LOCUS       sequence-161169-        5634 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  5' entry vector containing the EF1a/B-globin fused to zebrafish 
            b-actin 2 enhancer/promoter for strong semi-ubiquitous expression.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5634)
  AUTHORS   Fowler DK, Stewart S, Seredick S, Eisen JS, Stankunas K, Washbourne 
            P
  TITLE     A MultiSite Gateway Toolkit for Rapid Cloning of Vertebrate 
            Expression Constructs with Diverse Research Applications.
  JOURNAL   PLoS One. 2016 Aug 8;11(8):e0159277. doi: 
            10.1371/journal.pone.0159277. eCollection 2016.
  PUBMED    27500400
REFERENCE   2  (bases 1 to 5634)
  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: "PLoS One.";
            date: "2016-08-8"; volume: "11(8):e0159277. doi"; pages: " 
            10.1371/journal.pone.0159277. eCollection 2016"
COMMENT     Sequence Label: p5E-EF1a/b-actin
FEATURES             Location/Qualifiers
     source          1..5634
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     92..109
                     /label=L4440
                     /note="L4440 vector, forward primer"
     terminator      complement(268..295)
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB 
                     gene"
     terminator      complement(387..473)
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     primer_bind     522..544
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     536..553
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     537..553
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    593..688
                     /gene="mutant version of attL"
                     /label=attL4
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     promoter        690..708
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     intron          2974..3546
                     /label=beta-globin intron
                     /note="intron from rabbit beta-globin gene"
     primer_bind     complement(3608..3624)
                     /label=SK primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3608..3624)
                     /label=pBluescriptSK
                     /note="For pBluescript vector"
     promoter        complement(3661..3679)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     protein_bind    3680..3803
                     /gene="mutant version of attR"
                     /label=attR1
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     promoter        complement(3876..3894)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(3899..3915)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3899..3915)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4017..4036)
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     CDS             4028..4837
                     /codon_start=1
                     /gene="aph(3')-Ia"
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYGKP
                     DAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGKTA
                     FQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDASD
                     FDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGIAD
                     RYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     primer_bind     complement(4095..4114)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     rep_origin      4984..5572
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5473..5492
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 ctttcctgcg ttatcccctg attctgtgga taaccgtatt accgcctttg agtgagctga
