LOCUS       sequence-259217-        6727 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Firefly-luciferase single-strand annealing assay cloning backbone.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6727)
  AUTHORS   Cradick TJ, Antico CJ, Bao G
  TITLE     High-throughput cellular screening of engineered nuclease activity 
            using the single-strand annealing assay and luciferase reporter.
  JOURNAL   Methods Mol Biol. 2014;1114:339-52. doi: 
            10.1007/978-1-62703-761-7_22.
  PUBMED    24557914
REFERENCE   2  (bases 1 to 6727)
  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: "Methods Mol
            Biol."; date: "2014"; volume: "1114:339-52. doi"; pages: " 
            10.1007/978-1-62703-761-7_22"
COMMENT     Sequence Label: 1647_pGL4.51 TLV41 SSA-Luc cloning vector_Circ...
FEATURES             Location/Qualifiers
     source          1..6727
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        77..380
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        381..584
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     535..555
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     581..605
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     regulatory      703..712
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     polyA_signal    2774..2895
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2785..2804)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     2839..2858
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     promoter        3089..3446
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        3089..3280
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(3112..3132)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      3297..3432
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     3359..3378
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             3477..4271
                     /codon_start=1
                     /product="aminoglycoside phosphotransferase"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     polyA_signal    4296..4344
                     /note="synthetic polyadenylation signal"
     primer_bind     complement(4500..4517)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4671..5259)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4751..4770)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5459..6250,6251..6319))
                     /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    6424..6472
                     /note="synthetic polyadenylation signal"
     misc_feature    6486..6577
                     /label=pause site
                     /note="RNA polymerase II transcriptional pause signal from 
                     the human alpha-2 globin gene"
     primer_bind     6526..6545
                     /label=RVprimer3
                     /note="pGL3 vector, forward primer"
ORIGIN
        1 ggcattgatt attgactagt tattaatagt aatcaattac ggggtcatta gttcatagcc
       61 catatatgga gttccgcgtt acataactta cggtaaatgg cccgcctggc tgaccgccca
      121 acgacccccg cccattgacg tcaataatga cgtatgttcc catagtaacg ccaataggga
      181 ctttccattg acgtcaatgg gtggagtatt tacggtaaac tgcccacttg gcagtacatc
      241 aagtgtatca tatgccaagt ccgcccccta ttgacgtcaa tgacggtaaa tggcccgcct
      301 ggcattatgc ccagtacatg accttacggg actttcctac ttggcagtac atctacgtat
      361 tagtcatcgc tattaccatg gtgatgcggt tttggcagta caccaatggg cgtggatagc
      421 ggtttgactc acggggattt ccaagtctcc accccattga cgtcaatggg agtttgtttt
      481 ggcaccaaaa tcaacgggac tttccaaaat gtcgtaataa ccccgccccg ttgacgcaaa
      541 tgggcggtag gcgtgtacgg tgggaggtct atataagcag agctcgttta gtgaaccgtc
      601 agatcactag aagctttatt gcggtagttt atcacagtta aattgctaac gcagtcagtg
      661 ggcctcggcg gccaagcttg gcaatccggt actgttggta aagccaccat ggaagatgcc
