LOCUS       sequence-472679-        6722 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Express COLA1A in luciferase reporter assay.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6722)
  AUTHORS   Promega
  TITLE     Promega Plasmid Collection
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 6722)
  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            "Unpublished"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: pGL4 [luc2P/COLA1A/Hygro] Vector
FEATURES             Location/Qualifiers
     source          1..6722
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     regulatory      847..856
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             853..2502
                     /codon_start=1
                     /product="firefly luciferase"
                     /label=luciferase
                     /note="synthetic luc2 version of the luciferase gene"
                     /translation="MEDAKNIKKGPAPFYPLEDGTAGEQLHKAMKRYALVPGTIAFTDA
                     HIEVDITYAEYFEMSVRLAEAMKRYGLNTNHRIVVCSENSLQFFMPVLGALFIGVAVAP
                     ANDIYNERELLNSMGISQPTVVFVSKKGLQKILNVQKKLPIIQKIIIMDSKTDYQGFQS
                     MYTFVTSHLPPGFNEYDFVPESFDRDKTIALIMNSSGSTGLPKGVALPHRTACVRFSHA
                     RDPIFGNQIIPDTAILSVVPFHHGFGMFTTLGYLICGFRVVLMYRFEEELFLRSLQDYK
                     IQSALLVPTLFSFFAKSTLIDKYDLSNLHEIASGGAPLSKEVGEAVAKRFHLPGIRQGY
                     GLTETTSAILITPEGDDKPGAVGKVVPFFEAKVVDLDTGKTLGVNQRGELCVRGPMIMS
                     GYVNNPEATNALIDKDGWLHSGDIAYWDEDEHFFIVDRLKSLIKYKGYQVAPAELESIL
                     LQHPNIFDAGVAGLPDDDAGELPAAVVVLEHGKTMTEKEIVDYVASQVTTAKKLRGGVV
                     FVDEVPKGLTGKLDARKIREILIKAKKGGKIAV"
     CDS             2506..2625
                     /codon_start=1
                     /product="PEST degradation sequence from mouse ornithine 
                     decarboxylase"
                     /label=hPEST
                     /note="human codon-optimized"
                     /translation="SHGFPPEVEEQAAGTLPMSCAQESGMDRHPAACASARINV"
     polyA_signal    2677..2798
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2688..2707)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     2742..2761
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     promoter        2992..3349
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        2992..3183
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(3015..3035)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      3200..3335
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     3262..3281
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             3380..4417
                     /codon_start=1
                     /product="hygromycin B phosphotransferase"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="MKKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRG
                     YVLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLP
                     ETELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQ
                     TVMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMF
                     GDSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDG
                     NFDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAK
                     EVGRV"
     RBS             4396..4404
                     /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    4441..4489
                     /note="synthetic polyadenylation signal"
     primer_bind     complement(4645..4662)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4816..5404)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4896..4915)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5604..6395,6396..6464))
                     /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    6569..6617
                     /note="synthetic polyadenylation signal"
     misc_feature    6631..6722
                     /label=pause site
                     /note="RNA polymerase II transcriptional pause signal from 
                     the human alpha-2 globin gene"
     primer_bind     6671..6690
                     /label=RVprimer3
                     /note="pGL3 vector, forward primer"
ORIGIN
        1 ggcctaactg gccggtacca cggccagccg gccagccgac gtggctccct ccccttctgt
       61 tccttttttt tcccctttgc cttcgttgca caaaaccagc tgggggaggg cgtggagagg
      121 ggcgggggga ggcaatggaa tcttggatgg tttgggggag gcgggactcc ccgcttccac
      181 gtttgcagct ctggagcacc cggggtgggg agctgcacag gagggagaga aatgaacagg
      241 gcactgcaag gagaccccca ggccttctct cagccctaca gagtttctca ggacgaggta
      301 gattggggtt gaggcagagc cttgttgggg gaatgggaca tggaggaaga aaggacgtgg
      361 agttctagag ccatcttcct tagatatagc ctgctgtcct tcgggtcccc agaccctttc
      421 agagtgtaca gatgattctc tctggttgct gcacccctac cacagcacct ctggcccagg
      481 ctgggctggg gggctgggga ggcagagctg cgaagagggg agatgtgggg tggactccct
      541 tccctcctcc tccccctctc cattccaact cccaaattgg gggccgggcc aggcagctct
      601 gattggctgg ggcacgggcg gccggctccc cctctccgag gggcagggtt cctccctgct
      661 ctccatcagg acagtataaa aggggcccgg gccagtcgtc ggagcagacg ggagtttctc
      721 ctcggggtcg gagcaggagg cacgcggagt gtgaggccac gcatgagcgg acgctaaccc
      781 cctccccagc cacaaagagt ctacatgtct agggtctaag cttggcaatc cggtactgtt
      841 ggtaaagcca ccatggaaga tgccaaaaac attaagaagg gcccagcgcc attctaccca
      901 ctcgaagacg ggaccgccgg cgagcagctg cacaaagcca tgaagcgcta cgccctggtg
      961 cccggcacca tcgcctttac cgacgcacat atcgaggtgg acattaccta cgccgagtac
     1021 ttcgagatga gcgttcggct ggcagaagct atgaagcgct atgggctgaa tacaaaccat
     1081 cggatcgtgg tgtgcagcga gaatagcttg cagttcttca tgcccgtgtt gggtgccctg
     1141 ttcatcggtg tggctgtggc cccagctaac gacatctaca acgagcgcga gctgctgaac
     1201 agcatgggca tcagccagcc caccgtcgta ttcgtgagca agaaagggct gcaaaagatc
     1261 ctcaacgtgc aaaagaagct accgatcata caaaagatca tcatcatgga tagcaagacc
     1321 gactaccagg gcttccaaag catgtacacc ttcgtgactt cccatttgcc acccggcttc
     1381 aacgagtacg acttcgtgcc cgagagcttc gaccgggaca aaaccatcgc cctgatcatg
     1441 aacagtagtg gcagtaccgg attgcccaag ggcgtagccc taccgcaccg caccgcttgt
     1501 gtccgattca gtcatgcccg cgaccccatc ttcggcaacc agatcatccc cgacaccgct
     1561 atcctcagcg tggtgccatt tcaccacggc ttcggcatgt tcaccacgct gggctacttg
     1621 atctgcggct ttcgggtcgt gctcatgtac cgcttcgagg aggagctatt cttgcgcagc
