LOCUS       sequence-471826-        4959 bp    DNA     circular SYN 18-JUN-2025
DEFINITION  Express LEF response element in luciferase reporter assay.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4959)
  TITLE     Promega Early Access Collection
REFERENCE   2  (bases 1 to 4959)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Jun 18, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"
COMMENT     Sequence Label: pNL [NlucP/TCF/LEF-RE/Hygro] Vector
FEATURES             Location/Qualifiers
     source          1..4959
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        163..194
                     /label=minP
                     /note="minimal TATA-box promoter with low basal activity"
     regulatory      221..230
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             227..739
                     /codon_start=1
                     /product="NanoLuc(R) luciferase"
                     /label=Nluc
                     /note="human codon-optimized"
                     /translation="MVFTLEDFVGDWRQTAGYNLDQVLEQGGVSSLFQNLGVSVTPIQR
                     IVLSGENGLKIDIHVIIPYEGLSGDQMGQIEKIFKVVYPVDDHHFKVILHYGTLVIDGV
                     TPNMIDYFGRPYEGIAVFDGKKITVTGTLWNGNKIIDERLINPDGSLLFRVTINGVTGW
                     RLCERILA"
     CDS             743..862
                     /codon_start=1
                     /product="PEST degradation sequence from mouse ornithine 
                     decarboxylase"
                     /label=hPEST
                     /note="human codon-optimized"
                     /translation="SHGFPPEVEEQAAGTLPMSCAQESGMDRHPAACASARINV"
     polyA_signal    914..1035
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(925..944)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     979..998
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     promoter        1229..1586
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        1229..1420
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(1252..1272)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      1437..1572
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     1499..1518
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             1617..2654
                     /codon_start=1
                     /product="hygromycin B phosphotransferase"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="MKKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRG
                     YVLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLP
                     ETELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQ
                     TVMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMF
                     GDSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDG
                     NFDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAK
                     EVGRV"
     RBS             2633..2641
                     /label=Shine-Dalgarno sequence
                     /note="full consensus sequence for ribosome-binding sites 
                     upstream of start codons in E. coli; complementary to a 
                     region in the 3' end of the 16S rRNA (Chen et al., 1994)"
     polyA_signal    2678..2726
                     /note="synthetic polyadenylation signal"
     primer_bind     complement(2882..2899)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3053..3641)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3133..3152)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(3841..4632,4633..4701))
