LOCUS       sequence-390641-        7045 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Part of Renilla reporter system, encodes Firefly luciferase.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7045)
  AUTHORS   Luo EC, Nathanson JL, Tan FE, Schwartz JL, Schmok JC, Shankar A, 
            Markmiller S, Yee BA, Sathe S, Pratt GA, Scaletta DB, Ha Y, Hill DE,
            Aigner S, Yeo GW
  TITLE     Large-scale tethered function assays identify factors that regulate 
            mRNA stability and translation.
  JOURNAL   Nat Struct Mol Biol. 2020 Oct;27(10):989-1000. doi: 
            10.1038/s41594-020-0477-6. Epub 2020 Aug 17.
  PUBMED    32807991
REFERENCE   2  (bases 1 to 7045)
  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"; doi: 
            "10.1038/s41594-020-0477-6"; journalName: "Nat Struct Mol Biol"; 
            date: "2020-10"; volume: "27"; issue: "10"; pages: "989-1000"
COMMENT     Sequence Label: pTET2-lucCP
FEATURES             Location/Qualifiers
     source          1..7045
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(44..63)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     236..256
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        238..256
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     protein_bind    join(298..316,317..339,340..358,359..381,382..400,401..423,
                     424..442,443..465,466..484,485..507,508..526,527..549,
                     550..568)
                     /label=tetracycline response element
                     /bound_moiety="tetracycline repressor TetR"
                     /note="contains seven copies of the tetracycline operator 
                     tetO"
     promoter        601..639
                     /label=minimal CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     636..660
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     regulatory      691..700
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             697..2346
                     /codon_start=1
                     /gene="luc+"
                     /product="firefly luciferase"
                     /label=luciferase
                     /note="enhanced luc+ version of the luciferase gene"
                     /db_xref="GI:42718124"
                     /protein_id="AAS38485.1"
                     /translation="MADAKNIKKGPAPFYPLEDGTAGEQLHKAMKRYALVPGTIAFTDA
                     HIEVDITYAEYFEMSVRLAEAMKRYGLNTNHRIVVCSENSLQFFMPVLGALFIGVAVAP
                     ANDIYNERELLNSMGISQPTVVFVSKKGLQKILNVQKKLPIIQKIIIMDSKTDYQGFQS
                     MYTFVTSHLPPGFNEYDFVPESFDRDKTIALIMNSSGSTGLPKGVALPHRTACVRFSHA
                     RDPIFGNQIIPDTAILSVVPFHHGFGMFTTLGYLICGFRVVLMYRFEEELFLRSLQDYK
                     IQSALLVPTLFSFFAKSTLIDKYDLSNLHEIASGGAPLSKEVGEAVAKRFHLPGIRQGY
                     GLTETTSAILITPEGDDKPGAVGKVVPFFEAKVVDLDTGKTLGVNQRGELCVRGPMIMS
                     GYVNNPEATNALIDKDGWLHSGDIAYWDEDEHFFIVDRLKSLIKYKGYQVAPAELESIL
                     LQHPNIFDAGVAGLPDDDAGELPAAVVVLEHGKTMTEKEIVDYVASQVTTAKKLRGGVV
                     FVDEVPKGLTGKLDARKIREILIKAKKGGKIAV"
     RBS             1656..1664
                     /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)"
     CDS             2353..2400
                     /codon_start=1
                     /product="non-ORF yeast peptide conferring 
                     ubiquitin-dependent degradation"
                     /label=hCL1
                     /note="human codon-optimized"
                     /translation="ACKNWFSSLSHFVIHL"
     CDS             2404..2523
                     /codon_start=1
                     /product="PEST degradation sequence from mouse ornithine 
                     decarboxylase"
                     /label=hPEST
                     /note="human codon-optimized"
                     /translation="SHGFPPEVEEQAAGTLPMSCAQESGMDRHPAACASARINV"
     primer_bind     complement(2636..2653)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    2642..2866
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      2912..3340
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2999..3018)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3209..3230
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(3349..3369)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        3354..3683
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      3534..3669
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     3596..3615
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             3750..4544
                     /codon_start=1
                     /gene="aph(3')-II (or nptII)"
                     /product="aminoglycoside phosphotransferase from Tn5"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(3804..3823)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4414..4433
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    4718..4839
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4755..4774)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4809..4828
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(4888..4904)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4888..4904)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4901..4923)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4912..4928
                     /label=lac operator
                     /bound_moiety="lac repressor encoded by lacI"
                     /note="The lac repressor binds to the lac operator to 
                     inhibit transcription in E. coli. This inhibition can be 
                     relieved by adding lactose or 
                     isopropyl-beta-D-thiogalactopyranoside (IPTG)."
