LOCUS       sequence-391831-        8987 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expression of Need1-rCM1 harboring a (F75A) mutation in the rat (r) 
            Centrosomin motif 1 (CM1) and fused to mOrange2. F75A mutatuon 
            renders CM1 incapable of binding and activating gamma-TuRC.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8987)
  AUTHORS   Vinopal S, Dupraz S, Alfadil E, Pietralla T, Bendre S, Stiess M, 
            Falk S, Camargo Ortega G, Maghelli N, Tolic IM, Smejkal J, Gotz M, 
            Bradke F
  TITLE     Centrosomal microtubule nucleation regulates radial migration of 
            projection neurons independently of polarization in the developing 
            brain.
  JOURNAL   Neuron. 2023 Apr 19;111(8):1241-1263.e16. doi: 
            10.1016/j.neuron.2023.01.020. Epub 2023 Feb 15.
  PUBMED    36796357
REFERENCE   2  (bases 1 to 8987)
  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: 
            "10.1016/j.neuron.2023.01.020"; journalName: "Neuron"; date: 
            "2023-04-19- 19"; volume: "111"; issue: "8"; pages: "1241-1263.e16"
COMMENT     Sequence Label: pBactin-Nedd1-mOrange2-F75A
FEATURES             Location/Qualifiers
     source          1..8987
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     1179..1201
                     /label=pCAGGS-5
                     /note="Chimeric intron in CAG promoter, forward primer"
     CDS             3303..4010
                     /codon_start=1
                     /product="photostable monomeric orange derivative of DsRed 
                     fluorescent protein (Shaner et al., 2008)"
                     /label=mOrange2
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEENNMAIIKEFMRFKVRMEGSVNGHEFEIEGEGEGRPYEG
                     FQTAKLKVTKGGPLPFAWDILSPHFTYGSKAYVKHPADIPDYFKLSFPEGFKWERVMNY
                     EDGGVVTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASSERMYPEDGALK
                     GKIKMRLKLKDGGHYTSEVKTTYKAKKPVQLPGAYIVDIKLDITSHNEDYTIVEQYERA
                     EGRHSTGGMDELYK"
     polyA_signal    4383..4517
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4420..4439)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4474..4493
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(4531..4547)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4531..4547)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4544..4566)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4555..4571
                     /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(4579..4585,4586..4603,4604..4609))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4624..4645
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4762..4779)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4933..5521)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5013..5032)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5692..6483,6484..6552))
                     /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     6315..6334