       61 taccgctcgc cgcagccgaa cgaccgagcg cagcgagtca gtgagcgagg aagcggaaga
      121 gcgcccaata cgcaaaccgc ctctccccgc gcgttggccg attcattaat gcagctggca
      181 cgacaggttt cccgactgga aagcgggcag tgagcgcaac gcaattaata cgcgtaccgc
      241 tagccaggaa gagtttgtag aaacgcaaaa aggccatccg tcaggatggc cttctgctta
      301 gtttgatgcc tggcagttta tggcgggcgt cctgcccgcc accctccggg ccgttgcttc
      361 acaacgttca aatccgctcc cggcggattt gtcctactca ggagagcgtt caccgacaaa
      421 caacagataa aacgaaaggc ccagtcttcc gactgagcct ttcgttttat ttgatgcctg
      481 gcagttccct actctcgcgt taacgctagc atggatgttt tcccagtcac gacgttgtaa
      541 aacgacggcc agtcttaagc tcgggcccgc gttaacgcta ccatggagct ccaaataatg
      601 attttatttt gactgatagt gacctgttcg ttgcaacaaa ttgataagca atgctttttt
      661 ataatgccaa ctttgtatag aaaagttggt aatacgactc actatagggc gaattgggta
      721 ccgggccccc cctcgaggtc gagcaggggg atcatctaat caagcacaaa taaggggcgt
      781 gtaacacaaa agccagcgac cctttccaat gcaaatcaaa cttgcaattc tttgccgttt
      841 ttatcattta agtgtcggct taaggtccac tatcagatgt aaacagcctt atctaacaaa
      901 ggtatcatta cattctgaaa ttctcaggca tgcaagctag cttatgacgc actagggagt
      961 gccacccttc ctttcgccct aacttcgtga taactcgcgc gtttcactca acagctgcat
     1021 ccgccctagt gctactggga gttgtagtat acaagacgct tacaggctga atgttctgtc
     1081 aagaccccgc ctctagcact ttgggaattc tggacttgat gatgtcatgg ttaatccccg
     1141 cccagtagag gcggctatat aaagggtggt taaggcccgg ttcgctctct tcctcaccgg
     1201 gtctgcggcg agttctagct gaagcttcct gcaggtcgac aaagcaagtc tctcacataa
     1261 tatatcttct aaaagacaga aaacattact ttcagaccat attgtaatca aatgaacact
     1321 ttcatattag tcaataatat tacggaaatg tatttaaaaa ctgaataaac atatatttac
     1381 acctttgtct ttatttctag agtgctttta caatgtagat tgtgtcaaag cagcttaact
     1441 tagaggatta gtatagtaaa ttgaaactgt gtcagtccat tttttaagag tttaagttca
     1501 gttcagttta gttcagtgtg agttaatttt tactgctgag agtccaaaca ctaaagagca
     1561 aatccaaata cacaagtcaa taaataaact caatgatggg ctaaagatct ggggaaatct
     1621 gcagatttct gtgcgcacag attctgtgtg ggcctagtca ttacctatac tcaacgttta
     1681 ctctgagaaa cttccatatt ttagaggtaa aatgctcaga attgtaaaaa gtcttgtcaa
     1741 aatacttgga atttgccaaa aacaaaataa aagtcacatc acctcattgt ccttggtatc
     1801 agcaagcact tttaacacaa tcagctgttt ttgtttataa ctcccactga aggccatgtg
     1861 cattactccc aagttacaca gtaaaaagtc acatgtagtt tatcatgatc actgctgctc
     1921 aagcacagct ttatctgtta ttcagtgtgg caatatctga aatgtgtgca aaatattatt
     1981 tattgtagct gatcaccaac actttcagtg tttttttttt ctacttttaa tgagaattat
     2041 gttcttagga tttatcctta taaacctctt taaaaagaaa tgttatgtct gcatagacac
     2101 gacatgaaat tatgaacaga aaatttaact atttaagatc aagacttgac atataacatt
     2161 tctaatgaac attaaacctg aggtaggccc acttgcaaaa atgtaactgc tatcactcac
     2221 tttatacatc attgtgtaac ttgtacacac aaaaatagtg aagcaatgtg gcatggaatg
     2281 caggtcccat tcattcaaag ttattaaagc actttcttaa agtgcattac tactgagcta
     2341 agaccagcta gttgctcata caacatacta caagtagtgt catcctaatt tggataaaaa
     2401 aaatgtccat tttatacatg gaaaagtact cattagatcc cccattgttt gtattacggt
     2461 atttcgtgaa cacaagaggt aaatgaccta cagagctgct gttgtgttag attgaaaaca
     2521 caacacagga tcatggaggc atccatcatt atgacgcatt ctgccacttg taactctgca
     2581 cagacttttt gaaaagttta attgagtctt ggcgttgtcg cagaaaaata cgaaaaattc
     2641 tgtgttaaaa tttgcaaata aaaattccag ttttatagga aatcgccatg gccttgttct
     2701 ttaaccagtt aagcttcccc ttctttcact ctcaagttgc aagaagcaag tgtagcaatg
     2761 tgcacgcgac agccgggtgt gtgacgctgg accaatcaga gcacagagct ccgaaagttt
