LOCUS       sequence-342068-        6968 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 6968)
  AUTHORS   Shida T, Cueva JG, Xu Z, Goodman MB, Nachury MV
  TITLE     The major alpha-tubulin K40 acetyltransferase alphaTAT1 promotes 
            rapid ciliogenesis and efficient mechanosensation.
  JOURNAL   Proc Natl Acad Sci U S A. 2010 Dec 14;107(50):21517-22. doi: 
            10.1073/pnas.1013728107. Epub 2010 Nov 10.
  PUBMED    21068373
REFERENCE   2  (bases 1 to 6968)
  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: 
            "10.1073/pnas.1013728107"; journalName: "Proc Natl Acad Sci U S A"; 
            date: "2010-12-14- 14"; volume: "107"; issue: "50"; pages: 
            "21517-22"
COMMENT     Sequence Label: pEF5B-FRT-GFP-alpha-TAT1
FEATURES             Location/Qualifiers
     source          1..6968
                     /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"
     promoter        351..1529
                     /label=EF-1-alpha promoter
                     /note="strong constitutive promoter for human elongation 
                     factor EF-1-alpha"
     intron          582..1520
                     /label=EF-1-alpha intron A
                     /note="intron upstream of the start codon of human 
                     EF-1-alpha"
     primer_bind     1477..1497
                     /label=EF1a-F
                     /note="Human elongation factor-1a promoter, forward primer"
     primer_bind     1546..1565
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        1546..1564
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             join(1597..1599,1600..1602,1603..2313)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             join(1597..1599,1600..1602,1603..2313)
                     /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(1642..1663)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(1903..1922)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     2250..2271
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     CDS             2326..2346
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQG"
     CDS             2359..2403
                     /codon_start=1
                     /product="affinity and epitope tag derived from pancreatic 
                     ribonuclease A"
                     /label=S-Tag
                     /translation="KETAAAKFERQHMDS"
     protein_bind    2440..2464
                     /gene="mutant version of attB"
                     /label=attB1
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     protein_bind    complement(3460..3484)
                     /gene="mutant version of attB"
                     /label=attB2
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     CDS             3537..3578
                     /codon_start=1
                     /product="epitope tag from simian virus 5"
                     /label=V5 tag
                     /translation="GKPIPNPLLGLDST"
     primer_bind     complement(3616..3633)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    3622..3846
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(3979..3998)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     protein_bind    4130..4177
                     /label=FRT
                     /bound_moiety="FLP recombinase from the Saccharomyces 
                     cerevisiae 2u plasmid"
