LOCUS       sequence-330605-        3356 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Gateway attB4/attB1r entry clone for mammalian expression using the 
            human EF1alpha promoter.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 3356)
  AUTHORS   Wall VE, Garvey LA, Mehalko JL, Procter LV, Esposito D
  TITLE     Combinatorial assembly of clone libraries using site-specific 
            recombination.
  JOURNAL   Methods Mol Biol. 2014;1116:193-208. doi: 
            10.1007/978-1-62703-764-8_14.
  PUBMED    24395366
REFERENCE   2  (bases 1 to 3356)
  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: "Methods Mol
            Biol."; date: "2014"; volume: "1116:193-208. doi"; pages: " 
            10.1007/978-1-62703-764-8_14"
COMMENT     Sequence Label: C413-E19: attB4-mPol2p>-attB1r
FEATURES             Location/Qualifiers
     source          1..3356
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     23..45
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     37..54
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     38..54
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    94..189
                     /gene="mutant version of attL"
                     /label=attL4
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     protein_bind    903..1026
                     /gene="mutant version of attR"
                     /label=attR1
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     primer_bind     complement(1098..1117)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(1099..1117)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(1122..1138)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1122..1138)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(1240..1259)
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     CDS             1251..2060
                     /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(1318..1337)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     rep_origin      2207..2795
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     2696..2715
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     2949..2966
                     /label=L4440
                     /note="L4440 vector, forward primer"
     terminator      complement(3125..3152)
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB 
                     gene"
     terminator      complement(3244..3330)
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
ORIGIN
        1 ttaacgctag catggatgtt ttcccagtca cgacgttgta aaacgacggc cagtcttaag
       61 ctcgggcccg agttaacgct accatggagc tccaaataat gattttattt tgactgatag
      121 tgacctgttc gttgcaacaa attgataagc aatgcttttt tataatgcca actttgtata
      181 gaaaagttgg caagcttggg cttgaacatc gagcgccagg gctccgtaaa gctactagag
      241 cacaggcggt gccccaacgt cctggggcct ctccactaat aacggctact tccaattgat
      301 tggacgcgcc atcttgcctg ccttatgcat attcagcggt gaactgaata ttcatgaacg
      361 aggcccgtcc cgtccctccc tccttccccc cacccccgga acccgctccg gaggacccga
      421 agggccccgc cttcattacc gatgcgtagg acaaaccatt ttcccgatgt gtgtgggggg
      481 atactaatga gagactttag ctgaaaatga gcctgaactc cgaagctgag taaaaatggc
      541 ctaactttat cctccgttct gtaagtcctc ggtttgagtg cacgggaaac ccgaaaggag
      601 gacgacagga ccaggaacat tctcctcctc ctgtcgcgtc agaaagaaca cccaaccagg
      661 gagccggagc cctagcgtca acaactccgc cgcgcgcgct ccgtgtaggc cggtgcgggc