      721 aaaaacatta agaagggccc agcgccattc tacccactcg aagacgggac cgccggcgag
      781 cagctgcaca aagccatgaa gcgctacgcc ctggtgcccg gcaccatcgc ctttaccgac
      841 gcacatatcg aggtggacat tacctacgcc gagtacttcg agatgagcgt tcggctggca
      901 gaagctatga agcgctatgg gctgaataca aaccatcgga tcgtggtgtg cagcgagaat
      961 agcttgcagt tcttcatgcc cgtgttgggt gccctgttca tcggtgtggc tgtggcccca
     1021 gctaacgaca tctacaacga gcgcgagctg ctgaacagca tgggcatcag ccagcccacc
     1081 gtcgtattcg tgagcaagaa agggctgcaa aagatcctca acgtgcaaaa gaagctaccg
     1141 atcatacaaa agatcatcat catggatagc aagaccgact accagggctt ccaaagcatg
     1201 tacaccttcg tgacttccca tttgccaccc ggcttcaacg agtacgactt cgtgcccgaa
     1261 taatgaggcg cgaccatctt cttcaaggac gacggcgcgc ctgggatcct gcaggtgtgc
     1321 agcgagaata gcttgcagtt cttcatgccc gtgttgggtg ccctgttcat cggtgtggct
     1381 gtggccccag ctaacgacat ctacaacgag cgcgagctgc tgaacagcat gggcatcagc
     1441 cagcccaccg tcgtattcgt gagcaagaaa gggctgcaaa agatcctcaa cgtgcaaaag
     1501 aagctaccga tcatacaaaa gatcatcatc atggatagca agaccgacta ccagggcttc
     1561 caaagcatgt acaccttcgt gacttcccat ttgccacccg gcttcaacga gtacgacttc
     1621 gtgcccgaga gcttcgaccg ggacaaaacc atcgccctga tcatgaacag tagtggcagt
     1681 accggattgc ccaagggcgt agccctaccg caccgcaccg cttgtgtccg attcagtcat
     1741 gcccgcgacc ccatcttcgg caaccagatc atccccgaca ccgctatcct cagcgtggtg
     1801 ccatttcacc acggcttcgg catgttcacc acgctgggct acttgatctg cggctttcgg
     1861 gtcgtgctca tgtaccgctt cgaggaggag ctattcttgc gcagcttgca agactataag
     1921 attcaatctg ccctgctggt gcccacacta tttagcttct tcgctaagag cactctcatc
     1981 gacaagtacg acctaagcaa cttgcacgag atcgccagcg gcggggcgcc gctcagcaag
     2041 gaggtaggtg aggccgtggc caaacgcttc cacctaccag gcatccgcca gggctacggc
     2101 ctgacagaaa caaccagcgc cattctgatc acccccgaag gggacgacaa gcctggcgca
     2161 gtaggcaagg tggtgccctt cttcgaggct aaggtggtgg acttggacac cggtaagaca
     2221 ctgggtgtga accagcgcgg cgagctgtgc gtccgtggcc ccatgatcat gagcggctac
     2281 gttaacaacc ccgaggctac aaacgctctc atcgacaagg acggctggct gcacagcggc
     2341 gacatcgcct actgggacga ggacgagcac ttcttcatcg tggaccggct gaagagcctg
     2401 atcaaataca agggctacca ggtagcccca gccgaactgg agagcatcct gctgcaacac
     2461 cccaacatct tcgacgccgg ggtcgccggc ctgcccgacg acgatgccgg cgagctgccc
     2521 gccgcagtcg tcgtgctgga acacggtaaa accatgaccg agaaggagat cgtggactat
     2581 gtggccagcc aggttacaac cgccaagaag ctgcgcggtg gtgttgtgtt cgtggacgag
     2641 gtgcctaaag gactgaccgg caagttggac gcccgcaaga tccgcgagat tctcattaag
     2701 gccaagaagg gcggcaagat cgccgtgtaa taattctaga gtcggggcgg ccggccgctt
     2761 cgagcagaca tgataagata cattgatgag tttggacaaa ccacaactag aatgcagtga
     2821 aaaaaatgct ttatttgtga aatttgtgat gctattgctt tatttgtaac cattataagc
     2881 tgcaataaac aagttaacaa caacaattgc attcatttta tgtttcaggt tcagggggag
     2941 gtgtgggagg ttttttaaag caagtaaaac ctctacaaat gtggtaaaat cgataaggat
     3001 ccgtttgcgt attgggcgct cttccgctga tctgcgcagc accatggcct gaaataacct
     3061 ctgaaagagg aacttggtta gctaccttct gaggcggaaa gaaccagctg tggaatgtgt
     3121 gtcagttagg gtgtggaaag tccccaggct ccccagcagg cagaagtatg caaagcatgc
     3181 atctcaatta gtcagcaacc aggtgtggaa agtccccagg ctccccagca ggcagaagta
     3241 tgcaaagcat gcatctcaat tagtcagcaa ccatagtccc gcccctaact ccgcccatcc
     3301 cgcccctaac tccgcccagt tccgcccatt ctccgcccca tggctgacta atttttttta
     3361 tttatgcaga ggccgaggcc gcctctgcct ctgagctatt ccagaagtag tgaggaggct
     3421 tttttggagg cctaggcttt tgcaaaaagc tcgattcttc tgacactagc gccaccatga
     3481 tcgaacaaga cggcctccat gctggcagtc ccgcagcttg ggtcgaacgc ttgttcgggt
     3541 acgactgggc ccagcagacc atcggatgta gcgatgcggc cgtgttccgt ctaagcgctc
     3601 aaggccggcc cgtgctgttc gtgaagaccg acctgagcgg cgccctgaac gagcttcaag
     3661 acgaggctgc ccgcctgagc tggctggcca ccaccggcgt accctgcgcc gctgtgttgg