                     /note="FLP-mediated recombination occurs in the 8-bp core 
                     sequence TCTAGAAA (Turan and Bode, 2011)."
     CDS             4185..4580
                     /codon_start=1
                     /gene="Aspergillus terreus BSD"
                     /product="blasticidin S deaminase"
                     /label=BSD
                     /note="confers resistance to blasticidin"
                     /translation="AKPLSQEESTLIERATATINSIPISEDYSVASAALSSDGRIFTGV
                     NVYHFTGGPCAELVVLGTAAAAAAGNLTCIVAIGNENRGILSPCGRCRQVLLDLHPGIK
                     AIVKDSDGQPTAVGIRELLPSGYVWEG"
     polyA_signal    4643..4762
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4678..4697)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4732..4751
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(4811..4827)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4811..4827)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4824..4846)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4835..4851
                     /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(4859..4865,4866..4883,4884..4889))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4904..4925
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5042..5059)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5213..5801)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5293..5312)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5972..6763,6764..6832))
                     /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     6595..6614
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6833..6937)
                     /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 taggcttttg caaaaagctt tgcaaagatg gataaagttt taaacagaga
      301 ggaatctttg cagctaatgg accttctagg tcttgaaagg agtgggaatt ggctccggtg
      361 cccgtcagtg ggcagagcgc acatcgccca cagtccccga gaagttgggg ggaggggtcg
      421 gcaattgaac cggtgcctag agaaggtggc gcggggtaaa ctgggaaagt gatgtcgtgt
      481 actggctccg cctttttccc gagggtgggg gagaaccgta tataagtgca gtagtcgccg
      541 tgaacgttct ttttcgcaac gggtttgccg ccagaacaca ggtaagtgcc gtgtgtggtt
      601 cccgcgggcc tggcctcttt acgggttatg gcccttgcgt gccttgaatt acttccacct
      661 ggctgcagta cgtgattctt gatcccgagc ttcgggttgg aagtgggtgg gagagttcga
      721 ggccttgcgc ttaaggagcc ccttcgcctc gtgcttgagt tgaggcctgg cctgggcgct
      781 ggggccgccg cgtgcgaatc tggtggcacc ttcgcgcctg tctcgctgct ttcgataagt
      841 ctctagccat ttaaaatttt tgatgacctg ctgcgacgct ttttttctgg caagatagtc
      901 ttgtaaatgc gggccaagat ctgcacactg gtatttcggt ttttggggcc gcgggcggcg
      961 acggggcccg tgcgtcccag cgcacatgtt cggcgaggcg gggcctgcga gcgcggccac
     1021 cgagaatcgg acgggggtag tctcaagctg gccggcctgc tctggtgcct ggcctcgcgc
     1081 cgccgtgtat cgccccgccc tgggcggcaa ggctggcccg gtcggcacca gttgcgtgag
     1141 cggaaagatg gccgcttccc ggccctgctg cagggagctc aaaatggagg acgcggcgct
     1201 cgggagagcg ggcgggtgag tcacccacac aaaggaaaag ggcctttccg tcctcagccg
     1261 tcgcttcatg tgactccacg gagtaccggg cgccgtccag gcacctcgat tagttctcga
     1321 gcttttggag tacgtcgtct ttaggttggg gggaggggtt ttatgcgatg gagtttcccc
     1381 acactgagtg ggtggagact gaagttaggc cagcttggca cttgatgtaa ttctccttgg
     1441 aatttgccct ttttgagttt ggatcttggt tcattctcaa gcctcagaca gtggttcaaa
     1501 gtttttttct tccatttcag gtgtcgtgag gaattagctt ggtactaata cgactcacta
     1561 tagggagacc caagctggct aggtaagctt ggtaccatgg tgagcaaggg cgaggagctg
