LOCUS       sequence-12825-e        6970 bp    DNA     circular SYN 14-APR-2025
DEFINITION  synthetic circular DNA
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6970)
  AUTHORS   Shida T, Cueva JG, Xu Z, Goodman MB, Nachury MV
  TITLE     The major {alpha}-tubulin K40 acetyltransferase {alpha}TAT1 promotes
            rapid ciliogenesis and efficient mechanosensation.
  JOURNAL   Proc Natl Acad Sci U S A. 2010 Nov 10. ():.
  PUBMED    21068373
REFERENCE   2  (bases 1 to 6970)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Apr 14, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Proc Natl 
            Acad Sci U S A. 2010 Nov 10. ():."
COMMENT     Sequence Label: pEF5B-FRT-GFP-alpha-TAT1[D157N]
FEATURES             Location/Qualifiers
     source          1..6970
                     /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        353..1531
                     /label=EF-1-alpha promoter
                     /note="strong constitutive promoter for human elongation 
                     factor EF-1-alpha"
     intron          584..1522
                     /label=EF-1-alpha intron A
                     /note="intron upstream of the start codon of human 
                     EF-1-alpha"
     primer_bind     1479..1499
                     /label=EF1a-F
                     /note="Human elongation factor-1a promoter, forward primer"
     primer_bind     1548..1567
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        1548..1566
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             join(1599..1601,1602..1604,1605..2315)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             join(1599..1601,1602..1604,1605..2315)
                     /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(1644..1665)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(1905..1924)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     2252..2273
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     CDS             2328..2348
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQG"
     CDS             2361..2405
                     /codon_start=1
                     /product="affinity and epitope tag derived from pancreatic 
                     ribonuclease A"
                     /label=S-Tag
                     /translation="KETAAAKFERQHMDS"
     protein_bind    2442..2466
                     /gene="mutant version of attB"
                     /label=attB1
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     protein_bind    complement(3462..3486)
                     /gene="mutant version of attB"
                     /label=attB2
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     primer_bind     complement(3618..3635)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    3624..3848
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(3981..4000)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     protein_bind    4132..4179
                     /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             4187..4582
                     /codon_start=1
                     /gene="Aspergillus terreus BSD"
                     /product="blasticidin S deaminase"
                     /label=BSD