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6553..6657)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     6725..6743
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(6781..6803)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     6903..6922
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     7116..7138
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     7130..7147
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     7131..7147
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     promoter        join(8948..8987,1..236)
                     /label=chicken beta-actin promoter
ORIGIN
        1 ccccccctcc ccacccccaa ttttgtattt atttattttt taattatttt gtgcagcgat
       61 gggggsgggg gggggggggg cgcgcgccag gcggggcggg gcggggcgag gggcggggcg
      121 gggcgaggcg gagaggtgcg gcggcagcca atcagagcgg cgcgctccga aagtttcctt
      181 ttatggcgag gcggcggcgg cggcggccct ataaaaagcg aagcgcgcgg cgggcgggag
      241 tcgctgcgcg ctgccttcgc cccgtgcccc gctccgccgc cgcctcgcgc cgcccgcccc
      301 ggctctgact gaccgcgtta ctcccacagg tgagcgggcg ggacggccct tctcctccgg
      361 gctgtaatta gcgcttggtt taatgacggc tcgtttcttt tctgtggctg cgtgaaagcc
      421 ttaaagggct ccgggagggc cctttgtgcg ggggggagcg gctcgggggg tgcgtgcgtg
      481 tgtgtgtgcg tggggagcgc cgcgtgcggc tccgcgctgc ccggcggctg tgagcgctgc
      541 gggcgcggcg cggggctttg tgcgctccgc agtgtgcgcg aggggagcgc ggccgggggc
      601 ggtgccccgc ggtgcggggg gggctgcgag gggaacaaag gctgcgtgcg gggtgtgtgc
      661 gtgggggggt gagcaggggg tgtgggcgcg gcggtcgggc tgtaaccccc ccctgcaccc
      721 ccctccccga gttgctgagc acggcccggc ttcgggtgcg gggctccgtg cggggcgtgg
      781 cgcggggctc gccgtgccgg gcggggggtg gcggcaggtg ggggtgccgg gcggggcggg
      841 gccgcctcgg gccggggagg gctcggggga ggggcgcggc ggcccccgga gcgccggcgg
      901 ctgtcgaggc gcggcgagcc gcagccattg ccttttatgg taatcgtgcg agagggcgca
      961 gggacttcct ttgtcccaaa tctgtgcgga gccgaaatct gggaggcgcc gccgcacccc
     1021 ctctagcggg cgcggggcga agcggtgcgg cgccggcagg aaggaaatgg gcggggaggg
     1081 ccttcgtgcg tcgccgcgcc gccgtcccct tctccatctc cagcctcggg gctgtccgca
     1141 gggggacggc tgccttcggg ggggacgggg cagggcgggg ttcggcttct ggcgtgtgac
     1201 cggcggggtt tatatcttcc cttctctgtt cctccgcagc ccccaagcta gcagccacca
     1261 tgcaggaaaa cctcagattt gcctcatcag gagatgatgt taaaatatgg gatgcttcgt
     1321 ttctgacatt ggtggataaa ttcaaccccc acacgtcacc gcatggaatc agttcaatct
     1381 gttggagcag caataataac ttcctagtga ctgcgtcttc tggtggtgac aaaatagtgg
     1441 tgtcaagttg caaatgtaag cccgttccac ttttggagct tgcagaaggg caaaagcaga
     1501 catgtgtcga tttaaattct acatcaatgt atttggcaag tggaggccta aataacactg
     1561 ttaatatttg ggatttaaag tcaaaacgac ttcatcggtc tcttaaggac cataaacggg
     1621 aagtaacttg tgtggcgtac aattggaatg attgttacat tgcttccggg tctctcagtg
     1681 gtgagattat cttacacagc gtaaccacta atacctccag tacgcctttc ggccatggga
     1741 gtaagcagcc tgtccggcac atcaagtact cgttgtttag gaagtctctc ttgggcagcg
     1801 tttcggataa tggagtagtg accctctggg atgtgaacag tcagagtgca taccacagct
