LOCUS       sequence-94539-e        5447 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Knocks down TACC3 in mammalian cells via shRNA and co-expresses 
            GFP..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5447)
  AUTHORS   Booth DG, Hood FE, Prior IA, Royle SJ
  TITLE     A TACC3/ch-TOG/clathrin complex stabilises kinetochore fibres by 
            inter-microtubule bridging.
  JOURNAL   EMBO J. 2011 Mar 2;30(5):906-19. doi: 10.1038/emboj.2011.15. Epub 
            2011 Feb 4.
  PUBMED    21297582
REFERENCE   2  (bases 1 to 5447)
  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/emboj.2011.15"; journalName: "EMBO J"; date: "2011-03-2- 
            2"; volume: "30"; issue: "5"; pages: "906-19"
COMMENT     Sequence Label: pBrain-GFP-shTACC3
FEATURES             Location/Qualifiers
     source          1..5447
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     64..86
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     78..95
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     79..95
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     102..121
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        102..120
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     153..169
                     /label=SK primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     153..169
                     /label=pBluescriptSK
                     /note="For pBluescript vector"
     promoter        189..403
                     /label=H1 promoter
                     /note="human H1 RNA promoter"
     primer_bind     324..343
                     /label=H1
                     /note="Human H1 promoter, forward primer"
     primer_bind     complement(476..492)
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(477..493)
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     promoter        complement(521..539)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(560..576)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(560..576)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(573..595)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    584..600
                     /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(608..614,615..632,633..638))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    653..674
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     enhancer        777..1080
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        1081..1284
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     1235..1255
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     regulatory      1323..1332
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(1329..1331,1332..1334,1335..2045)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             join(1329..1331,1332..1334,1335..2045)
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(1374..1395)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(1635..1654)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1982..2003
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     misc_feature    2046..2111
                     /label=MCS
                     /note="multiple cloning site"
     polyA_signal    2235..2356
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2272..2291)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     2326..2345
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(2363..2818)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2500..2521)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     2712..2731
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        2845..2949
                     /gene="bla"
                     /label=AmpR promoter
     promoter        2951..3308
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        2951..3142
