LOCUS       sequence-447208-        3068 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Gateway compatible 5' entry clone with human alpha 2 globin pause 
            site (to prevent transcription readthrough) and EF1a core promoter. 
            Lacks downstream intron for a lower-activity, compact promoter..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 3068)
  AUTHORS   Gillespie WB 3rd, Zhang Y, Ruiz OE, Cerda J 3rd, Ortiz-Guzman J, 
            Sherman M, Turner WD, Largoza G, Mosser LE, Fujimoto E, Chien CB, 
            Kwan KM, Arenkiel BR, Devine WP, Wythe JD
  TITLE     MultiSite Assembly of Gateway Induced Clones (MAGIC): a flexible 
            cloning toolbox for use in vertebrate model systems.
  JOURNAL   Development. 2025 Nov 15;152(22):dev204308. doi: 10.1242/dev.204308.
            Epub 2025 Nov 13.
  PUBMED    41230856
REFERENCE   2  (bases 1 to 3068)
  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: 
            "Development."; date: "2025-11-15"; volume: "152(22):dev204308. 
            doi"; pages: " 10.1242/dev.204308. Epub 2025 Nov 13"
COMMENT     Sequence Label: p5E-EFS (EF1a core) (JDW 1319)
FEATURES             Location/Qualifiers
     source          1..3068
                     /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"
     misc_feature    720..811
                     /label=pause site
                     /note="RNA polymerase II transcriptional pause signal from 
                     the human alpha-2 globin gene"
     primer_bind     760..779
                     /label=RVprimer3
                     /note="pGL3 vector, forward primer"
     promoter        838..1049
                     /label=EF-1-alpha core promoter
                     /note="core promoter for human elongation factor 
                     EF-1-alpha"
     protein_bind    1114..1237
                     /gene="mutant version of attR"
                     /label=attR1
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     primer_bind     complement(1309..1328)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(1310..1328)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(1333..1349)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1333..1349)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(1451..1470)
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     CDS             1462..2271
                     /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(1529..1548)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     rep_origin      2418..3006
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     2907..2926
                     /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 tcgggcccga gttaacgcta ccatggagct ccaaataatg
      601 attttatttt gactgatagt gacctgttcg ttgcaacaaa ttgataagca atgctttttt
      661 ataatgccaa ctttgtatag aaaagttgcc gaaaaacgcg tgaattccca tggaagctta
      721 acatacgctc tccatcaaaa caaaacgaaa caaaacaaac tagcaaaata ggctgtcccc
      781 agtgcaagtg caggtgccag aacatttctc tatcgattta attaagatat cactagtggg
      841 cagagcgcac atcgcccaca gtccccgaga agttgggggg aggggtcggc aattgaaccg
      901 gtgcctagag aaggtggcgc ggggtaaact gggaaagtga tgtcgtgtac tggctccgcc
      961 tttttcccga gggtggggga gaaccgtata taagtgcagt agtcgccgtg aacgttcttt
     1021 ttcgcaacgg gtttgccgcc agaacacagc agctggtcta gagtcgacgg atccgctagc
     1081 gcggccgcgg taccttttct actcgagcta ggtcaagttt gtacaaaaaa gttgaacgag
     1141 aaacgtaaaa tgatataaat atcaatatat taaattagat tttgcataaa aaacagacta
     1201 cataatactg taaaacacaa catatgcagt cactatgaac caactactta gatggtatta
     1261 gtgacctgta gaattcgagc tctagagctg cagggcggcc gcgatatccc ctatagtgag
     1321 tcgtattaca tggtcatagc tgtttcctgg cagctctggc ccgtgtctca aaatctctga
     1381 tgttacattg cacaagataa aaatatatca tcatgaacaa taaaactgtc tgcttacata
     1441 aacagtaata caaggggtgt tatgagccat attcaacggg aaacgtcgag gccgcgatta
     1501 aattccaaca tggatgctga tttatatggg tataaatggg ctcgcgataa tgtcgggcaa
     1561 tcaggtgcga caatctatcg cttgtatggg aagcccgatg cgccagagtt gtttctgaaa
     1621 catggcaaag gtagcgttgc caatgatgtt acagatgaga tggtcagact aaactggctg
     1681 acggaattta tgcctcttcc gaccatcaag cattttatcc gtactcctga tgatgcatgg
     1741 ttactcacca ctgcgatccc cggaaaaaca gcattccagg tattagaaga atatcctgat
     1801 tcaggtgaaa atattgttga tgcgctggca gtgttcctgc gccggttgca ttcgattcct
     1861 gtttgtaatt gtccttttaa cagcgatcgc gtatttcgtc tcgctcaggc gcaatcacga
     1921 atgaataacg gtttggttga tgcgagtgat tttgatgacg agcgtaatgg ctggcctgtt
     1981 gaacaagtct ggaaagaaat gcataaactt ttgccattct caccggattc agtcgtcact
     2041 catggtgatt tctcacttga taaccttatt tttgacgagg ggaaattaat aggttgtatt
     2101 gatgttggac gagtcggaat cgcagaccga taccaggatc ttgccatcct atggaactgc
     2161 ctcggtgagt tttctccttc attacagaaa cggctttttc aaaaatatgg tattgataat
     2221 cctgatatga ataaattgca gtttcatttg atgctcgatg agtttttcta atcagaattg
     2281 gttaattggt tgtaacactg gcagagcatt acgctgactt gacgggacgg cgcaagctca
     2341 tgaccaaaat cccttaacgt gagttacgcg tcgttccact gagcgtcaga ccccgtagaa
     2401 aagatcaaag gatcttcttg agatcctttt tttctgcgcg taatctgctg cttgcaaaca
     2461 aaaaaaccac cgctaccagc ggtggtttgt ttgccggatc aagagctacc aactcttttt
     2521 ccgaaggtaa ctggcttcag cagagcgcag ataccaaata ctgttcttct agtgtagccg
     2581 tagttaggcc accacttcaa gaactctgta gcaccgccta catacctcgc tctgctaatc
     2641 ctgttaccag tggctgctgc cagtggcgat aagtcgtgtc ttaccgggtt ggactcaaga
     2701 cgatagttac cggataaggc gcagcggtcg ggctgaacgg ggggttcgtg cacacagccc
     2761 agcttggagc gaacgaccta caccgaactg agatacctac agcgtgagct atgagaaagc
     2821 gccacgcttc ccgaagggag aaaggcggac aggtatccgg taagcggcag ggtcggaaca
     2881 ggagagcgca cgagggagct tccaggggga aacgcctggt atctttatag tcctgtcggg
     2941 tttcgccacc tctgacttga gcgtcgattt ttgtgatgct cgtcaggggg gcggagccta
     3001 tggaaaaacg ccagcaacgc ggccttttta cggttcctgg ccttttgctg gccttttgct
     3061 cacatgtt
//