     1861 ttgacagtac acacaaagcc ccagcttcag gcctctgctt ctctcctgtc aacgaactgc
     1921 tgtttgtgac cataggcctg gataaaagaa tcatcctgta tgacacttca agtaagaagc
     1981 tggtgaaaac tttagtggct gacactcctc tgactgctgt agattttatg ccagatggag
     2041 ccagtttggc tattggatcc tcccgtggga aaatatatca gtatgactta aggatgctga
     2101 aatctcccgt gaaaaccatc agcgcccaca agacatctgt gcagtgtata gcatttcagt
     2161 actccacatc tctgacgaag gcaagtttaa ataaaggctg ctcaaataag gccacggcag
     2221 cgaataaacg gagtgttcct gtgggcagca gcagtggggc tgcccagaat tctggaattg
     2281 tcagagaagc acccagccct tccattgcca cagttctccc acagcccgtg acgacagctg
     2341 tgggcaaagg agcaggggct gctcaggacg aagcaggttt gcctcgaagc aaaaatacag
     2401 atatcttatc taaggaaact gatgctggaa aaagtcaaga cttctccagc tttgatgaca
     2461 cggggaagag tagtttaggg gacatgtttt caccaattag agatgatgct gtagtcaaca
     2521 aaggaggtga cgagtctgta ggcaaaggag atggccttga ctttctacca cagctaaact
     2581 cggtgttccc cctgaggaag aatgcaggct cctccagcac tctggtgctg cactccagcc
     2641 cttttaatgt ccttatgggg acagggaagg aagagcatga gagtcacgag ctgtcagccg
     2701 agtctaagaa ggcacacctt gggaagccgg agcctaagga ctccttgaag cagtttacaa
     2761 agttgatctc ctctggtgct gaacctggaa ttctaaacac ctgtccatca tctaaccaag
     2821 caagaaatct tgaaaagttt gaaaagccag aaaaagatat tgaagtccag ttgatgcacg
     2881 aaccctcagt caatggaccc tcaaccccag taccaaaggc agcctcctcc gtcactgctg
     2941 gagtagccag ttcactctca gaaaaaatag tcgacaccct tgggaacagt cggccggccg
     3001 cgccacttac ctctgtccag atccgcttca ttcagaacat gatacaagaa accctggatg
     3061 acttcaggga agcttgccat agggacattg tgaatctgca agtggaaatg attaaacagt
     3121 ttcatatgca actgaatgaa atgcactctt tgctggaaag atactcagtg aatgaaggct
     3181 tagtggctga gatcgaaagg ctgcgagaag aaaacaaaag gctgcgggcc cacttctcaa
     3241 gcctcctcga cggcagcaag cttggtggag gcagcgcatc aggcggaagc ggaagcagat
     3301 ctatggtgag caagggcgag gagaataaca tggccatcat caaggagttc atgcgcttca
     3361 aggtgcgcat ggagggctcc gtgaacggcc acgagttcga gatcgagggc gagggcgagg
     3421 gccgccccta cgagggcttt cagaccgcta agctgaaggt gaccaagggt ggccccctgc
     3481 ccttcgcctg ggacatcctg tcccctcatt tcacctacgg ctccaaggcc tacgtgaagc
     3541 accccgccga catccccgac tacttcaagc tgtccttccc cgagggcttc aagtgggagc
     3601 gcgtgatgaa ctacgaggac ggcggcgtgg tgaccgtgac ccaggactcc tccctgcagg
     3661 acggcgagtt catctacaag gtgaagctgc gcggcaccaa cttcccctcc gacggccccg
     3721 tgatgcagaa gaagaccatg ggctgggagg cctcctccga gcggatgtac cccgaggacg
     3781 gtgccctgaa gggcaagatc aagatgaggc tgaagctgaa ggacggcggc cactacacct
     3841 ccgaggtcaa gaccacctac aaggccaaga agcccgtgca gctgcccggc gcctacatcg
     3901 tcgacatcaa gttggacatc acctcccaca acgaggacta caccatcgtg gaacagtacg
     3961 aacgcgccga gggccgccac tccaccggcg gcatggacga gctgtacaag tctggcacag
     4021 gtggaagcat cgataaggtc tctcccacca gagcccggaa catgaaggac tttgaaaacc
     4081 aaatcactga gctgaagaag gagaacgcca atctaaagct ccggatctac ttccttgagg
     4141 agaggataca gcaggagttt gctggcccca cagagcacat ctactaagcg gccgccatat
     4201 gcatcctagg cctattaata ttccggagta tacgtagccg gctaacgtta acaaccggta
