LOCUS       sequence-222818-        7895 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  synthetic circular DNA
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7895)
  AUTHORS   Tulu US, Rusan NM, Wadsworth P
  TITLE     Peripheral, non-centrosome-associated microtubules contribute to 
            spindle formation in centrosome-containing cells.
  JOURNAL   Curr Biol. 2003 Oct 28. 13(21):1894-9.
  PUBMED    14588246
REFERENCE   2  (bases 1 to 7895)
  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: "Curr Biol. 
            2003 Oct 28. 13(21):1894-9."
COMMENT     Sequence Label: pIREShyg-PA-GFP-alpha Tubulin
FEATURES             Location/Qualifiers
     source          1..7895
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(44..63)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        235..614
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        615..818
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     769..789
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     863..882
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        863..881
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     regulatory      950..959
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(956..958,959..961,962..1672)
                     /codon_start=1
                     /product="photoactivatable GFP variant (Patterson et al., 
                     2002)"
                     /label=PA-GFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTFSYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     ANFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSHQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(1001..1022)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(1262..1281)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1609..1630
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     intron          3129..3358
                     /label=chimeric intron
                     /note="chimera between introns from adenovirus and 
                     immunoglobulin heavy chain genes"
     primer_bind     3341..3360
                     /label=pMT2-F
                     /note="Synthetic intron, forward primer"
     misc_feature    join(3441..4015,4016..4018,4019..4027)
                     /label=IRES2
                     /note="internal ribosome entry site (IRES) of the 
                     encephalomyocarditis virus (EMCV)"
     primer_bind     complement(3608..3625)
                     /label=IRES reverse
                     /note="IRES internal ribosome entry site, reverse primer. 
                     Also called pCDH-rev"
     primer_bind     3835..3854
                     /label=IRES-F
                     /note="IRES internal ribosome entry site, forward primer"
     CDS             4053..5072
                     /codon_start=1
                     /product="hygromycin B phosphotransferase"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="KPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRGYV
                     LRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLPET
                     ELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQTV
                     MDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMFGD
                     SQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDGNF
                     DDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAKE"
     CDS             4053..5072
                     /codon_start=1
                     /gene="aph(4)-Ia"
                     /product="aminoglycoside phosphotransferase from E. coli"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="KPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRGYV
                     LRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLPET
