LOCUS       sequence-251297-        3062 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  NanoLuc CDS middle entry gateway clone.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 3062)
  AUTHORS   Thierer JH, Ekker SC, Farber SA
  TITLE     The LipoGlo reporter system for sensitive and specific monitoring of
            atherogenic lipoproteins.
  JOURNAL   Nat Commun. 2019 Jul 31;10(1):3426. doi: 10.1038/s41467-019-11259-w.
  PUBMED    31366908
REFERENCE   2  (bases 1 to 3062)
  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: "Nat 
            Commun."; date: "2019-07-31"; volume: "10(1):3426. doi"; pages: " 
            10.1038/s41467-019-11259-w"
COMMENT     Sequence Label: pDONR_221_middle-entry_NanoLuc
FEATURES             Location/Qualifiers
     source          1..3062
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     84..101
                     /label=L4440
                     /note="L4440 vector, forward primer"
     terminator      complement(260..287)
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB 
                     gene"
     terminator      complement(379..465)
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     primer_bind     514..536
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     528..545
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     529..545
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    561..660
                     /gene="mutant version of attL"
                     /label=attL1
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     CDS             663..1178
                     /codon_start=1
                     /product="NanoLuc(R) luciferase"
                     /label=Nluc
                     /note="human codon-optimized"
                     /translation="MVFTLEDFVGDWRQTAGYNLDQVLEQGGVSSLFQNLGVSVTPIQR
                     IVLSGENGLKIDIHVIIPYEGLSGDQMGQIEKIFKVVYPVDDHHFKVILHYGTLVIDGV
                     TPNMIDYFGRPYEGIAVFDGKKITVTGTLWNGNKIIDERLINPDGSLLFRVTINGVTGW
                     RLCERILA"
     protein_bind    complement(1179..1278)
                     /gene="mutant version of attL"
                     /label=attL2
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     primer_bind     complement(1295..1314)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(1296..1314)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(1319..1335)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1319..1335)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(1437..1456)
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     CDS             1448..2257
                     /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(1515..1534)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     rep_origin      2404..2992
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     2893..2912
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 cgttatcccc tgattctgtg gataaccgta ttaccgcctt tgagtgagct gataccgctc
       61 gccgcagccg aacgaccgag cgcagcgagt cagtgagcga ggaagcggaa gagcgcccaa
      121 tacgcaaacc gcctctcccc gcgcgttggc cgattcatta atgcagctgg cacgacaggt
      181 ttcccgactg gaaagcgggc agtgagcgca acgcaattaa tacgcgtacc gctagccagg
      241 aagagtttgt agaaacgcaa aaaggccatc cgtcaggatg gccttctgct tagtttgatg
      301 cctggcagtt tatggcgggc gtcctgcccg ccaccctccg ggccgttgct tcacaacgtt
      361 caaatccgct cccggcggat ttgtcctact caggagagcg ttcaccgaca aacaacagat
