LOCUS       sequence-439376-        5907 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses His-tagged NCAN-ELS to produce recombinant protein.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5907)
  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 5907)
  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 ELS
FEATURES             Location/Qualifiers
     source          1..5907
                     /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             1830..1859
                     /codon_start=1
                     /product="Myc (human c-Myc proto-oncogene) epitope tag"
                     /label=Myc
                     /translation="EQKLISEEDL"
     CDS             1875..1892
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(1915..1932)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    1921..2145
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      2191..2619
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2278..2297)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2488..2509
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(2628..2648)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        2633..2962
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      2813..2948
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     2875..2894
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     promoter        3010..3057
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             3076..3450
                     /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    3580..3701
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3617..3636)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3671..3690
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(3750..3766)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3750..3766)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3763..3785)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    3774..3790
                     /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(3798..3804,3805..3822,3823..3828))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3843..3864
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(3981..3998)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4152..4740)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4232..4251)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4911..5702,5703..5771))
                     /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     5534..5553
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5772..5876)
                     /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 atggaggtgc actcagatcc ctgtgagaac aacccttgtc
     1021 ttcatggagg gacatgtaat gccaatggca ccatgtatgg ctgtagctgt gatcagggct
     1081 tcgccgggga gaactgtgag attgacattg atgactgcct ctgcagcccc tgtgagaatg
     1141 gaggcacctg tattgatgag gtcaatggct ttgtctgcct ttgcctcccc agctatgggg
     1201 gcagcttttg tgagaaagac accgagggct gtgaccgcgg ctggcataag ttccagggcc
     1261 actgttaccg ctattttgcc caccggaggg catgggaaga tgccgagaag gactgccgcc
     1321 gccgctccgg ccacctgacc agcgtccact caccggagga acacagcttc attaatagct
     1381 ttgggcatga aaacacgtgg atcggcctga acgacaggat cgtggagaga gatttccagt
     1441 ggacggacaa caccgggctg caatttgaga actggcgaga gaaccagccg gacaatttct
     1501 tcgcgggtgg cgaggactgt gtggtgatgg tggcgcatga aagcgggcgc tggaacgatg
     1561 tcccctgcaa ctacaaccta ccctatgtct gcaagaaggg cacagtgctc tgtggtcccc
     1621 ctccggcagt ggagaatgcc tcactcatcg gtgcccgcaa ggccaagtac aatgtccatg
     1681 ccactgtaag gtaccagtgc aatgaaggat ttgcccagca ccatgtggcc accattcgat
     1741 gccggagcaa tggcaagtgg gacaggcccc aaattgtctg caccaaaccc agacgttcac
     1801 atcggatgcg gcgatctcta gaagggcccg aacaaaaact catctcagaa gaggatctga
     1861 atagcgccgt cgaccatcat catcatcatc attgagttta aacccgctga tcagcctcga
     1921 ctgtgccttc tagttgccag ccatctgttg tttgcccctc ccccgtgcct tccttgaccc
     1981 tggaaggtgc cactcccact gtcctttcct aataaaatga ggaaattgca tcgcattgtc
     2041 tgagtaggtg tcattctatt ctggggggtg gggtggggca ggacagcaag ggggaggatt
     2101 gggaagacaa tagcaggcat gctggggatg cggtgggctc tatggcttct gaggcggaaa
     2161 gaaccagctg gggctctagg gggtatcccc acgcgccctg tagcggcgca ttaagcgcgg
     2221 cgggtgtggt ggttacgcgc agcgtgaccg ctacacttgc cagcgcccta gcgcccgctc
     2281 ctttcgcttt cttcccttcc tttctcgcca cgttcgccgg ctttccccgt caagctctaa
     2341 atcgggggct ccctttaggg ttccgattta gtgctttacg gcacctcgac cccaaaaaac