     1681 ttgcaagact ataagattca atctgccctg ctggtgccca cactatttag cttcttcgct
     1741 aagagcactc tcatcgacaa gtacgaccta agcaacttgc acgagatcgc cagcggcggg
     1801 gcgccgctca gcaaggaggt aggtgaggcc gtggccaaac gcttccacct accaggcatc
     1861 cgccagggct acggcctgac agaaacaacc agcgccattc tgatcacccc cgaaggggac
     1921 gacaagcctg gcgcagtagg caaggtggtg cccttcttcg aggctaaggt ggtggacttg
     1981 gacaccggta agacactggg tgtgaaccag cgcggcgagc tgtgcgtccg tggccccatg
     2041 atcatgagcg gctacgttaa caaccccgag gctacaaacg ctctcatcga caaggacggc
     2101 tggctgcaca gcggcgacat cgcctactgg gacgaggacg agcacttctt catcgtggac
     2161 cggctgaaga gcctgatcaa atacaagggc taccaggtag ccccagccga actggagagc
     2221 atcctgctgc aacaccccaa catcttcgac gccggggtcg ccggcctgcc cgacgacgat
     2281 gccggcgagc tgcccgccgc agtcgtcgtg ctggaacacg gtaaaaccat gaccgagaag
     2341 gagatcgtgg actatgtggc cagccaggtt acaaccgcca agaagctgcg cggtggtgtt
     2401 gtgttcgtgg acgaggtgcc taaaggactg accggcaagt tggacgcccg caagatccgc
     2461 gagattctca ttaaggccaa gaagggcggc aagatcgccg tgaattctca cggcttccct
     2521 cccgaggtgg aggagcaggc cgccggcacc ctgcccatga gctgcgccca ggagagcggc
     2581 atggatagac accctgctgc ttgcgccagc gccaggatca acgtctaagg ccgcgactct
     2641 agagtcgggg cggccggccg cttcgagcag acatgataag atacattgat gagtttggac
     2701 aaaccacaac tagaatgcag tgaaaaaaat gctttatttg tgaaatttgt gatgctattg
     2761 ctttatttgt aaccattata agctgcaata aacaagttaa caacaacaat tgcattcatt
     2821 ttatgtttca ggttcagggg gaggtgtggg aggtttttta aagcaagtaa aacctctaca
     2881 aatgtggtaa aatcgataag gatccgtttg cgtattgggc gctcttccgc tgatctgcgc
     2941 agcaccatgg cctgaaataa cctctgaaag aggaacttgg ttagctacct tctgaggcgg
     3001 aaagaaccag ctgtggaatg tgtgtcagtt agggtgtgga aagtccccag gctccccagc
     3061 aggcagaagt atgcaaagca tgcatctcaa ttagtcagca accaggtgtg gaaagtcccc
     3121 aggctcccca gcaggcagaa gtatgcaaag catgcatctc aattagtcag caaccatagt
     3181 cccgccccta actccgccca tcccgcccct aactccgccc agttccgccc attctccgcc
     3241 ccatggctga ctaatttttt ttatttatgc agaggccgag gccgcctctg cctctgagct
     3301 attccagaag tagtgaggag gcttttttgg aggcctaggc ttttgcaaaa agctcgattc
     3361 ttctgacact agcgccacca tgaagaagcc cgaactcacc gctaccagcg ttgaaaaatt
     3421 tctcatcgag aagttcgaca gtgtgagcga cctgatgcag ttgtcggagg gcgaagagag
     3481 ccgagccttc agcttcgatg tcggcggacg cggctatgta ctgcgggtga atagctgcgc
     3541 tgatggcttc tacaaagacc gctacgtgta ccgccacttc gccagcgctg cactacccat
     3601 ccccgaagtg ttggacatcg gcgagttcag cgagagcctg acatactgca tcagtagacg
     3661 cgcccaaggc gttactctcc aagacctccc cgaaacagag ctgcctgctg tgttacagcc
     3721 tgtcgccgaa gctatggatg ctattgccgc cgccgacctc agtcaaacca gcggcttcgg
     3781 cccattcggg ccccaaggca tcggccagta cacaacctgg cgggatttca tttgcgccat
     3841 tgctgatccc catgtctacc actggcagac cgtgatggac gacaccgtgt ccgccagcgt
     3901 agctcaagcc ctggacgaac tgatgctgtg ggccgaagac tgtcccgagg tgcgccacct
     3961 cgtccatgcc gacttcggca gcaacaacgt cctgaccgac aacggccgca tcaccgccgt
     4021 aatcgactgg tccgaagcta tgttcgggga cagtcagtac gaggtggcca acatcttctt
     4081 ctggcggccc tggctggctt gcatggagca gcagactcgc tacttcgagc gccggcatcc
     4141 cgagctggcc ggcagccctc gtctgcgagc ctacatgctg cgcatcggcc tggatcagct