      421 aaaacgaaag gcccagtctt ccgactgagc ctttcgtttt atttgatgcc tggcagttcc
      481 ctactctcgc gttaacgcta gcatggatgt tttcccagtc acgacgttgt aaaacgacgg
      541 ccagtcttaa gctcgggccc caaataatga ttttattttg actgatagtg acctgttcgt
      601 tgcaacaaat tgatgagcaa tgctttttta taatgccaac tttgtacaaa aaagcaggct
      661 tgatggtctt cacactcgaa gatttcgttg gggactggcg acagacagcc ggctacaacc
      721 tggaccaagt ccttgaacag ggaggtgtgt ccagtttgtt tcagaatctc ggggtgtccg
      781 taactccgat ccaaaggatt gtcctgagcg gtgaaaatgg gctgaagatc gacatccatg
      841 tcatcatccc gtatgaaggt ctgagcggcg accaaatggg ccagatcgaa aaaattttta
      901 aggtggtgta ccctgtggat gatcatcact ttaaggtgat cctgcactat ggcacactgg
      961 taatcgacgg ggttacgccg aacatgatcg actatttcgg acggccgtat gaaggcatcg
     1021 ccgtgttcga cggcaaaaag atcactgtaa cagggaccct gtggaacggc aacaaaatta
     1081 tcgacgagcg cctgatcaac cccgacggct ccctgctgtt ccgagtaacc atcaacggag
     1141 tgaccggctg gcggctgtgc gaacgcattc tggcgtaaac ccagctttct tgtacaaagt
     1201 tggcattata agaaagcatt gcttatcaat ttgttgcaac gaacaggtca ctatcagtca
     1261 aaataaaatc attatttgcc atccagctga tatcccctat agtgagtcgt attacatggt
     1321 catagctgtt tcctggcagc tctggcccgt gtctcaaaat ctctgatgtt acattgcaca
     1381 agataaaaat atatcatcat gaacaataaa actgtctgct tacataaaca gtaatacaag
     1441 gggtgttatg agccatattc aacgggaaac gtcgaggccg cgattaaatt ccaacatgga
     1501 tgctgattta tatgggtata aatgggctcg cgataatgtc gggcaatcag gtgcgacaat
     1561 ctatcgcttg tatgggaagc ccgatgcgcc agagttgttt ctgaaacatg gcaaaggtag
     1621 cgttgccaat gatgttacag atgagatggt cagactaaac tggctgacgg aatttatgcc
     1681 tcttccgacc atcaagcatt ttatccgtac tcctgatgat gcatggttac tcaccactgc
     1741 gatccccgga aaaacagcat tccaggtatt agaagaatat cctgattcag gtgaaaatat
     1801 tgttgatgcg ctggcagtgt tcctgcgccg gttgcattcg attcctgttt gtaattgtcc
     1861 ttttaacagc gatcgcgtat ttcgtctcgc tcaggcgcaa tcacgaatga ataacggttt
     1921 ggttgatgcg agtgattttg atgacgagcg taatggctgg cctgttgaac aagtctggaa
     1981 agaaatgcat aaacttttgc cattctcacc ggattcagtc gtcactcatg gtgatttctc
     2041 acttgataac cttatttttg acgaggggaa attaataggt tgtattgatg ttggacgagt
     2101 cggaatcgca gaccgatacc aggatcttgc catcctatgg aactgcctcg gtgagttttc
     2161 tccttcatta cagaaacggc tttttcaaaa atatggtatt gataatcctg atatgaataa
     2221 attgcagttt catttgatgc tcgatgagtt tttctaatca gaattggtta attggttgta
     2281 acactggcag agcattacgc tgacttgacg ggacggcgca agctcatgac caaaatccct
     2341 taacgtgagt tacgcgtcgt tccactgagc gtcagacccc gtagaaaaga tcaaaggatc
     2401 ttcttgagat cctttttttc tgcgcgtaat ctgctgcttg caaacaaaaa aaccaccgct
     2461 accagcggtg gtttgtttgc cggatcaaga gctaccaact ctttttccga aggtaactgg
     2521 cttcagcaga gcgcagatac caaatactgt tcttctagtg tagccgtagt taggccacca
     2581 cttcaagaac tctgtagcac cgcctacata cctcgctctg ctaatcctgt taccagtggc
     2641 tgctgccagt ggcgataagt cgtgtcttac cgggttggac tcaagacgat agttaccgga
     2701 taaggcgcag cggtcgggct gaacgggggg ttcgtgcaca cagcccagct tggagcgaac
     2761 gacctacacc gaactgagat acctacagcg tgagctatga gaaagcgcca cgcttcccga
     2821 agggagaaag gcggacaggt atccggtaag cggcagggtc ggaacaggag agcgcacgag
     2881 ggagcttcca gggggaaacg cctggtatct ttatagtcct gtcgggtttc gccacctctg
     2941 acttgagcgt cgatttttgt gatgctcgtc aggggggcgg agcctatgga aaaacgccag
     3001 caacgcggcc tttttacggt tcctggcctt ttgctggcct tttgctcaca tgttctttcc
     3061 tg
//