                     ELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQTV
                     MDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMFGD
                     SQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDGNF
                     DDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAKE"
     primer_bind     complement(5382..5399)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    5388..5612
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     5659..5678
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(5738..5754)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5738..5754)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(5751..5773)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5762..5778
                     /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(5786..5792,5793..5810,5811..5816))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5831..5852
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5969..5986)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(6140..6728)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(6220..6239)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6899..7690,7691..7759))
                     /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     7522..7541
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7760..7864)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 gacggatcgg gagatctccc gatcccctat ggtgcactct cagtacaatc tgctctgatg
       61 ccgcatagtt aagccagtat ctgctccctg cttgtgtgtt ggaggtcgct gagtagtgcg
      121 cgagcaaaat ttaagctaca acaaggcaag gcttgaccga caattgcatg aagaatctgc
      181 ttagggttag gcgttttgcg ctgcttcgcg atgtacgggc cagatatacg cgttgacatt
      241 gattattgac tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata
      301 tggagttccg cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc
      361 cccgcccatt gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc
      421 attgacgtca atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt
      481 atcatatgcc aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt
      541 atgcccagta catgacctta tgggactttc ctacttggca gtacatctac gtattagtca
      601 tcgctattac catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg
      661 actcacgggg atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc
      721 aaaatcaacg ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg
      781 gtaggcgtgt acggtgggag gtctatataa gcagagctct ctggctaact agagaaccca
      841 ctgcttactg gcttatcgaa attaatacga ctcactatag ggagacccaa gcttggtacc
      901 gagctcggat ctgtacaggc cttaagccgg cgcgctagcg ctaccggtcg ccaccatggt
      961 gagcaagggc gaggagctgt tcaccggggt ggtgcccatc ctggtcgagc tggacggcga
     1021 cgtaaacggc cacaagttca gcgtgtccgg cgagggcgag ggcgatgcca cctacggcaa
     1081 gctgaccctg aagttcatct gcaccaccgg caagctgccc gtgccctggc ccaccctcgt
     1141 gaccaccttc agctacggcg tgcagtgctt cagccgctac cccgaccaca tgaagcagca
     1201 cgacttcttc aagtccgcca tgcccgaagg ctacgtccag gagcgcacca tcttcttcaa
     1261 ggacgacggc aactacaaga cccgcgccga ggtgaagttc gagggcgaca ccctggtgaa
     1321 ccgcatcgag ctgaagggca tcgacttcaa ggaggacggc aacatcctgg ggcacaagct
     1381 ggagtacaac tacaacagcc acaacgtcta tatcatggcc gacaagcaga agaacggcat
     1441 caaggccaac ttcaagatcc gccacaacat cgaggacggc agcgtgcagc tcgccgacca
     1501 ctaccagcag aacaccccca tcggcgacgg ccccgtgctg ctgcccgaca accactacct
     1561 gagccaccag tccgccctga gcaaagaccc caacgagaag cgcgatcaca tggtcctgct
     1621 ggagttcgtg accgccgccg ggatcactct cggcatggac gagctgtaca agtccggact
     1681 cagatctcga gtgcgtgagt gcatctccat ccacgttggc caggctggtg tccagattgg
     1741 caatgcctgc tgggagctct actgcctgga acacggcatc cagcccgatg gccagatgcc
     1801 aagtgacaag accattgggg gaggagatga ctccttcaac accttcttca gtgagacggg