     1621 ttcaccgggg tggtgcccat cctggtcgag ctggacggcg acgtaaacgg ccacaagttc
     1681 agcgtgtccg gcgagggcga gggcgatgcc acctacggca agctgaccct gaagttcatc
     1741 tgcaccaccg gcaagctgcc cgtgccctgg cccaccctcg tgaccaccct gacctacggc
     1801 gtgcagtgct tcagccgcta ccccgaccac atgaagcagc acgacttctt caagtccgcc
     1861 atgcccgaag gctacgtcca ggagcgcacc atcttcttca aggacgacgg caactacaag
     1921 acccgcgccg aggtgaagtt cgagggcgac accctggtga accgcatcga gctgaagggc
     1981 atcgacttca aggaggacgg caacatcctg gggcacaagc tggagtacaa ctacaacagc
     2041 cacaacgtct atatcatggc cgacaagcag aagaacggca tcaaggtgaa cttcaagatc
     2101 cgccacaaca tcgaggacgg cagcgtgcag ctcgccgacc actaccagca gaacaccccc
     2161 atcggcgacg gccccgtgct gctgcccgac aaccactacc tgagcaccca gtccgccctg
     2221 agcaaagacc ccaacgagaa gcgcgatcac atggtcctgc tggagttcgt gaccgccgcc
     2281 gggatcactc tcggcatgga cgagctgtac aagggcgggg gcggggagaa tctttatttt
     2341 cagggcggcg ggggcgggaa agaaaccgct gctgctaaat tcgaacgcca gcacatggac
     2401 agcggactag tccagtgtgg tggaattctg cagatatcaa caagtttgta caaaaaagtt
     2461 gatgagttcc cgttcgatgt ggacgcgctg ttcccggagc ggatcacggt gctggaccag
     2521 cacctgaggc ccccagcccg ccgacccgga accacaacgc cggcccgtgt tgatctacag
     2581 cagcaaatta tgaccattat agatgaactg ggcaaggctt ctgccaaggc ccagaatctt
     2641 tccgctccta tcactagtgc atcaaggatg cagagtaacc gccatgttgt ttatattctc
     2701 aaagacagtt cagcccgacc ggctggaaaa ggagccatta ttggtttcat caaagttgga
     2761 tacaagaagc tctttgtact ggatgatcgt gaggctcata atgaggtaga accactttgc
     2821 atcctggact tttacatcca tgagtctgtg caacgccatg gccatgggcg agaactcttc
     2881 cagtatatgt tgcagaagga gcgagtggaa ccgcaccaac tggcaattga ccgaccctca
     2941 cagaagctgc tgaaattcct gaataagcac tacaatctgg agaccacagt cccacaggtg
     3001 aacaactttg tgatctttga aggcttcttt gcccatcaac atcggccccc tgctccctct
     3061 ctgagggcaa ctcgacactc tcgtgctgct gcagtcgatc ccacgcccgc tgctccagca
     3121 aggaagctgc cacccaagag agcagaggga gacatcaagc catactcctc tagtgaccga
     3181 gaatttctga aggtagctgt ggagcctcct tggcccctaa acagggcccc tcgccgcgcc
     3241 acacctccag cccacccacc cccccgctcc agcagcctgg gaaactcacc agaacgaggt
     3301 cccctccgcc cctttgtgcc agagcaggag ctgctgcgtt ccttgcgcct ctgcccccca
     3361 caccctaccg cccgccttct gttggctgct gaccctgggg gcagcccagc tcaacgtcgt
     3421 cgcaccagct cccttccccg ctctgaggag agtcgatact aacaactttc ttgtacaaag
     3481 tggttgatat ccagcacagt ggcggccgct cgagtctaga gggcccgcgg ttcgaaggta
     3541 agcctatccc taaccctctc ctcggtctcg attctacgcg taccggttag taatgagttt
     3601 aaacccgctg atcagcctcg actgtgcctt ctagttgcca gccatctgtt gtttgcccct
     3661 cccccgtgcc ttccttgacc ctggaaggtg ccactcccac tgtcctttcc taataaaatg
     3721 aggaaattgc atcgcattgt ctgagtaggt gtcattctat tctggggggt ggggtggggc
     3781 aggacagcaa gggggaggat tgggaagaca atagcaggca tgctggggat gcggtgggct
     3841 ctatggcttc tgaggcggaa agaaccagct ggggctctag ggggtatccc cacgcgccct
     3901 gtagcggcgc attaagcgcg gcgggtgtgg tggttacgcg cagcgtgacc gctacacttg
     3961 ccagcgccct agcgcccgct cctttcgctt tcttcccttc ctttctcgcc acgttcgccg
     4021 gctttccccg tcaagctcta aatcgggggc tccctttagg gttccgattt agtgctttac
     4081 ggcacctcga ccccaaaaaa cttgattagg gtgatggttc acgtacctag aagttcctat
     4141 tccgaagttc ctattctcta gaaagtatag gaacttcctt ggccgccaag cctttgtctc
     4201 aagaagaatc caccctcatt gaaagagcaa cggctacaat caacagcatc cccatctctg