     4261 cctctagata agtaatgatc ataatcagcc atatcacatc tgtagaggtt ttacttgctt
     4321 taaaaaacct cccacacctc cccctgaacc tgaaacataa aatgaatgca attgttgttg
     4381 ttaacttgtt tattgcagct tataatggtt acaaataaag caatagcatc acaaatttca
     4441 caaataaagc atttttttca ctgcattcta gttgtggttt gtccaaactc atcaatgtat
     4501 cttatcatgt ctggatcaat tcgtaatcat gtcatagctg tttcctgtgt gaaattgtta
     4561 tccgctcaca attccacaca acatacgagc cggaagcata aagtgtaaag cctggggtgc
     4621 ctaatgagtg agctaactca cattaattgc gttgcgctca ctgcccgctt tccagtcggg
     4681 aaacctgtcg tgccagctgc attaatgaat cggccaacgc gcggggagag gcggtttgcg
     4741 tattgggcgc tcttccgctt cctcgctcac tgactcgctg cgctcggtcg ttcggctgcg
     4801 gcgagcggta tcagctcact caaaggcggt aatacggtta tccacagaat caggggataa
     4861 cgcaggaaag aacatgtgag caaaaggcca gcaaaaggcc aggaaccgta aaaaggccgc
     4921 gttgctggcg tttttccata ggctccgccc ccctgacgag catcacaaaa atcgacgctc
     4981 aagtcagagg tggcgaaacc cgacaggact ataaagatac caggcgtttc cccctggaag
     5041 ctccctcgtg cgctctcctg ttccgaccct gccgcttacc ggatacctgt ccgcctttct
     5101 cccttcggga agcgtggcgc tttctcatag ctcacgctgt aggtatctca gttcggtgta
     5161 ggtcgttcgc tccaagctgg gctgtgtgca cgaacccccc gttcagcccg accgctgcgc
     5221 cttatccggt aactatcgtc ttgagtccaa cccggtaaga cacgacttat cgccactggc
     5281 agcagccact ggtaacagga ttagcagagc gaggtatgta ggcggtgcta cagagttctt
     5341 gaagtggtgg cctaactacg gctacactag aagaacagta tttggtatct gcgctctgct
     5401 gaagccagtt accttcggaa aaagagttgg tagctcttga tccggcaaac aaaccaccgc
     5461 tggtagcggt ggtttttttg tttgcaagca gcagattacg cgcagaaaaa aaggatctca
     5521 agaagatcct ttgatctttt ctacggggtc tgacgctcag tggaacgaaa actcacgtta
     5581 agggattttg gtcatgagat tatcaaaaag gatcttcacc tagatccttt taaattaaaa
     5641 atgaagtttt aaatcaatct aaagtatata tgagtaaact tggtctgaca gttaccaatg
     5701 cttaatcagt gaggcaccta tctcagcgat ctgtctattt cgttcatcca tagttgcctg
     5761 actccccgtc gtgtagataa ctacgatacg ggagggctta ccatctggcc ccagtgctgc
     5821 aatgataccg cgagacccac gctcaccggc tccagattta tcagcaataa accagccagc
     5881 cggaagggcc gagcgcagaa gtggtcctgc aactttatcc gcctccatcc agtctattaa
     5941 ttgttgccgg gaagctagag taagtagttc gccagttaat agtttgcgca acgttgttgc
     6001 cattgctaca ggcatcgtgg tgtcacgctc gtcgtttggt atggcttcat tcagctccgg
     6061 ttcccaacga tcaaggcgag ttacatgatc ccccatgttg tgcaaaaaag cggttagctc
     6121 cttcggtcct ccgatcgttg tcagaagtaa gttggccgca gtgttatcac tcatggttat
     6181 ggcagcactg cataattctc ttactgtcat gccatccgta agatgctttt ctgtgactgg
     6241 tgagtactca accaagtcat tctgagaata gtgtatgcgg cgaccgagtt gctcttgccc
     6301 ggcgtcaata cgggataata ccgcgccaca tagcagaact ttaaaagtgc tcatcattgg
     6361 aaaacgttct tcggggcgaa aactctcaag gatcttaccg ctgttgagat ccagttcgat
     6421 gtaacccact cgtgcaccca actgatcttc agcatctttt actttcacca gcgtttctgg
     6481 gtgagcaaaa acaggaaggc aaaatgccgc aaaaaaggga ataagggcga cacggaaatg
     6541 ttgaatactc atactcttcc tttttcaata ttattgaagc atttatcagg gttattgtct