     1861 cgctggcaag cacgtgcccc gggctgtgtt tgtagacttg gaacccacag tcattgatga
     1921 agttcgcact ggcacctacc gccagctctt ccaccctgag cagctcatca caggcaagga
     1981 agatgctgcc aataactatg cccgagggca ctacaccatt ggcaaggaga tcattgacct
     2041 tgtgttggac cgaattcgca agctggctga ccagtgcacc ggtcttcagg gcttcttggt
     2101 tttccacagc tttggtgggg gaactggttc tgggttcacc tccctgctca tggaacgtct
     2161 ctcagttgat tatggcaaga agtccaagct ggagttctcc atttacccag caccccaggt
     2221 ttccacagct gtagttgagc cctacaactc catcctcacc acccacacca ccctggagca
     2281 ctctgattgt gccttcatgg tagacaatga ggccatctat gacatctgtc gtagaaacct
     2341 cgatatcgag cgcccaacct acactaacct taaccgcctt attagccaga ttgtgtcctc
     2401 catcactgct tccctgagat ttgatggagc cctgaatgtt gacctgacag aattccagac
     2461 caacctggtg ccctaccccc gcatccactt ccctctggcc acatatgccc ctgtcatctc
     2521 tgctgagaaa gcctaccatg aacagctttc tgtagcagag atcaccaatg cttgctttga
     2581 gccagccaac cagatggtga aatgtgaccc tcgccatggt aaatacatgg cttgctgcct
     2641 gttgtaccgt ggtgacgtgg ttcccaaaga tgtcaatgct gccattgcca ccatcaaaac
     2701 caagcgcagc atccagtttg tggattggtg ccccactggc ttcaaggttg gcatcaacta
     2761 ccagcctccc actgtggtgc ctggtggaga cctggccaag gtacagagag ctgtgtgcat
     2821 gctgagcaac accacagcca ttgctgaggc ctgggctcgc ctggaccaca agtttgacct
     2881 gatgtatgcc aagcgtgcct ttgttcactg gtacgtgggt gaggggatgg aggaaggcga
     2941 gttttcagag gcccgtgaag atatggctgc ccttgagaag gattatgagg aggttggtgt
     3001 ggattctgtt gaaggagagg gtgaggaaga aggagaggaa tactaaggat ccgatatcgt
     3061 acgttaacat agataactga tccagtgtgc tggaattaat tcgctgtctg cgagggccag
     3121 ctgttggggt gagtactccc tctcaaaagc gggcatgact tctgcgctaa gattgtcagt
     3181 ttccaaaaac gaggaggatt tgatattcac ctggcccgcg gtgatgcctt tgagggtggc
     3241 cgcgtccatc tggtcagaaa agacaatctt tttgttgtca agcttgaggt gtggcaggct
     3301 tgagatctgg ccatacactt gagtgacaat gacatccact ttgcctttct ctccacaggt
     3361 gtccactccc aggtccaact gcaggtcgag catgcatcta gggcggccgc actagaggaa
     3421 ttccgcccct ctcccccccc cccctctccc tccccccccc ctaacgttac tggccgaagc
     3481 cgcttggaat aaggccggtg tgcgtttgtc tatatgttat tttccaccat attgccgtct
     3541 tttggcaatg tgagggcccg gaaacctggc cctgtcttct tgacgagcat tcctaggggt
     3601 ctttcccctc tcgccaaagg aatgcaaggt ctgttgaatg tcgtgaagga agcagttcct
     3661 ctggaagctt cttgaagaca aacaacgtct gtagcgaccc tttgcaggca gcggaacccc
     3721 ccacctggcg acaggtgcct ctgcggccaa aagccacgtg tataagatac acctgcaaag
     3781 gcggcacaac cccagtgcca cgttgtgagt tggatagttg tggaaagagt caaatggctc
     3841 tcctcaagcg tattcaacaa ggggctgaag gatgcccaga aggtacccca ttgtatggga
     3901 tctgatctgg ggcctcggtg cacatgcttt acatgtgttt agtcgaggtt aaaaaaacgt
     3961 ctaggccccc cgaaccacgg ggacgtggtt ttcctttgaa aaacacgatg ataagcttgc
     4021 cacaacccgt accaaagatg gatagatccg gaaagcctga actcaccgcg acgtctgtcg
     4081 agaagtttct gatcgaaaag ttcgacagcg tctccgacct gatgcagctc tcggagggcg
     4141 aagaatctcg tgctttcagc ttcgatgtag gagggcgtgg atatgtcctg cgggtaaata
     4201 gctgcgccga tggtttctac aaagatcgtt atgtttatcg gcactttgca tcggccgcgc
     4261 tcccgattcc ggaagtgctt gacattgggg aattcagcga gagcctgacc tattgcatct
     4321 cccgccgtgc acagggtgtc acgttgcaag acctgcctga aaccgaactg cccgctgttc
     4381 tgcagccggt cgcggaggcc atggatgcga tcgctgcggc cgatcttagc cagacgagcg
     4441 ggttcggccc attcggaccg caaggaatcg gtcaatacac tacatggcgt gatttcatat
     4501 gcgcgattgc tgatccccat gtgtatcact ggcaaactgt gatggacgac accgtcagtg
     4561 cgtccgtcgc gcaggctctc gatgagctga tgctttgggc cgaggactgc cccgaagtcc
     4621 ggcacctcgt gcacgcggat ttcggctcca acaatgtcct gacggacaat ggccgcataa
     4681 cagcggtcat tgactggagc gaggcgatgt tcggggattc ccaatacgag gtcgccaaca
     4741 tcttcttctg gaggccgtgg ttggcttgta tggagcagca gacgcgctac ttcgagcgga
     4801 ggcatccgga gcttgcagga tcgccgcggc tccgggcgta tatgctccgc attggtcttg
     4861 accaactcta tcagagcttg gttgacggca atttcgatga tgcagcttgg gcgcagggtc
