatgca
     5701 aagcatgcat ctcaattagt cagcaaccag gtgtggaaag tccccaggct ccccagcagg
     5761 cagaagtatg caaagcatgc atctcaatta gtcagcaacc atagtcccgc ccctaactcc
     5821 gcccatcccg cccctaactc cgcccagttc cgcccattct ccgccccatg gctgactaat
     5881 tttttttatt tatgcagagg ccgaggccgc ctctgcctct gagctattcc agaagtagtg
     5941 aggaggcttt tttggaggcc taggcttttg caaaaagctc ccgggagctt gtatatccat
     6001 tttcggatct gatcagcacg tgttgacaat taatcatcgg catagtatat cggcatagta
     6061 taatacgaca aggtgaggaa ctaaaccatg gccaagttga ccagtgccgt tccggtgctc
     6121 accgcgcgcg acgtcgccgg agcggtcgag ttctggaccg accggctcgg gttctcccgg
     6181 gacttcgtgg aggacgactt cgccggtgtg gtccgggacg acgtgaccct gttcatcagc
     6241 gcggtccagg accaggtggt gccggacaac accctggcct gggtgtgggt gcgcggcctg
     6301 gacgagctgt acgccgagtg gtcggaggtc gtgtccacga acttccggga cgcctccggg
     6361 ccggccatga ccgagatcgg cgagcagccg tgggggcggg agttcgccct gcgcgacccg
     6421 gccggcaact gcgtgcactt cgtggccgag gagcaggact gacacgtgct acgagatttc
     6481 gattccaccg ccgccttcta tgaaaggttg ggcttcggaa tcgttttccg ggacgccggc
     6541 tggatgatcc tccagcgcgg ggatctcatg ctggagttct tcgcccaccc caacttgttt
     6601 attgcagctt ataatggtta caaataaagc aatagcatca caaatttcac aaataaagca
     6661 tttttttcac tgcattctag ttgtggtttg tccaaactca tcaatgtatc ttatcatgtc
     6721 tgtataccgt cgacctctag ctagagcttg gcgtaatcat ggtcatagct gtttcctgtg
     6781 tgaaattgtt atccgctcac aattccacac aacatacgag ccggaagcat aaagtgtaaa
     6841 gcctggggtg cctaatgagt gagctaactc acattaattg cgttgcgctc actgcccgct
     6901 ttccagtcgg gaaacctgtc gtgccagctg cattaatgaa tcggccaacg cgcggggaga
     6961 ggcggtttgc gtattgggcg ctcttccgct tcctcgctca ctgactcgct gcgctcggtc
     7021 gttcggctgc ggcgagcggt atcagctcac tcaaaggcgg taatacggtt atccacagaa
     7081 tcaggggata acgcaggaaa gaacatgtga gcaaaaggcc agcaaaaggc caggaaccgt
     7141 aaaaaggccg cgttgctggc gtttttccat aggctccgcc cccctgacga gcatcacaaa
     7201 aatcgacgct caagtcagag gtggcgaaac ccgacaggac tataaagata ccaggcgttt
     7261 ccccctggaa gctccctcgt gcgctctcct gttccgaccc tgccgcttac cggatacctg
     7321 tccgcctttc tcccttcggg aagcgtggcg ctttctcata gctcacgctg taggtatctc
     7381 agttcggtgt aggtcgttcg ctccaagctg ggctgtgtgc acgaaccccc cgttcagccc
     7441 gaccgctgcg ccttatccgg taactatcgt cttgagtcca acccggtaag acacgactta
     7501 tcgccactgg cagcagccac tggtaacagg attagcagag cgaggtatgt aggcggtgct
     7561 acagagttct tgaagtggtg gcctaactac ggctacacta gaagaacagt atttggtatc
     7621 tgcgctctgc tgaagccagt taccttcgga aaaagagttg gtagctcttg atccggcaaa
     7681 caaaccaccg ctggtagcgg tggttttttt gtttgcaagc agcagattac gcgcagaaaa
     7741 aaaggatctc aagaagatcc tttgatcttt tctacggggt ctgacgctca gtggaacgaa
     7801 aactcacgtt aagggatttt ggtcatgaga ttatcaaaaa ggatcttcac ctagatcctt
     7861 ttaaattaaa aatgaagttt taaatcaatc taaagtatat atgagtaaac ttggtctgac
     7921 agttaccaat gcttaatcag tgaggcacct atctcagcga tctgtctatt tcgttcatcc
     7981 atagttgcct gactccccgt cgtgtagata actacgatac gggagggctt accatctggc
     8041 cccagtgctg caatgatacc gcgagaccca cgctcaccgg ctccagattt atcagcaata
     8101 aaccagccag ccggaagggc cgagcgcaga agtggtcctg caactttatc cgcctccatc
     8161 cagtctatta attgttgccg ggaagctaga gtaagtagtt cgccagttaa tagtttgcgc
     8221 aacgttgttg ccattgctac aggcatcgtg gtgtcacgct cgtcgtttgg tatggcttca
     8281 ttcagctccg gttcccaacg atcaaggcga gttacatgat cccccatgtt gtgcaaaaaa
     8341 gcggttagct ccttcggtcc tccgatcgtt gtcagaagta agttggccgc agtgttatca
     8401 ctcatggtta tggcagcact gcataattct cttactgtca tgccatccgt aagatgcttt
     8461 tctgtgactg gtgagtactc aaccaagtca ttctgagaat agtgtatgcg gcgaccgagt
     8521 tgctcttgcc cggcgtcaat acgggataat accgcgccac atagcagaac tttaaaagtg
     8581 ctcatcattg gaaaacgttc ttcggggcga aaactctcaa ggatcttacc gctgttgaga
     8641 tccagttcga tgtaacccac tcgtgcaccc aactgatctt cagcatcttt tactttcacc
     8701 agcgtttctg ggtgagcaaa aacaggaagg caaaatgccg caaaaaaggg aataagggcg
     8761 acacggaaat gttgaatact catactcttc ctttttcaat attattgaag catttatcag
     8821 ggttattgtc tcatgagcgg atacatattt gaatgtattt agaaaaataa acaaataggg
     8881 gttccgcgca catttccccg aaaagtgcca cctgacgtc
//