     promoter        complement(join(4936..4942,4943..4960,4961..4966))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4981..5002
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5119..5136)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5290..5878)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5370..5389)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6049..6840,6841..6909))
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     6672..6691
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6910..7014)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 gacggatcgg gagatctccc gatcccctat ggtgcactct cagtacaatc tgctctgatg
       61 ccgcatagtt aagccagtat ctgctccctg cttgtgtgtt ggaggtcgct gagtagtgcg
      121 cgagcaaaat ttaagctaca acaaggcaag gcttgaccga caattgcatg aagaatctgc
      181 ttagggttag gcgttttgcg ctgcttcgcg atgtacgggc cagatatacg cgtgcgcaat
      241 taaccctcac taaagggaac aaaagctggg taccgggccc cccctcgagt ttaccactcc
      301 ctatcagtga tagagaaaag tgaaagtcga gtttaccact ccctatcagt gatagagaaa
      361 agtgaaagtc gagtttacca ctccctatca gtgatagaga aaagtgaaag tcgagtttac
      421 cactccctat cagtgataga gaaaagtgaa agtcgagttt accactccct atcagtgata
      481 gagaaaagtg aaagtcgagt ttaccactcc ctatcagtga tagagaaaag tgaaagtcga
      541 gtttaccact ccctatcagt gatagagaaa agtgaaagtc gagctcggta cccgggtcga
      601 ggtaggcgtg tacggtggga ggcctatata agcagagctc gtttagtgaa ccgtcagatc
      661 gcctggagac gccatccacg ctgaagctta gccaccatgg ccgatgctaa gaacattaag
      721 aagggccctg ctcccttcta ccctctggag gatggcaccg ctggcgagca gctgcacaag
      781 gccatgaaga ggtatgccct ggtgcctggc accattgcct tcaccgatgc ccacattgag
      841 gtggacatca cctatgccga gtacttcgag atgtctgtgc gcctggccga ggccatgaag
      901 aggtacggcc tgaacaccaa ccaccgcatc gtggtgtgct ctgagaactc tctgcagttc
      961 ttcatgccag tgctgggcgc cctgttcatc ggagtggccg tggcccctgc taacgacatt
     1021 tacaacgagc gcgagctgct gaacagcatg ggcatttctc agcctaccgt ggtgttcgtg
     1081 tctaagaagg gcctgcagaa gatcctgaac gtgcagaaga agctgcctat catccagaag
     1141 atcatcatca tggactctaa gaccgactac cagggcttcc agagcatgta cacattcgtg
     1201 acatctcatc tgcctcctgg cttcaacgag tacgacttcg tgccagagtc tttcgacagg
     1261 gacaaaacca ttgccctgat catgaacagc tctgggtcta ccggcctgcc taagggcgtg
     1321 gccctgcctc atcgcaccgc ctgtgtgcgc ttctctcacg cccgcgaccc tattttcggc
     1381 aaccagatca tccccgacac cgctattctg agcgtggtgc cattccacca cggcttcggc
     1441 atgttcacca ccctgggcta cctgatttgc ggctttcggg tggtgctgat gtaccgcttc
     1501 gaggaggagc tgttcctgcg cagcctgcaa gactacaaaa ttcagtctgc cctgctggtg
     1561 ccaaccctgt tcagcttctt cgctaagagc accctgatcg acaagtacga cctgtctaac
     1621 ctgcacgaga ttgcctctgg cggcgcccca ctgtctaagg aggtgggcga agccgtggcc
     1681 aagcgctttc atctgccagg catccgccag ggctacggcc tgaccgagac aaccagcgcc
     1741 attctgatta ccccagaggg cgacgacaag cctggcgccg tgggcaaggt ggtgccattc
     1801 ttcgaggcca aggtggtgga cctggacacc ggcaagaccc tgggagtgaa ccagcgcggc
     1861 gagctgtgtg tgcgcggccc tatgattatg tccggctacg tgaataaccc tgaggccaca
     1921 aacgccctga tcgacaagga cggctggctg cactctggcg acattgccta ctgggacgag
     1981 gacgagcact tcttcatcgt ggaccgcctg aagtctctga tcaagtacaa gggctaccag