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(2974..2994)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      3159..3294
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             3343..4137
                     /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             3343..4137
                     /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(3397..3416)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4007..4026
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(4325..4344)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    4369..4416
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      4745..5333
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5234..5253
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 tagttattct ggcgaaaggg ggatgtgctg caaggcgatt aagttgggta acgccagggt
       61 tttcccagtc acgacgttgt aaaacgacgg ccagtgaatt gtaatacgac tcactatagg
      121 gcgaattgga gctccaccgc ggtggcggcc gctctagaac tagtggatcc cccgggctgc
      181 aggaattcga acgctgacgt catcaacccg ctccaaggaa tcgcgggccc agtgtcacta
      241 ggcgggaaca cccagcgcgc gtgcgccctg gcaggaagat ggctgtgagg gacaggggag
      301 tggcgccctg caatatttgc atgtcgctat gtgttctggg aaatcaccat aaacgtgaaa
      361 tgtctttgga tttgggaatc ttataagttc tgtatgagac cacagatccc cgttaccgga
      421 agatcgtctg ttcaagagac agacgatctt ccggtaactt tttggaaaag cttatcgata
      481 ccgtcgacct cgaggggggg cccggtacca gcttttgttc cctttagtga gggttaattt
      541 cgagcttggc gtaatcatgg tcatagctgt ttcctgtgtg aaattgttat ccgctcacaa
      601 ttccacacaa catacgagcc ggaagcataa agtgtaaagc ctggggtgcc taatgagtga
      661 gctaactcac attaattgcg ttgcgctcac tgcccgcttt ccagtcggga aacctgtcgt
      721 gccagatagt aatcaattac ggggtcatta gttcatagcc catatatgga gttccgcgtt
      781 acataactta cggtaaatgg cccgcctggc tgaccgccca acgacccccg cccattgacg
      841 tcaataatga cgtatgttcc catagtaacg ccaataggga ctttccattg acgtcaatgg
      901 gtggagtatt tacggtaaac tgcccacttg gcagtacatc aagtgtatca tatgccaagt
      961 acgcccccta ttgacgtcaa tgacggtaaa tggcccgcct ggcattatgc ccagtacatg
     1021 accttatggg actttcctac ttggcagtac atctacgtat tagtcatcgc tattaccatg
     1081 gtgatgcggt tttggcagta catcaatggg cgtggatagc ggtttgactc acggggattt
     1141 ccaagtctcc accccattga cgtcaatggg agtttgtttt ggcaccaaaa tcaacgggac
     1201 tttccaaaat gtcgtaacaa ctccgcccca ttgacgcaaa tgggcggtag gcgtgtacgg
     1261 tgggaggtct atataagcag agctggttta gtgaaccgtc agatccgcta gcgctaccgg
     1321 tcgccaccat ggtgagcaag ggcgaggagc tgttcaccgg ggtggtgccc atcctggtcg
     1381 agctggacgg cgacgtaaac ggccacaagt tcagcgtgtc cggcgagggc gagggcgatg
     1441 ccacctacgg caagctgacc ctgaagttca tctgcaccac cggcaagctg cccgtgccct
     1501 ggcccaccct cgtgaccacc ctgacctacg gcgtgcagtg cttcagccgc taccccgacc
     1561 acatgaagca gcacgacttc ttcaagtccg ccatgcccga aggctacgtc caggagcgca
     1621 ccatcttctt caaggacgac ggcaactaca agacccgcgc cgaggtgaag ttcgagggcg
     1681 acaccctggt gaaccgcatc gagctgaagg gcatcgactt caaggaggac ggcaacatcc
     1741 tggggcacaa gctggagtac aactacaaca gccacaacgt ctatatcatg gccgacaagc
     1801 agaagaacgg catcaaggtg aacttcaaga tccgccacaa catcgaggac ggcagcgtgc
     1861 agctcgccga ccactaccag cagaacaccc ccatcggcga cggccccgtg ctgctgcccg
     1921 acaaccacta cctgagcacc cagtccgccc tgagcaaaga ccccaacgag aagcgcgatc
     1981 acatggtcct gctggagttc gtgaccgccg ccgggatcac tctcggcatg gacgagctgt
     2041 acaagtccgg actcagatct cgagctcaag cttcgaattc tgcagtcgac ggtaccgcgg
     2101 gcccgggatc caccggatct agataactga tcataatcag ccataccaca tttgtagagg
     2161 ttttacttgc tttaaaaaac ctcccacacc tccccctgaa cctgaaacat aaaatgaatg
     2221 caattgttgt tgttaacttg tttattgcag cttataatgg ttacaaataa agcaatagca
     2281 tcacaaattt cacaaataaa gcattttttt cactgcattc tagttgtggt ttgtccaaac
     2341 tcatcaatgt atcttaacgc gtaaattgta agcgttaata ttttgttaaa attcgcgtta
     2401 aatttttgtt aaatcagctc attttttaac caataggccg aaatcggcaa aatcccttat