     2401 ttgattaggg tgatggttca cgtagtgggc catcgccctg atagacggtt tttcgccctt
     2461 tgacgttgga gtccacgttc tttaatagtg gactcttgtt ccaaactgga acaacactca
     2521 accctatctc ggtctattct tttgatttat aagggatttt gccgatttcg gcctattggt
     2581 taaaaaatga gctgatttaa caaaaattta acgcgaatta attctgtgga atgtgtgtca
     2641 gttagggtgt ggaaagtccc caggctcccc agcaggcaga agtatgcaaa gcatgcatct
     2701 caattagtca gcaaccaggt gtggaaagtc cccaggctcc ccagcaggca gaagtatgca
     2761 aagcatgcat ctcaattagt cagcaaccat agtcccgccc ctaactccgc ccatcccgcc
     2821 cctaactccg cccagttccg cccattctcc gccccatggc tgactaattt tttttattta
     2881 tgcagaggcc gaggccgcct ctgcctctga gctattccag aagtagtgag gaggcttttt
     2941 tggaggccta ggcttttgca aaaagctccc gggagcttgt atatccattt tcggatctga
     3001 tcagcacgtg ttgacaatta atcatcggca tagtatatcg gcatagtata atacgacaag
     3061 gtgaggaact aaaccatggc caagttgacc agtgccgttc cggtgctcac cgcgcgcgac
     3121 gtcgccggag cggtcgagtt ctggaccgac cggctcgggt tctcccggga cttcgtggag
     3181 gacgacttcg ccggtgtggt ccgggacgac gtgaccctgt tcatcagcgc ggtccaggac
     3241 caggtggtgc cggacaacac cctggcctgg gtgtgggtgc gcggcctgga cgagctgtac
     3301 gccgagtggt cggaggtcgt gtccacgaac ttccgggacg cctccgggcc ggccatgacc
     3361 gagatcggcg agcagccgtg ggggcgggag ttcgccctgc gcgacccggc cggcaactgc
     3421 gtgcacttcg tggccgagga gcaggactga cacgtgctac gagatttcga ttccaccgcc
     3481 gccttctatg aaaggttggg cttcggaatc gttttccggg acgccggctg gatgatcctc
     3541 cagcgcgggg atctcatgct ggagttcttc gcccacccca acttgtttat tgcagcttat
     3601 aatggttaca aataaagcaa tagcatcaca aatttcacaa ataaagcatt tttttcactg
     3661 cattctagtt gtggtttgtc caaactcatc aatgtatctt atcatgtctg tataccgtcg
     3721 acctctagct agagcttggc gtaatcatgg tcatagctgt ttcctgtgtg aaattgttat
     3781 ccgctcacaa ttccacacaa catacgagcc ggaagcataa agtgtaaagc ctggggtgcc
     3841 taatgagtga gctaactcac attaattgcg ttgcgctcac tgcccgcttt ccagtcggga
     3901 aacctgtcgt gccagctgca ttaatgaatc ggccaacgcg cggggagagg cggtttgcgt
     3961 attgggcgct cttccgcttc ctcgctcact gactcgctgc gctcggtcgt tcggctgcgg
     4021 cgagcggtat cagctcactc aaaggcggta atacggttat ccacagaatc aggggataac
     4081 gcaggaaaga acatgtgagc aaaaggccag caaaaggcca ggaaccgtaa aaaggccgcg
     4141 ttgctggcgt ttttccatag gctccgcccc cctgacgagc atcacaaaaa tcgacgctca
     4201 agtcagaggt ggcgaaaccc gacaggacta taaagatacc aggcgtttcc ccctggaagc
     4261 tccctcgtgc gctctcctgt tccgaccctg ccgcttaccg gatacctgtc cgcctttctc
     4321 ccttcgggaa gcgtggcgct ttctcatagc tcacgctgta ggtatctcag ttcggtgtag
     4381 gtcgttcgct ccaagctggg ctgtgtgcac gaaccccccg ttcagcccga ccgctgcgcc
     4441 ttatccggta actatcgtct tgagtccaac ccggtaagac acgacttatc gccactggca
     4501 gcagccactg gtaacaggat tagcagagcg aggtatgtag gcggtgctac agagttcttg
     4561 aagtggtggc ctaactacgg ctacactaga agaacagtat ttggtatctg cgctctgctg
     4621 aagccagtta ccttcggaaa aagagttggt agctcttgat ccggcaaaca aaccaccgct
     4681 ggtagcggtg gtttttttgt ttgcaagcag cagattacgc gcagaaaaaa aggatctcaa
     4741 gaagatcctt tgatcttttc tacggggtct gacgctcagt ggaacgaaaa ctcacgttaa
     4801 gggattttgg tcatgagatt atcaaaaagg atcttcacct agatcctttt aaattaaaaa
     4861 tgaagtttta aatcaatcta aagtatatat gagtaaactt ggtctgacag ttaccaatgc
     4921 ttaatcagtg aggcacctat ctcagcgatc tgtctatttc gttcatccat agttgcctga
     4981 ctccccgtcg tgtagataac tacgatacgg gagggcttac catctggccc cagtgctgca
     5041 atgataccgc gagacccacg ctcaccggct ccagatttat cagcaataaa ccagccagcc
     5101 ggaagggccg agcgcagaag tggtcctgca actttatccg cctccatcca gtctattaat
     5161 tgttgccggg aagctagagt aagtagttcg ccagttaata gtttgcgcaa cgttgttgcc
     5221 attgctacag gcatcgtggt gtcacgctcg tcgtttggta tggcttcatt cagctccggt
     5281 tcccaacgat caaggcgagt tacatgatcc cccatgttgt gcaaaaaagc ggttagctcc
     5341 ttcggtcctc cgatcgttgt cagaagtaag ttggccgcag tgttatcact catggttatg
     5401 gcagcactgc ataattctct tactgtcatg ccatccgtaa gatgcttttc tgtgactggt
     5461 gagtactcaa ccaagtcatt ctgagaatag tgtatgcggc gaccgagttg ctcttgcccg
     5521 gcgtcaatac gggataatac cgcgccacat agcagaactt taaaagtgct catcattgga
     5581 aaacgttctt cggggcgaaa actctcaagg atcttaccgc tgttgagatc cagttcgatg
     5641 taacccactc gtgcacccaa ctgatcttca gcatctttta ctttcaccag cgtttctggg
     5701 tgagcaaaaa caggaaggca aaatgccgca aaaaagggaa taagggcgac acggaaatgt
     5761 tgaatactca tactcttcct ttttcaatat tattgaagca tttatcaggg ttattgtctc
     5821 atgagcggat acatatttga atgtatttag aaaaataaac aaataggggt tccgcgcaca
     5881 tttccccgaa aagtgccacc tgacgtc
//
