The pSP-luc+ Vector is a luciferase cassette vector containing the engineered firefly luciferase gene, luc+, which has been optimized for genetic reporter applications. The pSP-luc+ Vector is not itself intended for the expression of luciferase in eukaryotic cells because it does not contain eukaryotic promoters, enhancers or polyadenylation signals. However, the pGL3 Luciferase Reporter Vectors contain luc+ and are configured for expression analysis in mammalian cells (1).

The luc+ gene is positioned downstream of an SP6 promoter and ribosome binding site which, in the presence of SP6 polymerase, drive in vivo and in vitro luciferase expression. An opposing T7 promoter* is located immediately downstream of luc+. Thus, the pSP-luc+ Vector provides a convenient template for the in vitro synthesis of both sense and anti-sense luciferase transcripts for studies involving in situ hybridization, RNA processing, RNA transfection or coupled in vitro transcription/translation and protein folding. Multiple cloning regions containing recognition sequences for commonly used restriction enzymes are positioned at the 5 and 3 ends of luc+ to provide maximum flexibility in cloning. Subcloning luc+ into expression vectors provides a useful genetic reporter with exceptional sensitivity. Luciferase enzymatic activity can be assayed most efficiently using Promega's Luciferase Assay System.

*U.S. Patent No. 4,766,072 has been issued to Promega Corporation for transcription vectors having two different promoter sequences separated by a series of unique restriction sites into which foreign DNA can be inserted.

Luciferase is a 62kDa monomeric protein that does not require post-translational modifications for enzymatic activity. Thus, it can function as a genetic reporter immediately upon translation (2-4). Luciferase synthesized by in vitro translation can be labeled with 35S, as the protein contains 4 cysteine and 14 methionine residues. To ensure full enzymatic activity of luciferase, no more than 5 codons can be deleted from either the 5 or 3 end of the coding region.

 back Return to this vector's summary.