                     /note="confers resistance to blasticidin"
                     /translation="AKPLSQEESTLIERATATINSIPISEDYSVASAALSSDGRIFTGV
                     NVYHFTGGPCAELVVLGTAAAAAAGNLTCIVAIGNENRGILSPCGRCRQVLLDLHPGIK
                     AIVKDSDGQPTAVGIRELLPSGYVWEG"
     polyA_signal    4645..4764
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4680..4699)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4734..4753
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(4813..4829)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4813..4829)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4826..4848)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4837..4853
                     /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(4861..4867,4868..4885,4886..4891))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4906..4927
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5044..5061)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5215..5803)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5295..5314)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5974..6765,6766..6834))
                     /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     6597..6616
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6835..6939)
                     /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 agtgcctcgt gaggctccgg
      361 tgcccgtcag tgggcagagc gcacatcgcc cacagtcccc gagaagttgg ggggaggggt
      421 cggcaattga accggtgcct agagaaggtg gcgcggggta aactgggaaa gtgatgtcgt
      481 gtactggctc cgcctttttc ccgagggtgg gggagaaccg tatataagtg cagtagtcgc
      541 cgtgaacgtt ctttttcgca acgggtttgc cgccagaaca caggtaagtg ccgtgtgtgg
      601 ttcccgcggg cctggcctct ttacgggtta tggcccttgc gtgccttgaa ttacttccac
      661 ctggctgcag tacgtgattc ttgatcccga gcttcgggtt ggaagtgggt gggagagttc
      721 gaggccttgc gcttaaggag ccccttcgcc tcgtgcttga gttgaggcct ggcctgggcg
      781 ctggggccgc cgcgtgcgaa tctggtggca ccttcgcgcc tgtctcgctg ctttcgataa
      841 gtctctagcc atttaaaatt tttgatgacc tgctgcgacg ctttttttct ggcaagatag
      901 tcttgtaaat gcgggccaag atctgcacac tggtatttcg gtttttgggg ccgcgggcgg
      961 cgacggggcc cgtgcgtccc agcgcacatg ttcggcgagg cggggcctgc gagcgcggcc
     1021 accgagaatc ggacgggggt agtctcaagc tggccggcct gctctggtgc ctggcctcgc
     1081 gccgccgtgt atcgccccgc cctgggcggc aaggctggcc cggtcggcac cagttgcgtg
     1141 agcggaaaga tggccgcttc ccggccctgc tgcagggagc tcaaaatgga ggacgcggcg
     1201 ctcgggagag cgggcgggtg agtcacccac acaaaggaaa agggcctttc cgtcctcagc
     1261 cgtcgcttca tgtgactcca cggagtaccg ggcgccgtcc aggcacctcg attagttctc
     1321 gagcttttgg agtacgtcgt ctttaggttg gggggagggg ttttatgcga tggagtttcc
     1381 ccacactgag tgggtggaga ctgaagttag gccagcttgg cacttgatgt aattctcctt
     1441 ggaatttgcc ctttttgagt ttggatcttg gttcattctc aagcctcaga cagtggttca
     1501 aagttttttt cttccatttc aggtgtcgtg aggaattagc ttggtactaa tacgactcac
     1561 tatagggaga cccaagctgg ctaggtaagc ttggtaccat ggtgagcaag ggcgaggagc
     1621 tgttcaccgg ggtggtgccc atcctggtcg agctggacgg cgacgtaaac ggccacaagt
     1681 tcagcgtgtc cggcgagggc gagggcgatg ccacctacgg caagctgacc ctgaagttca
     1741 tctgcaccac cggcaagctg cccgtgccct ggcccaccct cgtgaccacc ctgacctacg
     1801 gcgtgcagtg cttcagccgc taccccgacc acatgaagca gcacgacttc ttcaagtccg