     4201 ctaccagagc ctcgtggacg gcaacttcga cgatgctgcc tgggctcaag gccgctgcga
     4261 tgccatcgtc cgcagcgggg ccggcaccgt cggtcgcaca caaatcgctc gccggagcgc
     4321 agccgtatgg accgacggct gcgtcgaggt gctggccgac agcggcaacc gccggcccag
     4381 tacacgaccg cgcgctaagg aggtaggtcg agtttaaact ctagaaccgg tcatggccgc
     4441 aataaaatat ctttattttc attacatctg tgtgttggtt ttttgtgtgt tcgaactaga
     4501 tgctgtcgac cgatgccctt gagagccttc aacccagtca gctccttccg gtgggcgcgg
     4561 ggcatgacta tcgtcgccgc acttatgact gtcttcttta tcatgcaact cgtaggacag
     4621 gtgccggcag cgctcttccg cttcctcgct cactgactcg ctgcgctcgg tcgttcggct
     4681 gcggcgagcg gtatcagctc actcaaaggc ggtaatacgg ttatccacag aatcagggga
     4741 taacgcagga aagaacatgt gagcaaaagg ccagcaaaag gccaggaacc gtaaaaaggc
     4801 cgcgttgctg gcgtttttcc ataggctccg cccccctgac gagcatcaca aaaatcgacg
     4861 ctcaagtcag aggtggcgaa acccgacagg actataaaga taccaggcgt ttccccctgg
     4921 aagctccctc gtgcgctctc ctgttccgac cctgccgctt accggatacc tgtccgcctt
     4981 tctcccttcg ggaagcgtgg cgctttctca tagctcacgc tgtaggtatc tcagttcggt
     5041 gtaggtcgtt cgctccaagc tgggctgtgt gcacgaaccc cccgttcagc ccgaccgctg
     5101 cgccttatcc ggtaactatc gtcttgagtc caacccggta agacacgact tatcgccact
     5161 ggcagcagcc actggtaaca ggattagcag agcgaggtat gtaggcggtg ctacagagtt
     5221 cttgaagtgg tggcctaact acggctacac tagaagaaca gtatttggta tctgcgctct
     5281 gctgaagcca gttaccttcg gaaaaagagt tggtagctct tgatccggca aacaaaccac
     5341 cgctggtagc ggtggttttt ttgtttgcaa gcagcagatt acgcgcagaa aaaaaggatc
     5401 tcaagaagat cctttgatct tttctacggg gtctgacgct cagtggaacg aaaactcacg
     5461 ttaagggatt ttggtcatga gattatcaaa aaggatcttc acctagatcc ttttaaatta
     5521 aaaatgaagt tttaaatcaa tctaaagtat atatgagtaa acttggtctg acagcggccg
     5581 caaatgctaa accactgcag tggttaccag tgcttgatca gtgaggcacc gatctcagcg
     5641 atctgcctat ttcgttcgtc catagtggcc tgactccccg tcgtgtagat cactacgatt
     5701 cgtgagggct taccatcagg ccccagcgca gcaatgatgc cgcgagagcc gcgttcaccg
     5761 gcccccgatt tgtcagcaat gaaccagcca gcagggaggg ccgagcgaag aagtggtcct
     5821 gctactttgt ccgcctccat ccagtctatg agctgctgtc gtgatgctag agtaagaagt
     5881 tcgccagtga gtagtttccg aagagttgtg gccattgcta ctggcatcgt ggtatcacgc
     5941 tcgtcgttcg gtatggcttc gttcaactct ggttcccagc ggtcaagccg ggtcacatga
     6001 tcacccatat tatgaagaaa tgcagtcagc tccttagggc ctccgatcgt tgtcagaagt
     6061 aagttggccg cggtgttgtc gctcatggta atggcagcac tacacaattc tcttaccgtc
     6121 atgccatccg taagatgctt ttccgtgacc ggcgagtact caaccaagtc gttttgtgag
     6181 tagtgtatac ggcgaccaag ctgctcttgc ccggcgtcta tacgggacaa caccgcgcca
     6241 catagcagta ctttgaaagt gctcatcatc gggaatcgtt cttcggggcg gaaagactca
     6301 aggatcttgc cgctattgag atccagttcg atatagccca ctcttgcacc cagttgatct
     6361 tcagcatctt ttactttcac cagcgtttcg gggtgtgcaa aaacaggcaa gcaaaatgcc
     6421 gcaaagaagg gaatgagtgc gacacgaaaa tgttggatgc tcatactcgt cctttttcaa
     6481 tattattgaa gcatttatca gggttactag tacgtctctc aaggataagt aagtaatatt
     6541 aaggtacggg aggtattgga caggccgcaa taaaatatct ttattttcat tacatctgtg
     6601 tgttggtttt ttgtgtgaat cgatagtact aacatacgct ctccatcaaa acaaaacgaa
     6661 acaaaacaaa ctagcaaaat aggctgtccc cagtgcaagt gcaggtgcca gaacatttct
     6721 ct
//