     3721 atgttgtgac cgaagccggc cgggactggc tgctgctggg cgaggtccct ggccaggatc
     3781 tgctgagcag ccaccttgcc cccgctgaga aggtttctat catggccgat gcaatgcggc
     3841 gcctgcacac cctggacccc gctacctgcc ccttcgacca ccaggctaag catcggatcg
     3901 agcgtgctcg gacccgcatg gaggccggcc tggtggacca ggacgacctg gacgaggagc
     3961 atcagggcct ggcccccgct gaactgttcg cccgactgaa agcccgcatg ccggacggtg
     4021 aggacctggt tgtcacacac ggagatgcct gcctccctaa catcatggtc gagaatggcc
     4081 gcttctccgg cttcatcgac tgcggtcgcc taggagttgc cgaccgctac caggacatcg
     4141 ccctggccac ccgcgacatc gctgaggagc ttggcggcga gtgggccgac cgcttcttag
     4201 tcttgtacgg catcgcagct cccgacagcc agcgcatcgc cttctaccgc ttgctcgacg
     4261 agttctttta atgatctaga accggtcatg gccgcaataa aatatcttta ttttcattac
     4321 atctgtgtgt tggttttttg tgtgttcgaa ctagatgctg tcgaccgatg cccttgagag
     4381 ccttcaaccc agtcagctcc ttccggtggg cgcggggcat gactatcgtc gccgcactta
     4441 tgactgtctt ctttatcatg caactcgtag gacaggtgcc ggcagcgctc ttccgcttcc
     4501 tcgctcactg actcgctgcg ctcggtcgtt cggctgcggc gagcggtatc agctcactca
     4561 aaggcggtaa tacggttatc cacagaatca ggggataacg caggaaagaa catgtgagca
     4621 aaaggccagc aaaaggccag gaaccgtaaa aaggccgcgt tgctggcgtt tttccatagg
     4681 ctccgccccc ctgacgagca tcacaaaaat cgacgctcaa gtcagaggtg gcgaaacccg
     4741 acaggactat aaagatacca ggcgtttccc cctggaagct ccctcgtgcg ctctcctgtt
     4801 ccgaccctgc cgcttaccgg atacctgtcc gcctttctcc cttcgggaag cgtggcgctt
     4861 tctcatagct cacgctgtag gtatctcagt tcggtgtagg tcgttcgctc caagctgggc
     4921 tgtgtgcacg aaccccccgt tcagcccgac cgctgcgcct tatccggtaa ctatcgtctt
     4981 gagtccaacc cggtaagaca cgacttatcg ccactggcag cagccactgg taacaggatt
     5041 agcagagcga ggtatgtagg cggtgctaca gagttcttga agtggtggcc taactacggc
     5101 tacactagaa gaacagtatt tggtatctgc gctctgctga agccagttac cttcggaaaa
     5161 agagttggta gctcttgatc cggcaaacaa accaccgctg gtagcggtgg tttttttgtt
     5221 tgcaagcagc agattacgcg cagaaaaaaa ggatctcaag aagatccttt gatcttttct
     5281 acggggtctg acgctcagtg gaacgaaaac tcacgttaag ggattttggt catgagatta
     5341 tcaaaaagga tcttcaccta gatcctttta aattaaaaat gaagttttaa atcaatctaa
     5401 agtatatatg agtaaacttg gtctgacagc ggccgcaaat gctaaaccac tgcagtggtt
     5461 accagtgctt gatcagtgag gcaccgatct cagcgatctg cctatttcgt tcgtccatag
     5521 tggcctgact ccccgtcgtg tagatcacta cgattcgtga gggcttacca tcaggcccca
     5581 gcgcagcaat gatgccgcga gagccgcgtt caccggcccc cgatttgtca gcaatgaacc
     5641 agccagcagg gagggccgag cgaagaagtg gtcctgctac tttgtccgcc tccatccagt
     5701 ctatgagctg ctgtcgtgat gctagagtaa gaagttcgcc agtgagtagt ttccgaagag
     5761 ttgtggccat tgctactggc atcgtggtat cacgctcgtc gttcggtatg gcttcgttca
     5821 actctggttc ccagcggtca agccgggtca catgatcacc catattatga agaaatgcag
     5881 tcagctcctt agggcctccg atcgttgtca gaagtaagtt ggccgcggtg ttgtcgctca
     5941 tggtaatggc agcactacac aattctctta ccgtcatgcc atccgtaaga tgcttttccg
     6001 tgaccggcga gtactcaacc aagtcgtttt gtgagtagtg tatacggcga ccaagctgct
     6061 cttgcccggc gtctatacgg gacaacaccg cgccacatag cagtactttg aaagtgctca
     6121 tcatcgggaa tcgttcttcg gggcggaaag actcaaggat cttgccgcta ttgagatcca
     6181 gttcgatata gcccactctt gcacccagtt gatcttcagc atcttttact ttcaccagcg
     6241 tttcggggtg tgcaaaaaca ggcaagcaaa atgccgcaaa gaagggaatg agtgcgacac
     6301 gaaaatgttg gatgctcata ctcgtccttt ttcaatatta ttgaagcatt tatcagggtt
     6361 actagtacgt ctctcaagga taagtaagta atattaaggt acgggaggta ttggacaggc
     6421 cgcaataaaa tatctttatt ttcattacat ctgtgtgttg gttttttgtg tgaatcgata
     6481 gtactaacat acgctctcca tcaaaacaaa acgaaacaaa acaaactagc aaaataggct
     6541 gtccccagtg caagtgcagg tgccagaaca tttctctggc ctaactggcc tcaatattgg
     6601 ccattagcca tattattcat tggttatata gcataaatca atattggcta ttggccattg
     6661 catacgttgt atctatatca taatatgtac atttatattg gctcatgtcc aatatgaccg
     6721 ccatgtt
//