     1861 ccatgcccga aggctacgtc caggagcgca ccatcttctt caaggacgac ggcaactaca
     1921 agacccgcgc cgaggtgaag ttcgagggcg acaccctggt gaaccgcatc gagctgaagg
     1981 gcatcgactt caaggaggac ggcaacatcc tggggcacaa gctggagtac aactacaaca
     2041 gccacaacgt ctatatcatg gccgacaagc agaagaacgg catcaaggtg aacttcaaga
     2101 tccgccacaa catcgaggac ggcagcgtgc agctcgccga ccactaccag cagaacaccc
     2161 ccatcggcga cggccccgtg ctgctgcccg acaaccacta cctgagcacc cagtccgccc
     2221 tgagcaaaga ccccaacgag aagcgcgatc acatggtcct gctggagttc gtgaccgccg
     2281 ccgggatcac tctcggcatg gacgagctgt acaagggcgg gggcggggag aatctttatt
     2341 ttcagggcgg cgggggcggg aaagaaaccg ctgctgctaa attcgaacgc cagcacatgg
     2401 acagcggact agtccagtgt ggtggaattc tgcagatatc aacaagtttg tacaaaaaag
     2461 ttgatgagtt cccgttcgat gtggacgcgc tgttcccgga gcggatcacg gtgctggacc
     2521 agcacctgag gcccccagcc cgccgacccg gaaccacaac gccggcccgt gttgatctac
     2581 agcagcaaat tatgaccatt atagatgaac tgggcaaggc ttctgccaag gcccagaatc
     2641 tttccgctcc tatcactagt gcatcaagga tgcagagtaa ccgccatgtt gtttatattc
     2701 tcaaagacag ttcagcccga ccggctggaa aaggagccat tattggtttc atcaaagttg
     2761 gatacaagaa gctctttgta ctggatgatc gtgaggctca taatgaggta gaaccacttt
     2821 gcatcctgga cttttacatc catgagtctg tgcaacgcca tggccatggg cgagaactct
     2881 tccagtatat gttgcagaag gagcgagtgg aaccgcacca actggcaatt aaccgaccct
     2941 cacagaagct gctgaaattc ctgaataagc actacaatct ggagaccaca gtcccacagg
     3001 tgaacaactt tgtgatcttt gaaggcttct ttgcccatca acatcggccc cctgctccct
     3061 ctctgagggc aactcgacac tctcgtgctg ctgcagtcga tcccacgccc gctgctccag
     3121 caaggaagct gccacccaag agagcagagg gagacatcaa gccatactcc tctagtgacc
     3181 gagaatttct gaaggtagct gtggagcctc cttggcccct aaacagggcc cctcgccgcg
     3241 ccacacctcc agcccaccca cccccccgct ccagcagcct gggaaactca ccagaacgag
     3301 gtcccctccg cccctttgtg ccagagcagg agctgctgcg ttccttgcgc ctctgccccc
     3361 cacaccctac cgcccgcctt ctgttggctg ctgaccctgg gggcagccca gctcaacgtc
     3421 gtcgcaccag ctcccttccc cgctctgagg agagtcgata ctaacaactt tcttgtacaa
     3481 agtggttgat atccagcaca gtggcggccg ctcgagtcta gagggcccgc ggttcgaagg
     3541 taagcctatc cctaaccctc tcctcggtct cgatctacgc ggtaccggtt agtaatgagt
     3601 ttaaacccgc tgatcagcct cgactgtgcc ttctagttgc cagccatctg ttgtttgccc
     3661 ctcccccgtg ccttccttga ccctggaagg tgccactccc actgtccttt cctaataaaa
     3721 tgaggaaatt gcatcgcatt gtctgagtag gtgtcattct attctggggg gtggggtggg
     3781 gcaggacagc aagggggagg attgggaaga caatagcagg catgctgggg atgcggtggg
     3841 ctctatggct tctgaggcgg aaagaaccag ctggggctct agggggtatc cccacgcgcc
     3901 ctgtagcggc gcattaagcg cggcgggtgt ggtggttacg cgcagcgtga ccgctacact
     3961 tgccagcgcc ctagcgcccg ctcctttcgc tttcttccct tcctttctcg ccacgttcgc
     4021 cggctttccc cgtcaagctc taaatcgggg gctcccttta gggttccgat ttagtgcttt
     4081 acggcacctc gaccccaaaa aacttgatta gggtgatggt tcacgtacct agaagttcct
     4141 attccgaagt tcctattctc tagaaagtat aggaacttcc ttggccgcca agcctttgtc
     4201 tcaagaagaa tccaccctca ttgaaagagc aacggctaca atcaacagca tccccatctc
     4261 tgaagactac agcgtcgcca gcgcagctct ctctagcgac ggccgcatct tcactggtgt
     4321 caatgtatat cattttactg ggggaccttg tgcagaactc gtggtgctgg gcactgctgc
     4381 tgctgcggca gctggcaacc tgacttgtat cgtcgcgatc ggaaatgaga acaggggcat