     2041 gtggccccag ccgagctgga gtctatcctg ctgcagcacc ctaacatttt cgacgccgga
     2101 gtggccggcc tgcccgacga cgatgccggc gagctgcctg ccgccgtcgt cgtgctggaa
     2161 cacggcaaga ccatgaccga gaaggagatc gtggactatg tggccagcca ggtgacaacc
     2221 gccaagaagc tgcgcggcgg agtggtgttc gtggacgagg tgcccaaggg cctgaccggc
     2281 aagctggacg cccgcaagat ccgcgagatc ctgatcaagg ctaagaaagg cggcaagatc
     2341 gccgtgaatt ctgcttgcaa gaactggttc agtagcttaa gccactttgt gatccacctt
     2401 aacagccacg gcttccctcc cgaggtggag gagcaggccg ccggcaccct gcccatgagc
     2461 tgcgcccagg agagcggcat ggatagacac cctgctgctt gcgccagcgc caggatcaac
     2521 gtctaatagt gagaattcac cccaccagtg caggctgcct atcagaaagt ggtggctggt
     2581 gtggctaatg ccctggccca caagtatcac taagcggcca gagctcgctg atcagcctcg
     2641 actgtgcctt ctagttgcca gccatctgtt gtttgcccct cccccgtgcc ttccttgacc
     2701 ctggaaggtg ccactcccac tgtcctttcc taataaaatg aggaaattgc atcgcattgt
     2761 ctgagtaggt gtcattctat tctggggggt ggggtggggc aggacagcaa gggggaggat
     2821 tgggaagaca atagcaggca tgctggggat gcggtgggct ctatggcttc tgaggcggaa
     2881 agaaccagct ggggctctag ggggtatccc cacgcgccct gtagcggcgc attaagcgcg
     2941 gcgggtgtgg tggttacgcg cagcgtgacc gctacacttg ccagcgccct agcgcccgct
     3001 cctttcgctt tcttcccttc ctttctcgcc acgttcgccg gctttccccg tcaagctcta
     3061 aatcgggggc tccctttagg gttccgattt agtgctttac ggcacctcga ccccaaaaaa
     3121 cttgattagg gtgatggttc acgtagtggg ccatcgccct gatagacggt ttttcgccct
     3181 ttgacgttgg agtccacgtt ctttaatagt ggactcttgt tccaaactgg aacaacactc
     3241 aaccctatct cggtctattc ttttgattta taagggattt tgccgatttc ggcctattgg
     3301 ttaaaaaatg agctgattta acaaaaattt aacgcgaatt aattctgtgg aatgtgtgtc
     3361 agttagggtg tggaaagtcc ccaggctccc cagcaggcag aagtatgcaa agcatgcatc
     3421 tcaattagtc agcaaccagg tgtggaaagt ccccaggctc cccagcaggc agaagtatgc
     3481 aaagcatgca tctcaattag tcagcaacca tagtcccgcc cctaactccg cccatcccgc
     3541 ccctaactcc gcccagttcc gcccattctc cgccccatgg ctgactaatt ttttttattt
     3601 atgcagaggc cgaggccgcc tctgcctctg agctattcca gaagtagtga ggaggctttt
     3661 ttggaggcct aggcttttgc aaaaagctcc cgggagcttg tatatccatt ttcggatctg
     3721 atcaagagac aggatgagga tcgtttcgca tgattgaaca agatggattg cacgcaggtt
     3781 ctccggccgc ttgggtggag aggctattcg gctatgactg ggcacaacag acaatcggct
     3841 gctctgatgc cgccgtgttc cggctgtcag cgcaggggcg cccggttctt tttgtcaaga
     3901 ccgacctgtc cggtgccctg aatgaactgc aggacgaggc agcgcggcta tcgtggctgg
     3961 ccacgacggg cgttccttgc gcagctgtgc tcgacgttgt cactgaagcg ggaagggact
     4021 ggctgctatt gggcgaagtg ccggggcagg atctcctgtc atctcacctt gctcctgccg
     4081 agaaagtatc catcatggct gatgcaatgc ggcggctgca tacgcttgat ccggctacct
     4141 gcccattcga ccaccaagcg aaacatcgca tcgagcgagc acgtactcgg atggaagccg
     4201 gtcttgtcga tcaggatgat ctggacgaag agcatcaggg gctcgcgcca gccgaactgt
     4261 tcgccaggct caaggcgcgc atgcccgacg gcgaggatct cgtcgtgacc catggcgatg
     4321 cctgcttgcc gaatatcatg gtggaaaatg gccgcttttc tggattcatc gactgtggcc
     4381 ggctgggtgt ggcggaccgc tatcaggaca tagcgttggc tacccgtgat attgctgaag
     4441 agcttggcgg cgaatgggct gaccgcttcc tcgtgcttta cggtatcgcc gctcccgatt
     4501 cgcagcgcat cgccttctat cgccttcttg acgagttctt ctgagcggga ctctggggtt