     4261 aagactacag cgtcgccagc gcagctctct ctagcgacgg ccgcatcttc actggtgtca
     4321 atgtatatca ttttactggg ggaccttgtg cagaactcgt ggtgctgggc actgctgctg
     4381 ctgcggcagc tggcaacctg acttgtatcg tcgcgatcgg aaatgagaac aggggcatct
     4441 tgagcccctg cggacggtgc cgacaggtgc ttctcgatct gcatcctggg atcaaagcca
     4501 tagtgaagga cagtgatgga cagccgacgg cagttgggat tcgtgaattg ctgccctctg
     4561 gttatgtgtg ggagggctaa gcacaattcg agctcggtac ctttaagacc aatgacttac
     4621 aaggcagctg tagatcttag ccaacttgtt tattgcagct taatggttac aaataaagca
     4681 atagcatcac aaatttcaca aataaagcat ttttttcact gcattctagt tgtggtttgt
     4741 ccaaactcat caatgtatct tatcatgtct gtataccgtc gacctctagc tagagcttgg
     4801 cgtaatcatg gtcatagctg tttcctgtgt gaaattgtta tccgctcaca attccacaca
     4861 acatacgagc cggaagcata aagtgtaaag cctggggtgc ctaatgagtg agctaactca
     4921 cattaattgc gttgcgctca ctgcccgctt tccagtcggg aaacctgtcg tgccagctgc
     4981 attaatgaat cggccaacgc gcggggagag gcggtttgcg tattgggcgc tcttccgctt
     5041 cctcgctcac tgactcgctg cgctcggtcg ttcggctgcg gcgagcggta tcagctcact
     5101 caaaggcggt aatacggtta tccacagaat caggggataa cgcaggaaag aacatgtgag
     5161 caaaaggcca gcaaaaggcc aggaaccgta aaaaggccgc gttgctggcg tttttccata
     5221 ggctccgccc ccctgacgag catcacaaaa atcgacgctc aagtcagagg tggcgaaacc
     5281 cgacaggact ataaagatac caggcgtttc cccctggaag ctccctcgtg cgctctcctg
     5341 ttccgaccct gccgcttacc ggatacctgt ccgcctttct cccttcggga agcgtggcgc
     5401 tttctcatag ctcacgctgt aggtatctca gttcggtgta ggtcgttcgc tccaagctgg
     5461 gctgtgtgca cgaacccccc gttcagcccg accgctgcgc cttatccggt aactatcgtc
     5521 ttgagtccaa cccggtaaga cacgacttat cgccactggc agcagccact ggtaacagga
     5581 ttagcagagc gaggtatgta ggcggtgcta cagagttctt gaagtggtgg cctaactacg
     5641 gctacactag aagaacagta tttggtatct gcgctctgct gaagccagtt accttcggaa
     5701 aaagagttgg tagctcttga tccggcaaac aaaccaccgc tggtagcggt ggtttttttg
     5761 tttgcaagca gcagattacg cgcagaaaaa aaggatctca agaagatcct ttgatctttt
     5821 ctacggggtc tgacgctcag tggaacgaaa actcacgtta agggattttg gtcatgagat
     5881 tatcaaaaag gatcttcacc tagatccttt taaattaaaa atgaagtttt aaatcaatct
     5941 aaagtatata tgagtaaact tggtctgaca gttaccaatg cttaatcagt gaggcaccta
     6001 tctcagcgat ctgtctattt cgttcatcca tagttgcctg actccccgtc gtgtagataa
     6061 ctacgatacg ggagggctta ccatctggcc ccagtgctgc aatgataccg cgagacccac
     6121 gctcaccggc tccagattta tcagcaataa accagccagc cggaagggcc gagcgcagaa
     6181 gtggtcctgc aactttatcc gcctccatcc agtctattaa ttgttgccgg gaagctagag
     6241 taagtagttc gccagttaat agtttgcgca acgttgttgc cattgctaca ggcatcgtgg
     6301 tgtcacgctc gtcgtttggt atggcttcat tcagctccgg ttcccaacga tcaaggcgag
     6361 ttacatgatc ccccatgttg tgcaaaaaag cggttagctc cttcggtcct ccgatcgttg
     6421 tcagaagtaa gttggccgca gtgttatcac tcatggttat ggcagcactg cataattctc
     6481 ttactgtcat gccatccgta agatgctttt ctgtgactgg tgagtactca accaagtcat
     6541 tctgagaata gtgtatgcgg cgaccgagtt gctcttgccc ggcgtcaata cgggataata
     6601 ccgcgccaca tagcagaact ttaaaagtgc tcatcattgg aaaacgttct tcggggcgaa
     6661 aactctcaag gatcttaccg ctgttgagat ccagttcgat gtaacccact cgtgcaccca
     6721 actgatcttc agcatctttt actttcacca gcgtttctgg gtgagcaaaa acaggaaggc
     6781 aaaatgccgc aaaaaaggga ataagggcga cacggaaatg ttgaatactc atactcttcc
     6841 tttttcaata ttattgaagc atttatcagg gttattgtct catgagcgga tacatatttg
     6901 aatgtattta gaaaaataaa caaatagggg ttccgcgcac atttccccga aaagtgccac
     6961 ctgacgtc
//