                     /codon_start=1
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     polyA_signal    4806..4854
                     /note="synthetic polyadenylation signal"
     misc_feature    4868..4959
                     /label=pause site
                     /note="RNA polymerase II transcriptional pause signal from 
                     the human alpha-2 globin gene"
     primer_bind     4908..4927
                     /label=RVprimer3
                     /note="pGL3 vector, forward primer"
ORIGIN
        1 ggcctaactg gccggtacct gagctcagat caaagggttt aagatcaaag ggcttaagat
       61 caaagggtat aagatcaaag ggcctaagat caaagggact aagatcaaag ggtttaagat
      121 caaagggctt aagatcaaag ggcctactcg aagcttagac actagagggt atataatgga
      181 agctcgactt ccagcttggc aatccggtac tgttggtaaa gccaccatgg tcttcacact
      241 cgaagatttc gttggggact ggcgacagac agccggctac aacctggacc aagtccttga
      301 acagggaggt gtgtccagtt tgtttcagaa tctcggggtg tccgtaactc cgatccaaag
      361 gattgtcctg agcggtgaaa atgggctgaa gatcgacatc catgtcatca tcccgtatga
      421 aggtctgagc ggcgaccaaa tgggccagat cgaaaaaatt tttaaggtgg tgtaccctgt
      481 ggatgatcat cactttaagg tgatcctgca ctatggcaca ctggtaatcg acggggttac
      541 gccgaacatg atcgactatt tcggacggcc gtatgaaggc atcgccgtgt tcgacggcaa
      601 aaagatcact gtaacaggga ccctgtggaa cggcaacaaa attatcgacg agcgcctgat
      661 caaccccgac ggctccctgc tgttccgagt aaccatcaac ggagtgaccg gctggcggct
      721 gtgcgaacgc attctggcga attctcacgg ctttccgcct gaggttgaag agcaagccgc
      781 cggtacattg cctatgtcct gcgcacaaga aagcggtatg gaccggcacc cagccgcttg
      841 tgcttcagct cgcatcaacg tctaaggccg cgactctaga gtcggggcgg ccggccgctt
      901 cgagcagaca tgataagata cattgatgag tttggacaaa ccacaactag aatgcagtga
      961 aaaaaatgct ttatttgtga aatttgtgat gctattgctt tatttgtaac cattataagc
     1021 tgcaataaac aagttaacaa caacaattgc attcatttta tgtttcaggt tcagggggag
     1081 gtgtgggagg ttttttaaag caagtaaaac ctctacaaat gtggtaaaat cgataaggat
     1141 ccgtttgcgt attgggcgct cttccgctga tctgcgcagc accatggcct gaaataacct
     1201 ctgaaagagg aacttggtta gctaccttct gaggcggaaa gaaccagctg tggaatgtgt
     1261 gtcagttagg gtgtggaaag tccccaggct ccccagcagg cagaagtatg caaagcatgc
     1321 atctcaatta gtcagcaacc aggtgtggaa agtccccagg ctccccagca ggcagaagta
     1381 tgcaaagcat gcatctcaat tagtcagcaa ccatagtccc gcccctaact ccgcccatcc
     1441 cgcccctaac tccgcccagt tccgcccatt ctccgcccca tggctgacta atttttttta
     1501 tttatgcaga ggccgaggcc gcctctgcct ctgagctatt ccagaagtag tgaggaggct
     1561 tttttggagg cctaggcttt tgcaaaaagc tcgattcttc tgacactagc gccaccatga
     1621 agaagcccga actcaccgct accagcgttg aaaaatttct catcgagaag ttcgacagtg
     1681 tgagcgacct gatgcagttg tcggagggcg aagagagccg agccttcagc ttcgatgtcg
     1741 gcggacgcgg ctatgtactg cgggtgaata gctgcgctga tggcttctac aaagaccgct
     1801 acgtgtaccg ccacttcgcc agcgctgcac tacccatccc cgaagtgttg gacatcggcg
     1861 agttcagcga gagcctgaca tactgcatca gtagacgcgc ccaaggcgtt actctccaag
     1921 acctccccga aacagagctg cctgctgtgt tacagcctgt cgccgaagct atggatgcta
     1981 ttgccgccgc cgacctcagt caaaccagcg gcttcggccc attcgggccc caaggcatcg
     2041 gccagtacac aacctggcgg gatttcattt gcgccattgc tgatccccat gtctaccact
     2101 ggcagaccgt gatggacgac accgtgtccg ccagcgtagc tcaagccctg gacgaactga
     2161 tgctgtgggc cgaagactgt cccgaggtgc gccacctcgt ccatgccgac ttcggcagca
