LOCUS       sequence-314736-        2267 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Provides Middle-to-End linker 2 for level 2 cloning.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 2267)
  AUTHORS   Andreou , Andreas I., Nirkko, Jessica , Ochoa-Villarreal, Marisol , 
            Nakayama, Naomi
  TITLE     Mobius Assembly for Plant Systems highlights promoter-terminator 
            interaction in gene regulation
  JOURNAL   bioRxiv, March 31, 2021
REFERENCE   2  (bases 1 to 2267)
  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: "bioRxiv, 
            March 31, 2021"
COMMENT     Sequence Label: Auxiliary 2
FEATURES             Location/Qualifiers
     source          1..2267
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     terminator      3..60
                     /label=his operon terminator
                     /note="This putative transcriptin terminator from the E. 
                     coli his operon has a 2-bp deletion introduced during 
                     synthesis. Its efficiency has not been determined."
     primer_bind     complement(84..101)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(255..843)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(335..354)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     terminator      complement(1026..1120)
                     /label=lambda t0 terminator
                     /note="transcription terminator from phage lambda"
     CDS             complement(1141..1956)
                     /codon_start=1
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     CDS             complement(1141..1956)
                     /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="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     primer_bind     1864..1883
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     2118..2136
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     terminator      complement(2140..2183)
                     /label=bacterial terminator
                     /note="putative bacterial transcription terminator"
ORIGIN
        1 agtccggcaa aaaagggcaa ggtgtcacca ccctgccctt tttctttaaa accgaaaaga
       61 ttacttcgcg ttatgcaggc ttcctcgctc actgactcgc tgcgctcggt cgttcggctg
      121 cggcgagcgg tatcagctca ctcaaaggcg gtaatacggt tatccacaga atcaggggat
      181 aacgcaggaa agaacatgtg agcaaaaggc cagcaaaagg ccaggaaccg taaaaaggcc
      241 gcgttgctgg cgtttttcca caggctccgc ccccctgacg agcatcacaa aaatcgacgc
      301 tcaagtcaga ggtggcgaaa cccgacagga ctataaagat accaggcgtt tccccctgga
      361 agctccctcg tgcgctctcc tgttccgacc ctgccgctta ccggatacct gtccgccttt
      421 ctcccttcgg gaagcgtggc gctttctcat agctcacgct gtaggtatct cagttcggtg
      481 taggtcgttc gctccaagct gggctgtgtg cacgaacccc ccgttcagcc cgaccgctgc
      541 gccttatccg gtaactatcg tcttgagtcc aacccggtaa gacacgactt atcgccactg
      601 gcagcagcca ctggtaacag gattagcaga gcgaggtatg taggcggtgc tacagagttc
      661 ttgaagtggt ggcctaacta cggctacact agaagaacag tatttggtat ctgcgctctg
      721 ctgaagccag ttaccttcgg aaaaagagtt ggtagctctt gatccggcaa acaaaccacc
      781 gctggtagcg gtggtttttt tgtttgcaag cagcagatta cgcgcagaaa aaaaggatct
      841 caagaagatc ctttgatctt ttctacgggg tctgacgctc agtggaacga aaactcacgt
      901 taagggattt tggtcatgag attatcaaaa aggatcttca cctagatcct tttaaattaa
      961 aaatgaagtt ttaaatcaat ctaaagtata tatgagtaaa cttggtctga cagctcgagg
     1021 cttggattct caccaataaa aaacgcccgg cggcaaccga gcgttctgaa caaatccaga
     1081 tggagttctg aggtcattac tggatctatc aacaggagtc caagcgagct cgatatcaaa
     1141 ttagaaaaac tcatcgagca tcaaatgaaa ctgcaattta ttcatatcag gattatcaat
     1201 accatatttt tgaaaaagcc gtttctgtaa tgaaggagaa aactcaccga ggcagttcca
     1261 taggatggca agatcctggt atcggtctgc gattccgact cgtccaacat caatacaacc
     1321 tattaatttc ccctcgtcaa aaataaggtt atcaagtgag aaatcaccat gagtgacgac
     1381 tgaatccggt gagaatggca aaagtttatg catttctttc cagacttgtt caacaggcca
     1441 gccattacgc tcgtcatcaa aatcactcgc atcaaccaaa ccgttattca ttcgtgattg
     1501 cgcctgagcg agacgaaata cgcgatcgct gttaaaagga caattacaaa caggaatcga
     1561 atgcaaccgg cgcaggaaca ctgccagcgc atcaacaata ttttcacctg aatcaggata
     1621 ttcttctaat acctggaatg ctgttttccc tgggatcgca gtggtgagta accatgcatc
     1681 atcaggagta cggataaaat gcttgatggt cggaagaggc ataaattccg tcagccagtt
     1741 tagtctgacc atctcatctg taacatcatt ggcaacgcta cctttgccat gtttcagaaa
     1801 caactctggc gcatcgggct tcccatacaa tcggtagatt gtcgcacctg attgcccgac
     1861 attatcgcga gcccatttat acccatataa atcagcatcc atgttggaat ttaatcgcgg
     1921 ccttgagcaa gacgtttccc gttgaatatg gctcatttta gcttccttag ctcctgaaaa
     1981 tctcgataac tcaaaaaata cgcccggtag tgatcttatt tcattatggt gaaagttgga
     2041 acctcttacg tgcccgatca actcgagtgc cacctgacgt ctaagaaacc attattatca
     2101 tgacattaac ctataaaaat aggcgtatca cgaggcagaa tttcagataa aaaaaatcct
     2161 tagctttcgc taaggatgat ttctggaatt ccacctgcat atgtcacgct agagaccgta
     2221 tgaggtggat ggtgtgctgc gtggccaaac ccatatgcag gtgctgc
//