     4441 cttgagcccc tgcggacggt gccgacaggt gcttctcgat ctgcatcctg ggatcaaagc
     4501 catagtgaag gacagtgatg gacagccgac ggcagttggg attcgtgaat tgctgccctc
     4561 tggttatgtg tgggagggct aagcacaatt cgagctcggt acctttaaga ccaatgactt
     4621 acaaggcagc tgtagatctt agccaacttg tttattgcag cttaatggtt acaaataaag
     4681 caatagcatc acaaatttca caaataaagc atttttttca ctgcattcta gttgtggttt
     4741 gtccaaactc atcaatgtat cttatcatgt ctgtataccg tcgacctcta gctagagctt
     4801 ggcgtaatca tggtcatagc tgtttcctgt gtgaaattgt tatccgctca caattccaca
     4861 caacatacga gccggaagca taaagtgtaa agcctggggt gcctaatgag tgagctaact
     4921 cacattaatt gcgttgcgct cactgcccgc tttccagtcg ggaaacctgt cgtgccagct
     4981 gcattaatga atcggccaac gcgcggggag aggcggtttg cgtattgggc gctcttccgc
     5041 ttcctcgctc actgactcgc tgcgctcggt cgttcggctg cggcgagcgg tatcagctca
     5101 ctcaaaggcg gtaatacggt tatccacaga atcaggggat aacgcaggaa agaacatgtg
     5161 agcaaaaggc cagcaaaagg ccaggaaccg taaaaaggcc gcgttgctgg cgtttttcca
     5221 taggctccgc ccccctgacg agcatcacaa aaatcgacgc tcaagtcaga ggtggcgaaa
     5281 cccgacagga ctataaagat accaggcgtt tccccctgga agctccctcg tgcgctctcc
     5341 tgttccgacc ctgccgctta ccggatacct gtccgccttt ctcccttcgg gaagcgtggc
     5401 gctttctcat agctcacgct gtaggtatct cagttcggtg taggtcgttc gctccaagct
     5461 gggctgtgtg cacgaacccc ccgttcagcc cgaccgctgc gccttatccg gtaactatcg
     5521 tcttgagtcc aacccggtaa gacacgactt atcgccactg gcagcagcca ctggtaacag
     5581 gattagcaga gcgaggtatg taggcggtgc tacagagttc ttgaagtggt ggcctaacta
     5641 cggctacact agaaggacag tatttggtat ctgcgctctg ctgaagccag ttaccttcgg
     5701 aaaaagagtt ggtagctctt gatccggcaa acaaaccacc gctggtagcg gtggtttttt
     5761 tgtttgcaag cagcagatta cgcgcagaaa aaaaggatct caagaagatc ctttgatctt
     5821 ttctacgggg tctgacgctc agtggaacga aaactcacgt taagggattt tggtcatgag
     5881 attatcaaaa aggatcttca cctagatcct tttaaattaa aaatgaagtt ttaaatcaat
     5941 ctaaagtata tatgagtaaa cttggtctga cagttaccaa tgcttaatca gtgaggcacc
     6001 tatctcagcg atctgtctat ttcgttcatc catagttgcc tgactccccg tcgtgtagat
     6061 aactacgata cgggagggct taccatctgg ccccagtgct gcaatgatac cgcgagaccc
     6121 acgctcaccg gctccagatt tatcagcaat aaaccagcca gccggaaggg ccgagcgcag
     6181 aagtggtcct gcaactttat ccgcctccat ccagtctatt aattgttgcc gggaagctag
     6241 agtaagtagt tcgccagtta atagtttgcg caacgttgtt gccattgcta caggcatcgt
     6301 ggtgtcacgc tcgtcgtttg gtatggcttc attcagctcc ggttcccaac gatcaaggcg
     6361 agttacatga tcccccatgt tgtgcaaaaa agcggttagc tccttcggtc ctccgatcgt
     6421 tgtcagaagt aagttggccg cagtgttatc actcatggtt atggcagcac tgcataattc
     6481 tcttactgtc atgccatccg taagatgctt ttctgtgact ggtgagtact caaccaagtc
     6541 attctgagaa tagtgtatgc ggcgaccgag ttgctcttgc ccggcgtcaa tacgggataa
     6601 taccgcgcca catagcagaa ctttaaaagt gctcatcatt ggaaaacgtt cttcggggcg
     6661 aaaactctca aggatcttac cgctgttgag atccagttcg atgtaaccca ctcgtgcacc
     6721 caactgatct tcagcatctt ttactttcac cagcgtttct gggtgagcaa aaacaggaag
     6781 gcaaaatgcc gcaaaaaagg gaataagggc gacacggaaa tgttgaatac tcatactctt
     6841 cctttttcaa tattattgaa gcatttatca gggttattgt ctcatgagcg gatacatatt
     6901 tgaatgtatt tagaaaaata aacaaatagg ggttccgcgc acatttcccc gaaaagtgcc
     6961 acctgacgtc
//