     2461 aaatcaaaag aatagaccga gatagggttg agtgttgttc cagtttggaa caagagtcca
     2521 ctattaaaga acgtggactc caacgtcaaa gggcgaaaaa ccgtctatca gggcgatggc
     2581 ccactacgtg aaccatcacc ctaatcaagt tttttggggt cgaggtgccg taaagcacta
     2641 aatcggaacc ctaaagggag cccccgattt agagcttgac ggggaaagcc ggcgaacgtg
     2701 gcgagaaagg aagggaagaa agcgaaagga gcgggcgcta gggcgctggc aagtgtagcg
     2761 gtcacgctgc gcgtaaccac cacacccgcc gcgcttaatg cgccgctaca gggcgcgtca
     2821 ggtggcactt ttcggggaaa tgtgcgcgga acccctattt gtttattttt ctaaatacat
     2881 tcaaatatgt atccgctcat gagacaataa ccctgataaa tgcttcaata atattgaaaa
     2941 aggaagagtc ctgaggcgga aagaaccagc tgtggaatgt gtgtcagtta gggtgtggaa
     3001 agtccccagg ctccccagca ggcagaagta tgcaaagcat gcatctcaat tagtcagcaa
     3061 ccaggtgtgg aaagtcccca ggctccccag caggcagaag tatgcaaagc atgcatctca
     3121 attagtcagc aaccatagtc ccgcccctaa ctccgcccat cccgccccta actccgccca
     3181 gttccgccca ttctccgccc catggctgac taattttttt tatttatgca gaggccgagg
     3241 ccgcctcggc ctctgagcta ttccagaagt agtgaggagg cttttttgga ggcctaggct
     3301 tttgcaaaga tcgatcaaga gacaggatga ggatcgtttc gcatgattga acaagatgga
     3361 ttgcacgcag gttctccggc cgcttgggtg gagaggctat tcggctatga ctgggcacaa
     3421 cagacaatcg gctgctctga tgccgccgtg ttccggctgt cagcgcaggg gcgcccggtt
     3481 ctttttgtca agaccgacct gtccggtgcc ctgaatgaac tgcaagacga ggcagcgcgg
     3541 ctatcgtggc tggccacgac gggcgttcct tgcgcagctg tgctcgacgt tgtcactgaa
     3601 gcgggaaggg actggctgct attgggcgaa gtgccggggc aggatctcct gtcatctcac
     3661 cttgctcctg ccgagaaagt atccatcatg gctgatgcaa tgcggcggct gcatacgctt
     3721 gatccggcta cctgcccatt cgaccaccaa gcgaaacatc gcatcgagcg agcacgtact
     3781 cggatggaag ccggtcttgt cgatcaggat gatctggacg aagagcatca ggggctcgcg
     3841 ccagccgaac tgttcgccag gctcaaggcg agcatgcccg acggcgagga tctcgtcgtg
     3901 acccatggcg atgcctgctt gccgaatatc atggtggaaa atggccgctt ttctggattc
     3961 atcgactgtg gccggctggg tgtggcggac cgctatcagg acatagcgtt ggctacccgt
     4021 gatattgctg aagagcttgg cggcgaatgg gctgaccgct tcctcgtgct ttacggtatc
     4081 gccgctcccg attcgcagcg catcgccttc tatcgccttc ttgacgagtt cttctgagcg
     4141 ggactctggg gttcgaaatg accgaccaag cgacgcccaa cctgccatca cgagatttcg
     4201 attccaccgc cgccttctat gaaaggttgg gcttcggaat cgttttccgg gacgccggct
     4261 ggatgatcct ccagcgcggg gatctcatgc tggagttctt cgcccaccct agggggaggc
     4321 taactgaaac acggaaggag acaataccgg aaggaacccg cgctatgacg gcaataaaaa
     4381 gacagaataa aacgcacggt gttgggtcgt ttgttcataa acgcggggtt cggtcccagg
     4441 gctggcactc tgtcgatacc ccaccgagac cccattgggg ccaatacgcc cgcgtttctt
     4501 ccttttcccc accccacccc ccaagttcgg gtgaaggccc agggctcgca gccaacgtcg
     4561 gggcggcagg ccctgccata gcctcaggtt actcatatat actttagatt gatttaaaac
     4621 ttcattttta atttaaaagg atctaggtga agatcctttt tgataatctc atgaccaaaa
     4681 tcccttaacg tgagttttcg ttccactgag cgtcagaccc cgtagaaaag atcaaaggat
     4741 cttcttgaga tccttttttt ctgcgcgtaa tctgctgctt gcaaacaaaa aaaccaccgc
     4801 taccagcggt ggtttgtttg ccggatcaag agctaccaac tctttttccg aaggtaactg
     4861 gcttcagcag agcgcagata ccaaatactg tccttctagt gtagccgtag ttaggccacc
     4921 acttcaagaa ctctgtagca ccgcctacat acctcgctct gctaatcctg ttaccagtgg
     4981 ctgctgccag tggcgataag tcgtgtctta ccgggttgga ctcaagacga tagttaccgg
     5041 ataaggcgca gcggtcgggc tgaacggggg gttcgtgcac acagcccagc ttggagcgaa
     5101 cgacctacac cgaactgaga tacctacagc gtgagctatg agaaagcgcc acgcttcccg
     5161 aagggagaaa ggcggacagg tatccggtaa gcggcagggt cggaacagga gagcgcacga
     5221 gggagcttcc agggggaaac gcctggtatc tttatagtcc tgtcgggttt cgccacctct
     5281 gacttgagcg tcgatttttg tgatgctcgt caggggggcg gagcctatgg aaaaacgcca
     5341 gcaacgcggc ctttttacgg ttcctggcct tttgctggcc ttttgctcac atgttctttc
     5401 ctgcgttatc ccctgattct gtggataacc gtattaccgc catgcat
//