     4561 cgaaatgacc gaccaagcga cgcccaacct gccatcacga gatttcgatt ccaccgccgc
     4621 cttctatgaa aggttgggct tcggaatcgt tttccgggac gccggctgga tgatcctcca
     4681 gcgcggggat ctcatgctgg agttcttcgc ccaccccaac ttgtttattg cagcttataa
     4741 tggttacaaa taaagcaata gcatcacaaa tttcacaaat aaagcatttt tttcactgca
     4801 ttctagttgt ggtttgtcca aactcatcaa tgtatcttat catgtctgta taccgtcgac
     4861 ctctagctag agcttggcgt aatcatggtc atagctgttt cctgtgtgaa attgttatcc
     4921 gctcacaatt ccacacaaca tacgagccgg aagcataaag tgtaaagcct ggggtgccta
     4981 atgagtgagc taactcacat taattgcgtt gcgctcactg cccgctttcc agtcgggaaa
     5041 cctgtcgtgc cagctgcatt aatgaatcgg ccaacgcgcg gggagaggcg gtttgcgtat
     5101 tgggcgctct tccgcttcct cgctcactga ctcgctgcgc tcggtcgttc ggctgcggcg
     5161 agcggtatca gctcactcaa aggcggtaat acggttatcc acagaatcag gggataacgc
     5221 aggaaagaac atgtgagcaa aaggccagca aaaggccagg aaccgtaaaa aggccgcgtt
     5281 gctggcgttt ttccataggc tccgcccccc tgacgagcat cacaaaaatc gacgctcaag
     5341 tcagaggtgg cgaaacccga caggactata aagataccag gcgtttcccc ctggaagctc
     5401 cctcgtgcgc tctcctgttc cgaccctgcc gcttaccgga tacctgtccg cctttctccc
     5461 ttcgggaagc gtggcgcttt ctcatagctc acgctgtagg tatctcagtt cggtgtaggt
     5521 cgttcgctcc aagctgggct gtgtgcacga accccccgtt cagcccgacc gctgcgcctt
     5581 atccggtaac tatcgtcttg agtccaaccc ggtaagacac gacttatcgc cactggcagc
     5641 agccactggt aacaggatta gcagagcgag gtatgtaggc ggtgctacag agttcttgaa
     5701 gtggtggcct aactacggct acactagaag aacagtattt ggtatctgcg ctctgctgaa
     5761 gccagttacc ttcggaaaaa gagttggtag ctcttgatcc ggcaaacaaa ccaccgctgg
     5821 tagcggtggt ttttttgttt gcaagcagca gattacgcgc agaaaaaaag gatctcaaga
     5881 agatcctttg atcttttcta cggggtctga cgctcagtgg aacgaaaact cacgttaagg
     5941 gattttggtc atgagattat caaaaaggat cttcacctag atccttttaa attaaaaatg
     6001 aagttttaaa tcaatctaaa gtatatatga gtaaacttgg tctgacagtt accaatgctt
     6061 aatcagtgag gcacctatct cagcgatctg tctatttcgt tcatccatag ttgcctgact
     6121 ccccgtcgtg tagataacta cgatacggga gggcttacca tctggcccca gtgctgcaat
     6181 gataccgcga gacccacgct caccggctcc agatttatca gcaataaacc agccagccgg
     6241 aagggccgag cgcagaagtg gtcctgcaac tttatccgcc tccatccagt ctattaattg
     6301 ttgccgggaa gctagagtaa gtagttcgcc agttaatagt ttgcgcaacg ttgttgccat
     6361 tgctacaggc atcgtggtgt cacgctcgtc gtttggtatg gcttcattca gctccggttc
     6421 ccaacgatca aggcgagtta catgatcccc catgttgtgc aaaaaagcgg ttagctcctt
     6481 cggtcctccg atcgttgtca gaagtaagtt ggccgcagtg ttatcactca tggttatggc
     6541 agcactgcat aattctctta ctgtcatgcc atccgtaaga tgcttttctg tgactggtga
     6601 gtactcaacc aagtcattct gagaatagtg tatgcggcga ccgagttgct cttgcccggc
     6661 gtcaatacgg gataataccg cgccacatag cagaacttta aaagtgctca tcattggaaa
     6721 acgttcttcg gggcgaaaac tctcaaggat cttaccgctg ttgagatcca gttcgatgta
     6781 acccactcgt gcacccaact gatcttcagc atcttttact ttcaccagcg tttctgggtg
     6841 agcaaaaaca ggaaggcaaa atgccgcaaa aaagggaata agggcgacac ggaaatgttg
     6901 aatactcata ctcttccttt ttcaatatta ttgaagcatt tatcagggtt attgtctcat
     6961 gagcggatac atatttgaat gtatttagaa aaataaacaa ataggggttc cgcgcacatt
     7021 tccccgaaaa gtgccacctg acgtc
//