     4921 gatgcgacgc aatcgtccga tccggagccg ggactgtcgg gcgtacacaa atcgcccgca
     4981 gaagcgcggc cgtctggacc gatggctgtg tagaagtact cgccgatagt ggaaaccgac
     5041 gccccagcac tcgtccgagg gcaaaggaat agagtagatg ccgaccgaac aagagctgat
     5101 ttcgagaacg cctcagccag caactcgcgc gagcctagca aggcaaatgc gagagaacgg
     5161 ccttacgctt ggtggcacag ttctcgtcca cagttcgcta agctcgctcg gctgggtcgc
     5221 gggagggccg gtcgcagtga ttcaggccct tctggattgt gttggtcccc agggcacgat
     5281 tgtcatgccc acgcactcgg gtgatctgac tgatcccgca gattggagat cgccgcccgt
     5341 gcctgccgat tgggtgcaga tctctagagc tcgctgatca gcctcgactg tgccttctag
     5401 ttgccagcca tctgttgttt gcccctcccc cgtgccttcc ttgaccctgg aaggtgccac
     5461 tcccactgtc ctttcctaat aaaatgagga aattgcatcg cattgtctga gtaggtgtca
     5521 ttctattctg gggggtgggg tggggcagga cagcaagggg gaggattggg aagacaatag
     5581 caggcatgct ggggatgcgg tgggctctat ggcttctgag gcggaaagaa ccagctgggg
     5641 ctcgagtgca ttctagttgt ggtttgtcca aactcatcaa tgtatcttat catgtctgta
     5701 taccgtcgac ctctagctag agcttggcgt aatcatggtc atagctgttt cctgtgtgaa
     5761 attgttatcc gctcacaatt ccacacaaca tacgagccgg aagcataaag tgtaaagcct
     5821 ggggtgccta atgagtgagc taactcacat taattgcgtt gcgctcactg cccgctttcc
     5881 agtcgggaaa cctgtcgtgc cagctgcatt aatgaatcgg ccaacgcgcg gggagaggcg
     5941 gtttgcgtat tgggcgctct tccgcttcct cgctcactga ctcgctgcgc tcggtcgttc
     6001 ggctgcggcg agcggtatca gctcactcaa aggcggtaat acggttatcc acagaatcag
     6061 gggataacgc aggaaagaac atgtgagcaa aaggccagca aaaggccagg aaccgtaaaa
     6121 aggccgcgtt gctggcgttt ttccataggc tccgcccccc tgacgagcat cacaaaaatc
     6181 gacgctcaag tcagaggtgg cgaaacccga caggactata aagataccag gcgtttcccc
     6241 ctggaagctc cctcgtgcgc tctcctgttc cgaccctgcc gcttaccgga tacctgtccg
     6301 cctttctccc ttcgggaagc gtggcgcttt ctcatagctc acgctgtagg tatctcagtt
     6361 cggtgtaggt cgttcgctcc aagctgggct gtgtgcacga accccccgtt cagcccgacc
     6421 gctgcgcctt atccggtaac tatcgtcttg agtccaaccc ggtaagacac gacttatcgc
     6481 cactggcagc agccactggt aacaggatta gcagagcgag gtatgtaggc ggtgctacag
     6541 agttcttgaa gtggtggcct aactacggct acactagaag aacagtattt ggtatctgcg
     6601 ctctgctgaa gccagttacc ttcggaaaaa gagttggtag ctcttgatcc ggcaaacaaa
     6661 ccaccgctgg tagcggtggt ttttttgttt gcaagcagca gattacgcgc agaaaaaaag
     6721 gatctcaaga agatcctttg atcttttcta cggggtctga cgctcagtgg aacgaaaact
     6781 cacgttaagg gattttggtc atgagattat caaaaaggat cttcacctag atccttttaa
     6841 attaaaaatg aagttttaaa tcaatctaaa gtatatatga gtaaacttgg tctgacagtt
     6901 accaatgctt aatcagtgag gcacctatct cagcgatctg tctatttcgt tcatccatag
     6961 ttgcctgact ccccgtcgtg tagataacta cgatacggga gggcttacca tctggcccca
     7021 gtgctgcaat gataccgcga gacccacgct caccggctcc agatttatca gcaataaacc
     7081 agccagccgg aagggccgag cgcagaagtg gtcctgcaac tttatccgcc tccatccagt
     7141 ctattaattg ttgccgggaa gctagagtaa gtagttcgcc agttaatagt ttgcgcaacg
     7201 ttgttgccat tgctacaggc atcgtggtgt cacgctcgtc gtttggtatg gcttcattca
     7261 gctccggttc ccaacgatca aggcgagtta catgatcccc catgttgtgc aaaaaagcgg
     7321 ttagctcctt cggtcctccg atcgttgtca gaagtaagtt ggccgcagtg ttatcactca
     7381 tggttatggc agcactgcat aattctctta ctgtcatgcc atccgtaaga tgcttttctg
     7441 tgactggtga gtactcaacc aagtcattct gagaatagtg tatgcggcga ccgagttgct
     7501 cttgcccggc gtcaatacgg gataataccg cgccacatag cagaacttta aaagtgctca
     7561 tcattggaaa acgttcttcg gggcgaaaac tctcaaggat cttaccgctg ttgagatcca
     7621 gttcgatgta acccactcgt gcacccaact gatcttcagc atcttttact ttcaccagcg
     7681 tttctgggtg agcaaaaaca ggaaggcaaa atgccgcaaa aaagggaata agggcgacac
     7741 ggaaatgttg aatactcata ctcttccttt ttcaatatta ttgaagcatt tatcagggtt
     7801 attgtctcat gagcggatac atatttgaat gtatttagaa aaataaacaa ataggggttc
     7861 cgcgcacatt tccccgaaaa gtgccacctg acgtc
//