LOCUS       CVU47122     4100 bp    DNA   circular  SYN       20-FEB-1996
DEFINITION  Cloning vector pSP-luc+, Luciferase cassette vector, complete
            sequence.
ACCESSION   U47122
NID         g1197679
KEYWORDS    .
SOURCE      Cloning vector pSP-luc+.
  ORGANISM  Cloning vector pSP-luc+
            artificial sequence; cloning vectors.
REFERENCE   1  (bases 1 to 4100)
  AUTHORS   Brondyk,W.H. and Groskreutz,D.J.
  TITLE     Direct Submission
  JOURNAL   Submitted (25-JAN-1996) Melinda Brady, R&D, Promega Corporation,
            5445 East Cheryl Parkway, Madison, WI 53711, USA
COMMENT
            NCBI gi: 1197679
FEATURES               Location/Qualifiers
     source          1..4100
                     /organism="Cloning vector pSP-luc+"
     misc_feature    1
                     /note="SP6 RNA polymerase initiation site"
     misc_feature    11..48
                     /note="upstream multiple cloning site"
     CDS             45..1697
                     /gene="luc+"
                     /note="firefly luciferase;  NCBI gi: 1197680"
                     /codon_start=1
                     /product="luciferase"
                     /db_xref="PID:g1197680"
                     /translation="MEDAKNIKKGPAPFYPLEDGTAGEQLHKAMKRYALVPGTIAFTD
                     AHIEVDITYAEYFEMSVRLAEAMKRYGLNTNHRIVVCSENSLQFFMPVLGALFIGVAV
                     APANDIYNERELLNSMGISQPTVVFVSKKGLQKILNVQKKLPIIQKIIIMDSKTDYQG
                     FQSMYTFVTSHLPPGFNEYDFVPESFDRDKTIALIMNSSGSTGLPKGVALPHRTACVR
                     FSHARDPIFGNQIIPDTAILSVVPFHHGFGMFTTLGYLICGFRVVLMYRFEEELFLRS
                     LQDYKIQSALLVPTLFSFFAKSTLIDKYDLSNLHEIASGGAPLSKEVGEAVAKRFHLP
                     GIRQGYGLTETTSAILITPEGDDKPGAVGKVVPFFEAKVVDLDTGKTLGVNQRGELCV
                     RGPMIMSGYVNNPEATNALIDKDGWLHSGDIAYWDEDEHFFIVDRLKSLIKYKGYQVA
                     PAELESILLQHPNIFDAGVAGLPDDDAGELPAAVVVLEHGKTMTEKEIVDYVASQVTT
                     AKKLRGGVVFVDEVPKGLTGKLDARKIREILIKAKKGGKIAV"
     primer_bind     complement(46..68)
                     /gene="luc+"
                     /note="GLprimer2 sequencing primer binding site"
     misc_feature    1699..1728
                     /note="downstream multiple cloning site"
     promoter        complement(1733..1756)
                     /note="T7 RNA polymerase promoter"
     misc_feature    1739
                     /note="T7 RNA polymerase initiation site"
     rep_origin      2011
                     /note="ColE1-derived plasmid replication origin"
     CDS             complement(2773..3633)
                     /gene="ampR"
                     /note="NCBI gi: 1197681"
                     /codon_start=1
                     /function="ampicillin resistance"
                     /product="beta-lactamase"
                     /db_xref="PID:g1197681"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGY
                     IELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVE
                     YSPVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRL
                     DRWEPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPL
                     LRSALPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIA
                     EIGASLIKHW"
     promoter        join(4084..4100,1..6)
                     /note="SP6 RNA polymerase promoter"
BASE COUNT     1076 a    972 c   1008 g   1044 t
ORIGIN      
        1 gaaccagatc ggtaccgcta gcaagatctc ctaggaagct ttccatggaa gacgccaaaa
       61 acataaagaa aggcccggcg ccattctatc cgctggaaga tggaaccgct ggagagcaac
      121 tgcataaggc tatgaagaga tacgccctgg ttcctggaac aattgctttt acagatgcac
      181 atatcgaggt ggacatcact tacgctgagt acttcgaaat gtccgttcgg ttggcagaag
      241 ctatgaaacg atatgggctg aatacaaatc acagaatcgt cgtatgcagt gaaaactctc
      301 ttcaattctt tatgccggtg ttgggcgcgt tatttatcgg agttgcagtt gcgcccgcga
      361 acgacattta taatgaacgt gaattgctca acagtatggg catttcgcag cctaccgtgg
      421 tgttcgtttc caaaaagggg ttgcaaaaaa ttttgaacgt gcaaaaaaag ctcccaatca
      481 tccaaaaaat tattatcatg gattctaaaa cggattacca gggatttcag tcgatgtaca
      541 cgttcgtcac atctcatcta cctcccggtt ttaatgaata cgattttgtg ccagagtcct
      601 tcgataggga caagacaatt gcactgatca tgaactcctc tggatctact ggtctgccta
      661 aaggtgtcgc tctgcctcat agaactgcct gcgtgagatt ctcgcatgcc agagatccta