     2221 acaacgtcct gaccgacaac ggccgcatca ccgccgtaat cgactggtcc gaagctatgt
     2281 tcggggacag tcagtacgag gtggccaaca tcttcttctg gcggccctgg ctggcttgca
     2341 tggagcagca gactcgctac ttcgagcgcc ggcatcccga gctggccggc agccctcgtc
     2401 tgcgagccta catgctgcgc atcggcctgg atcagctcta ccagagcctc gtggacggca
     2461 acttcgacga tgctgcctgg gctcaaggcc gctgcgatgc catcgtccgc agcggggccg
     2521 gcaccgtcgg tcgcacacaa atcgctcgcc ggagcgcagc cgtatggacc gacggctgcg
     2581 tcgaggtgct ggccgacagc ggcaaccgcc ggcccagtac acgaccgcgc gctaaggagg
     2641 taggtcgagt ttaaactcta gaaccggtca tggccgcaat aaaatatctt tattttcatt
     2701 acatctgtgt gttggttttt tgtgtgttcg aactagatgc tgtcgaccga tgcccttgag
     2761 agccttcaac ccagtcagct ccttccggtg ggcgcggggc atgactatcg tcgccgcact
     2821 tatgactgtc ttctttatca tgcaactcgt aggacaggtg ccggcagcgc tcttccgctt
     2881 cctcgctcac tgactcgctg cgctcggtcg ttcggctgcg gcgagcggta tcagctcact
     2941 caaaggcggt aatacggtta tccacagaat caggggataa cgcaggaaag aacatgtgag
     3001 caaaaggcca gcaaaaggcc aggaaccgta aaaaggccgc gttgctggcg tttttccata
     3061 ggctccgccc ccctgacgag catcacaaaa atcgacgctc aagtcagagg tggcgaaacc
     3121 cgacaggact ataaagatac caggcgtttc cccctggaag ctccctcgtg cgctctcctg
     3181 ttccgaccct gccgcttacc ggatacctgt ccgcctttct cccttcggga agcgtggcgc
     3241 tttctcatag ctcacgctgt aggtatctca gttcggtgta ggtcgttcgc tccaagctgg
     3301 gctgtgtgca cgaacccccc gttcagcccg accgctgcgc cttatccggt aactatcgtc
     3361 ttgagtccaa cccggtaaga cacgacttat cgccactggc agcagccact ggtaacagga
     3421 ttagcagagc gaggtatgta ggcggtgcta cagagttctt gaagtggtgg cctaactacg
     3481 gctacactag aagaacagta tttggtatct gcgctctgct gaagccagtt accttcggaa
     3541 aaagagttgg tagctcttga tccggcaaac aaaccaccgc tggtagcggt ggtttttttg
     3601 tttgcaagca gcagattacg cgcagaaaaa aaggatctca agaagatcct ttgatctttt
     3661 ctacggggtc tgacgctcag tggaacgaaa actcacgtta agggattttg gtcatgagat
     3721 tatcaaaaag gatcttcacc tagatccttt taaattaaaa atgaagtttt aaatcaatct
     3781 aaagtatata tgagtaaact tggtctgaca gcggccgcaa atgctaaacc actgcagtgg
     3841 ttaccagtgc ttgatcagtg aggcaccgat ctcagcgatc tgcctatttc gttcgtccat
     3901 agtggcctga ctccccgtcg tgtagatcac tacgattcgt gagggcttac catcaggccc
     3961 cagcgcagca atgatgccgc gagagccgcg ttcaccggcc cccgatttgt cagcaatgaa
     4021 ccagccagca gggagggccg agcgaagaag tggtcctgct actttgtccg cctccatcca
     4081 gtctatgagc tgctgtcgtg atgctagagt aagaagttcg ccagtgagta gtttccgaag
     4141 agttgtggcc attgctactg gcatcgtggt atcacgctcg tcgttcggta tggcttcgtt
     4201 caactctggt tcccagcggt caagccgggt cacatgatca cccatattat gaagaaatgc
     4261 agtcagctcc ttagggcctc cgatcgttgt cagaagtaag ttggccgcgg tgttgtcgct
     4321 catggtaatg gcagcactac acaattctct taccgtcatg ccatccgtaa gatgcttttc
     4381 cgtgaccggc gagtactcaa ccaagtcgtt ttgtgagtag tgtatacggc gaccaagctg
     4441 ctcttgcccg gcgtctatac gggacaacac cgcgccacat agcagtactt tgaaagtgct
     4501 catcatcggg aatcgttctt cggggcggaa agactcaagg atcttgccgc tattgagatc
     4561 cagttcgata tagcccactc ttgcacccag ttgatcttca gcatctttta ctttcaccag
     4621 cgtttcgggg tgtgcaaaaa caggcaagca aaatgccgca aagaagggaa tgagtgcgac
     4681 acgaaaatgt tggatgctca tactcgtcct ttttcaatat tattgaagca tttatcaggg
     4741 ttactagtac gtctctcaag gataagtaag taatattaag gtacgggagg tattggacag
     4801 gccgcaataa aatatcttta ttttcattac atctgtgtgt tggttttttg tgtgaatcga
     4861 tagtactaac atacgctctc catcaaaaca aaacgaaaca aaacaaacta gcaaaatagg
     4921 ctgtccccag tgcaagtgca ggtgccagaa catttctct
//