      721 ggccccgtag cgcaagggag ggcgggaaag gaaggggcgg gacacaaggg cgaatctata
      781 aagggcgtca ctcagccagt tctctcctca gaagcgccga gagcgcgacc gggacggttg
      841 gagaagaagg tggctcccgg aagggggaga gacaaactgc cgtaacctct gccgttcagg
      901 atcaactttg tacaaaaaag ttgaacgaga aacgtaaaat gatataaata tcaatatatt
      961 aaattagatt ttgcataaaa aacagactac ataatactgt aaaacacaac atatgcagtc
     1021 actatgaacc aactacttag atggtattag tgacctgtag aattcgagct ctagagctgc
     1081 agggcggccg cgatatcccc tatagtgagt cgtattacat ggtcatagct gtttcctggc
     1141 agctctggcc cgtgtctcaa aatctctgat gttacattgc acaagataaa aatatatcat
     1201 catgaacaat aaaactgtct gcttacataa acagtaatac aaggggtgtt atgagccata
     1261 ttcaacggga aacgtcgagg ccgcgattaa attccaacat ggatgctgat ttatatgggt
     1321 ataaatgggc tcgcgataat gtcgggcaat caggtgcgac aatctatcgc ttgtatggga
     1381 agcccgatgc gccagagttg tttctgaaac atggcaaagg tagcgttgcc aatgatgtta
     1441 cagatgagat ggtcagacta aactggctga cggaatttat gcctcttccg accatcaagc
     1501 attttatccg tactcctgat gatgcatggt tactcaccac tgcgatcccc ggaaaaacag
     1561 cattccaggt attagaagaa tatcctgatt caggtgaaaa tattgttgat gcgctggcag
     1621 tgttcctgcg ccggttgcat tcgattcctg tttgtaattg tccttttaac agcgatcgcg
     1681 tatttcgtct cgctcaggcg caatcacgaa tgaataacgg tttggttgat gcgagtgatt
     1741 ttgatgacga gcgtaatggc tggcctgttg aacaagtctg gaaagaaatg cataaacttt
     1801 tgccattctc accggattca gtcgtcactc atggtgattt ctcacttgat aaccttattt
     1861 ttgacgaggg gaaattaata ggttgtattg atgttggacg agtcggaatc gcagaccgat
     1921 accaggatct tgccatccta tggaactgcc tcggtgagtt ttctccttca ttacagaaac
     1981 ggctttttca aaaatatggt attgataatc ctgatatgaa taaattgcag tttcatttga
     2041 tgctcgatga gtttttctaa tcagaattgg ttaattggtt gtaacactgg cagagcatta
     2101 cgctgacttg acgggacggc gcaagctcat gaccaaaatc ccttaacgtg agttacgcgt
     2161 cgttccactg agcgtcagac cccgtagaaa agatcaaagg atcttcttga gatccttttt
     2221 ttctgcgcgt aatctgctgc ttgcaaacaa aaaaaccacc gctaccagcg gtggtttgtt
     2281 tgccggatca agagctacca actctttttc cgaaggtaac tggcttcagc agagcgcaga
     2341 taccaaatac tgttcttcta gtgtagccgt agttaggcca ccacttcaag aactctgtag
     2401 caccgcctac atacctcgct ctgctaatcc tgttaccagt ggctgctgcc agtggcgata
     2461 agtcgtgtct taccgggttg gactcaagac gatagttacc ggataaggcg cagcggtcgg
     2521 gctgaacggg gggttcgtgc acacagccca gcttggagcg aacgacctac accgaactga
     2581 gatacctaca gcgtgagcta tgagaaagcg ccacgcttcc cgaagggaga aaggcggaca
     2641 ggtatccggt aagcggcagg gtcggaacag gagagcgcac gagggagctt ccagggggaa
     2701 acgcctggta tctttatagt cctgtcgggt ttcgccacct ctgacttgag cgtcgatttt
     2761 tgtgatgctc gtcagggggg cggagcctat ggaaaaacgc cagcaacgcg gcctttttac
     2821 ggttcctggc cttttgctgg ccttttgctc acatgttctt tcctgcgtta tcccctgatt
     2881 ctgtggataa ccgtattacc gcctttgagt gagctgatac cgctcgccgc agccgaacga
     2941 ccgagcgcag cgagtcagtg agcgaggaag cggaagagcg cccaatacgc aaaccgcctc
     3001 tccccgcgcg ttggccgatt cattaatgca gctggcacga caggtttccc gactggaaag
     3061 cgggcagtga gcgcaacgca attaatacgc gtaccgctag ccaggaagag tttgtagaaa
     3121 cgcaaaaagg ccatccgtca ggatggcctt ctgcttagtt tgatgcctgg cagtttatgg
     3181 cgggcgtcct gcccgccacc ctccgggccg ttgcttcaca acgttcaaat ccgctcccgg
     3241 cggatttgtc ctactcagga gagcgttcac cgacaaacaa cagataaaac gaaaggccca
     3301 gtcttccgac tgagcctttc gttttatttg atgcctggca gttccctact ctcgcg
//