      721 tttttggcaa tcaaatcatt ccggatactg cgattttaag tgttgttcca ttccatcacg
      781 gttttggaat gtttactaca ctcggatatt tgatatgtgg atttcgagtc gtcttaatgt
      841 atagatttga agaagagctg tttctgagga gccttcagga ttacaagatt caaagtgcgc
      901 tgctggtgcc aaccctattc tccttcttcg ccaaaagcac tctgattgac aaatacgatt
      961 tatctaattt acacgaaatt gcttctggtg gcgctcccct ctctaaggaa gtcggggaag
     1021 cggttgccaa gaggttccat ctgccaggta tcaggcaagg atatgggctc actgagacta
     1081 catcagctat tctgattaca cccgaggggg atgataaacc gggcgcggtc ggtaaagttg
     1141 ttccattttt tgaagcgaag gttgtggatc tggataccgg gaaaacgctg ggcgttaatc
     1201 aaagaggcga actgtgtgtg agaggtccta tgattatgtc cggttatgta aacaatccgg
     1261 aagcgaccaa cgccttgatt gacaaggatg gatggctaca ttctggagac atagcttact
     1321 gggacgaaga cgaacacttc ttcatcgttg accgcctgaa gtctctgatt aagtacaaag
     1381 gctatcaggt ggctcccgct gaattggaat ccatcttgct ccaacacccc aacatcttcg
     1441 acgcaggtgt cgcaggtctt cccgacgatg acgccggtga acttcccgcc gccgttgttg
     1501 ttttggagca cggaaagacg atgacggaaa aagagatcgt ggattacgtc gccagtcaag
     1561 taacaaccgc gaaaaagttg cgcggaggag ttgtgtttgt ggacgaagta ccgaaaggtc
     1621 ttaccggaaa actcgacgca agaaaaatca gagagatcct cataaaggcc aagaagggcg
     1681 gaaagatcgc cgtgtaattc tagagaattc gatatcatcg atctcgaggc cggtctccct
     1741 atagtgagtc gtattaattt cgataagcca ggttaacctg cattaatgaa tcggccaacg
     1801 cgcggggaga ggcggtttgc gtattgggcg ctcttccgct tcctcgctca ctgactcgct
     1861 gcgctcggtc gttcggctgc ggcgagcggt atcagctcac tcaaaggcgg taatacggtt
     1921 atccacagaa tcaggggata acgcaggaaa gaacatgtga gcaaaaggcc agcaaaaggc
     1981 caggaaccgt aaaaaggccg cgttgctggc gtttttccat aggctccgcc cccctgacga
     2041 gcatcacaaa aatcgacgct caagtcagag gtggcgaaac ccgacaggac tataaagata
     2101 ccaggcgttt ccccctggaa gctccctcgt gcgctctcct gttccgaccc tgccgcttac
     2161 cggatacctg tccgcctttc tcccttcggg aagcgtggcg ctttctcaat gctcacgctg
     2221 taggtatctc agttcggtgt aggtcgttcg ctccaagctg ggctgtgtgc acgaaccccc
     2281 cgttcagccc gaccgctgcg ccttatccgg taactatcgt cttgagtcca acccggtaag
     2341 acacgactta tcgccactgg cagcagccac tggtaacagg attagcagag cgaggtatgt
     2401 aggcggtgct acagagttct tgaagtggtg gcctaactac ggctacacta gaaggacagt
     2461 atttggtatc tgcgctctgc tgaagccagt taccttcgga aaaagagttg gtagctcttg
     2521 atccggcaaa caaaccaccg ctggtagcgg tggttttttt gtttgcaagc agcagattac
     2581 gcgcagaaaa aaaggatctc aagaagatcc tttgatcttt tctacggggt ctgacgctca
     2641 gtggaacgaa aactcacgtt aagggatttt ggtcatgaga ttatcaaaaa ggatcttcac
     2701 ctagatcctt ttaaattaaa aatgaagttt taaatcaatc taaagtatat atgagtaaac
     2761 ttggtctgac agttaccaat gcttaatcag tgaggcacct atctcagcga tctgtctatt
     2821 tcgttcatcc atagttgcct gactccccgt cgtgtagata actacgatac gggagggctt
     2881 accatctggc cccagtgctg caatgatacc gcgagaccca cgctcaccgg ctccagattt
     2941 atcagcaata aaccagccag ccggaagggc cgagcgcaga agtggtcctg caactttatc
     3001 cgcctccatc cagtctatta attgttgccg ggaagctaga gtaagtagtt cgccagttaa
     3061 tagtttgcgc aacgttgttg ccattgctac aggcatcgtg gtgtcacgct cgtcgtttgg
     3121 tatggcttca ttcagctccg gttcccaacg atcaaggcga gttacatgat cccccatgtt
     3181 gtgcaaaaaa gcggttagct ccttcggtcc tccgatcgtt gtcagaagta agttggccgc
     3241 agtgttatca ctcatggtta tggcagcact gcataattct cttactgtca tgccatccgt
     3301 aagatgcttt tctgtgactg gtgagtactc aaccaagtca ttctgagaat agtgtatgcg
     3361 gcgaccgagt tgctcttgcc cggcgtcaat acgggataat accgcgccac atagcagaac
     3421 tttaaaagtg ctcatcattg gaaaacgttc ttcggggcga aaactctcaa ggatcttacc
     3481 gctgttgaga tccagttcga tgtaacccac tcgtgcaccc aactgatctt cagcatcttt
     3541 tactttcacc agcgtttctg ggtgagcaaa aacaggaagg caaaatgccg caaaaaaggg
     3601 aataagggcg acacggaaat gttgaatact catactcttc ctttttcaat attattgaag
     3661 catttatcag ggttattgtc tcatgagcgg atacatattt gaatgtattt agaaaaataa
     3721 acaaataggg gttccgcgca catttccccg aaaagtgcca cctgacgtct aagaaaccat
     3781 tattatcatg acattaacct ataaaaatag gcgtatcacg aggccctttc gtctcgcgcg
     3841 tttcggtgat gacggtgaaa acctctgaca catgcagctc ccggagacgg tcacagcttg
     3901 tctgtaagcg gatgccggga gcagacaagc ccgtcagggc gcgtcagcgg gtgttggcgg
     3961 gtgtcggggc tggcttaact atgcggcatc agagcagatt gtactgagag tgcaccatat
     4021 ggacatattg tcgttagaac gcggctacaa ttaatacata accttatgta tcatacacat
     4081 acgatttagg tgacactata 
//