     2821 accttttatg gctagagccg ggtatgtgcc gtcatataaa agagcgcgcc cagcttttca
     2881 gcctcacttt gagctcctcc acacgcagct agtgcggaat atcatctgct tgtaacccat
     2941 tctcttaagt cgaccgatcc tgagaacttc agggtgagtt tggggaccct tgattgttct
     3001 ttctttttcg ctattgtaaa attcatgtta tatggagggg gcaaagtttt cagggtgttg
     3061 tttagaatgg gaagatgtcc cttgtatcac catggaccct catgataatt ttgtttcttt
     3121 cactttctac tctgttgaca accattgtct cctcttattt tcttttcatt ttctgtaact
     3181 ttttcgttaa actttagctt gcatttgtaa cgaattttta aattcacttt tgtttatttg
     3241 tcagattgta agtactttct ctaatcactt ttttttcaag gcaatcaggg tatattatat
     3301 tgtacttcag cacagtttta gagaacaatt gttataatta aatgataagg tagaatattt
     3361 ctgcatataa attctggctg gcgtggaaat attcttattg gtagaaacaa ctacaccctg
     3421 gtcatcatcc tgcctttctc tttatggtta caatgatata cactgtttga gatgaggata
     3481 aaatactctg agtccaaacc gggcccctct gctaaccatg ttcatgcctt cttctctttc
     3541 ctacagctcc tgggcaacgt gctggttgtt gtgctgtctc atcattttgg caaagaattc
     3601 ctcgacggat ccactagttc tagagcggcc gccaccgcgg tggagctcca gcttttgttc
     3661 cctttagtga gggttaattc aagtttgtac aaaaaagttg aacgagaaac gtaaaatgat
     3721 ataaatatca atatattaaa ttagattttg cataaaaaac agactacata atactgtaaa
     3781 acacaacata tgcagtcact atgaaccaac tacttagatg gtattagtga cctgtagaat
     3841 tcgagctcta gagctgcagg gcggccgcga tatcccctat agtgagtcgt attacatggt
     3901 catagctgtt tcctggcagc tctggcccgt gtctcaaaat ctctgatgtt acattgcaca
     3961 agataaaaat atatcatcat gaacaataaa actgtctgct tacataaaca gtaatacaag
     4021 gggtgttatg agccatattc aacgggaaac gtcgaggccg cgattaaatt ccaacatgga
     4081 tgctgattta tatgggtata aatgggctcg cgataatgtc gggcaatcag gtgcgacaat
     4141 ctatcgcttg tatgggaagc ccgatgcgcc agagttgttt ctgaaacatg gcaaaggtag
     4201 cgttgccaat gatgttacag atgagatggt cagactaaac tggctgacgg aatttatgcc
     4261 tcttccgacc atcaagcatt ttatccgtac tcctgatgat gcatggttac tcaccactgc
     4321 gatccccgga aaaacagcat tccaggtatt agaagaatat cctgattcag gtgaaaatat
     4381 tgttgatgcg ctggcagtgt tcctgcgccg gttgcattcg attcctgttt gtaattgtcc
     4441 ttttaacagc gatcgcgtat ttcgtctcgc tcaggcgcaa tcacgaatga ataacggttt
     4501 ggttgatgcg agtgattttg atgacgagcg taatggctgg cctgttgaac aagtctggaa
     4561 agaaatgcat aaacttttgc cattctcacc ggattcagtc gtcactcatg gtgatttctc
     4621 acttgataac cttatttttg acgaggggaa attaataggt tgtattgatg ttggacgagt
     4681 cggaatcgca gaccgatacc aggatcttgc catcctatgg aactgcctcg gtgagttttc
     4741 tccttcatta cagaaacggc tttttcaaaa atatggtatt gataatcctg atatgaataa
     4801 attgcagttt catttgatgc tcgatgagtt tttctaatca gaattggtta attggttgta
     4861 acactggcag agcattacgc tgacttgacg ggacggcgca agctcatgac caaaatccct
     4921 taacgtgagt tacgcgtcgt tccactgagc gtcagacccc gtagaaaaga tcaaaggatc
     4981 ttcttgagat cctttttttc tgcgcgtaat ctgctgcttg caaacaaaaa aaccaccgct
     5041 accagcggtg gtttgtttgc cggatcaaga gctaccaact ctttttccga aggtaactgg
     5101 cttcagcaga gcgcagatac caaatactgt ccttctagtg tagccgtagt taggccacca
     5161 cttcaagaac tctgtagcac cgcctacata cctcgctctg ctaatcctgt taccagtggc
     5221 tgctgccagt ggcgataagt cgtgtcttac cgggttggac tcaagacgat agttaccgga
     5281 taaggcgcag cggtcgggct gaacgggggg ttcgtgcaca cagcccagct tggagcgaac
     5341 gacctacacc gaactgagat acctacagcg tgagcattga gaaagcgcca cgcttcccga
     5401 agggagaaag gcggacaggt atccggtaag cggcagggtc ggaacaggag agcgcacgag
     5461 ggagcttcca gggggaaacg cctggtatct ttatagtcct gtcgggtttc gccacctctg
     5521 acttgagcgt cgatttttgt gatgctcgtc aggggggcgg agcctatgga aaaacgccag
     5581 caacgcggcc tttttacggt tcctggcctt ttgctggcct tttgctcaca tgtt
//
