LOCUS       sequence-73126-e        3331 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Middle entry vector with TA-cloning linker.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 3331)
  TITLE     pME-TA
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 3331)
  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: 
            "Unpublished"
COMMENT     Sequence Label: pME-TA
FEATURES             Location/Qualifiers
     source          1..3331
                     /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    569..668
                     /gene="mutant version of attL"
                     /label=attL1
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     CDS             join(702..704,705..707,708..1421)
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             join(702..704,705..707,708..1418)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(747..768)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(1008..1027)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1355..1376
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     protein_bind    complement(1456..1555)
                     /gene="mutant version of attL"
                     /label=attL2
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     primer_bind     complement(1572..1591)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(1573..1591)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(1596..1612)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1596..1612)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(1714..1733)
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     CDS             1725..2534
                     /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(1792..1811)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     rep_origin      2681..3269
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3170..3189
                     /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 tcgggcccca aataatgatt ttattttgac tgatagtgac
      601 ctgttcgttg caacacattg atgagcaatg cttttttata atgccaactt tgtacaaaaa
      661 agcaggctcc gccaccagta ccaatacttg tatgggatat catggtgagc aagggcgagg
      721 agctgttcac cggggtggtg cccatcctgg tcgagctgga cggcgacgta aacggccaca
      781 agttcagcgt gtccggcgag ggcgagggcg atgccaccta cggcaagctg accctgaagt
      841 tcatctgcac caccggcaag ctgcccgtgc cctggcccac cctcgtgacc accctgacct
      901 acggcgtgca gtgcttcagc cgctaccccg accacatgaa gcagcacgac ttcttcaagt
      961 ccgccatgcc cgaaggctac gtccaggagc gcaccatctt cttcaaggac gacggcaact
     1021 acaagacccg cgccgaggtg aagttcgagg gcgacaccct ggtgaaccgc atcgagctga
     1081 agggcatcga cttcaaggag gacggcaaca tcctggggca caagctggag tacaactaca
     1141 acagccacaa cgtctatatc atggccgaca agcagaagaa cggcatcaag gtgaacttca
     1201 agatccgcca caacatcgag gacggcagcg tgcagctcgc cgaccactac cagcagaaca
     1261 cccccatcgg cgacggcccc gtgctgctgc ccgacaacca ctacctgagc acccagtccg
     1321 ccctgagcaa agaccccaac gagaagcgcg atcacatggt cctgctggag ttcgtgaccg
     1381 ccgccgggat cactctcggc atggacgagc tgtacaagta accatacaag tattggctcg
     1441 aggatgggga ctgacaccca gctttcttgt acaaagttgg cattataaga aagcattgct
     1501 tatcaatttg ttgcaacgaa caggtcacta tcagtcaaaa taaaatcatt atttgccatc
     1561 cagctgatat cccctatagt gagtcgtatt acatggtcat agctgtttcc tggcagctct
     1621 ggcccgtgtc tcaaaatctc tgatgttaca ttgcacaaga taaaataata tcatcatgaa
     1681 caataaaact gtctgcttac ataaacagta atacaagggg tgttatgagc catattcaac
     1741 gggaaacgtc gaggccgcga ttaaattcca acatggatgc tgatttatat gggtataaat
     1801 gggctcgcga taatgtcggg caatcaggtg cgacaatcta tcgcttgtat gggaagcccg
     1861 atgcgccaga gttgtttctg aaacatggca aaggtagcgt tgccaatgat gttacagatg
     1921 agatggtcag actaaactgg ctgacggaat ttatgcctct tccgaccatc aagcatttta
     1981 tccgtactcc tgatgatgca tggttactca ccactgcgat ccccggaaaa acagcattcc
     2041 aggtattaga agaatatcct gattcaggtg aaaatattgt tgatgcgctg gcagtgttcc
     2101 tgcgccggtt gcattcgatt cctgtttgta attgtccttt taacagcgat cgcgtatttc
     2161 gtctcgctca ggcgcaatca cgaatgaata acggtttggt tgatgcgagt gattttgatg
     2221 acgagcgtaa tggctggcct gttgaacaag tctggaaaga aatgcataaa cttttgccat
     2281 tctcaccgga ttcagtcgtc actcatggtg atttctcact tgataacctt atttttgacg
     2341 aggggaaatt aataggttgt attgatgttg gacgagtcgg aatcgcagac cgataccagg
     2401 atcttgccat cctatggaac tgcctcggtg agttttctcc ttcattacag aaacggcttt
     2461 ttcaaaaata tggtattgat aatcctgata tgaataaatt gcagtttcat ttgatgctcg
     2521 atgagttttt ctaatcagaa ttggttaatt ggttgtaaca ctggcagagc attacgctga
     2581 cttgacggga cggcgcaagc tcatgaccaa aatcccttaa cgtgagttac gcgtcgttcc
     2641 actgagcgtc agaccccgta gaaaagatca aaggatcttc ttgagatcct ttttttctgc
     2701 gcgtaatctg ctgcttgcaa acaaaaaaac caccgctacc agcggtggtt tgtttgccgg
     2761 atcaagagct accaactctt tttccgaagg taactggctt cagcagagcg cagataccaa
     2821 atactgttct tctagtgtag ccgtagttag gccaccactt caagaactct gtagcaccgc
     2881 ctacatacct cgctctgcta atcctgttac cagtggctgc tgccagtggc gataagtcgt
     2941 gtcttaccgg gttggactca agacgatagt taccggataa ggcgcagcgg tcgggctgaa
     3001 cggggggttc gtgcacacag cccagcttgg agcgaacgac ctacaccgaa ctgagatacc
     3061 tacagcgtga gctatgagaa agcgccacgc ttcccgaagg gagaaaggcg gacaggtatc
     3121 cggtaagcgg cagggtcgga acaggagagc gcacgaggga gcttccaggg ggaaacgcct
     3181 ggtatcttta tagtcctgtc gggtttcgcc acctctgact tgagcgtcga tttttgtgat
     3241 gctcgtcagg ggggcggagc ctatggaaaa acgccagcaa cgcggccttt ttacggttcc
     3301 tggccttttg ctggcctttt gctcacatgt t
//