     6601 catgagcgga tacatatttg aatgtattta gaaaaataaa caaatagggg ttccgcgcac
     6661 atttccccga aaagtgccac ctgacgtcta agaaaccatt attatcatga cattaaccta
     6721 taaaaatagg cgtatcacga ggccctttcg tctcgcgcgt ttcggtgatg acggtgaaaa
     6781 cctctgacac atgcagctcc cggagacggt cacagcttgt ctgtaagcgg atgccgggag
     6841 cagacaagcc cgtcagggcg cgtcagcggg tgttggcggg tgtcggggct ggcttaacta
     6901 tgcggcatca gagcagattg tactgagagt gcaccatatg cggtgtgaaa taccgcacag
     6961 atgcgtaagg agaaaatacc gcatcaggcg ccattcgcca ttcaggctgc gcaactgttg
     7021 ggaagggcga tcggtgcggg cctcttcgct attacgccag ctggcgaaag ggggatgtgc
     7081 tgcaaggcga ttaagttggg taacgccagg gttttcccag tcacgacgtt gtaaaacgac
     7141 ggccagtgcc aagttgggat ctttgcattg gcccacggct ctcaggatgg ggatgctccc
     7201 cttcagcacc cggttcccct tggaaactga tggtcctggc tctgtggcat ggcagtggca
     7261 ctgtgaggag ccccctacca gcagcacaca gtggggttgg cactgccacg ctccggatgc
     7321 cgcgctctga tccaacccca taatcaaggg aacccgaatt gccccatcat tgcccccacc
     7381 acccccatcc tgccgggccc tcacacccca cgctgccttc tggtgacatt ccccagccca
     7441 aacccacggc ttcatggcta ccgcggggca tttcccattg ccgccccatt atcagctctg
     7501 cacacctccc gctgtaccca tgcctcgtgg ctgcccttct ttgacgtata atcttctaat
     7561 taatacccgg ccttgtcaaa gtggagcaca aacgttaatt aattccccag caggcaggta
     7621 attaacagtg tgactccctt tttgctgcga gtggggctga tacagagaga tgtggcacta
     7681 tggagcccac ggggtcctgg cactgggtgc ccacggaggt ccccatgtgc tgcagtgtca
     7741 ccgcctccga ggagacagta ttgtccctgc ggtgtccctg cagctcagct ctgtccacag
     7801 ggccacctcc agtttggagg ggacacaatg cagccccgat gcaacccatc ctcgcagcat
     7861 cccagggaca aagaccccac tgcaagaccg cacacagggc tgggtcccgc tcccctaata
     7921 tctacagtgc ttcttgcatg gccccttaat caatgcagtt aatcagcatg cgctcatgca
     7981 ccgctctgga gctgcaaagc ccctcgcagc gctgctcacc aacaccgcgc accgccccgg
     8041 cccagcctgc agcacgcgct gcaaacagga aagaaacaaa atattgccca aatgtaggca
     8101 aaggcattcg gctgccttga cctccgccgg gccgggccct gcctgactca gctccttact
     8161 cagcgctcgc ttcctccctc cggctgccac cgccgcagcg cacaccctga caaagagtgg
     8221 cccttaacgg gctctgaggt gcacccagca gtgcactcag cagtccaagg gccggcctgg
     8281 aggtttgcac cgctacgtgc tgacattagc attgaacttg gccctgggta gtgctgcagg
     8341 ccgggcgggg tgggtgtaga gagtgcagcg cgcgttgcac ccggtgcccc ttcccctccc
     8401 ttgcatccca gcaggctgca ccccagcacc aggcccgtgc atgcatgctc ctggtgttat
     8461 tgcagcctgg tgcatgcatg cgtcttgatg gtgcagcgct gtgcatgcat cctccttggt
     8521 gtgtagcagt ctagtgcatg catacccctc ggtgttattg ctgctctgtg cacgcacgct
     8581 cattgtatca cttcatccca gtgcatgcac tcacactgga gcgattgctg ctcggtgcac
     8641 gcacactcat tgtatcactg cagctcagtg gctgcacgca caccggtgtt attgctgctc
     8701 ggtgcgtgca tgcacatcag tgtcgctgca gctcagtgca tgcacgctca ttgcccatcg
     8761 ctatccctgc ctctcctgct ggcgctcccc gggaggtgac ttcaagggga ccgcaggacc
     8821 acctcggggg tggggggagg gctgcacacg cggaccccgc tccccctccc caacaaagca
     8881 ctgtggaatc aaaaaggggg gaggggggat ggaggggcgc gtcacacccc cgccccacac
     8941 cctcacctcg aggtgagccc cacgttctgc ttcactctcc ccatctc
//
