LOCUS       sequence-383682-        6371 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses the wild-type I-SceI homing endonuclease from the T7 
            promoter.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6371)
  AUTHORS   Niu Y, Tenney K, Li H, Gimble FS
  TITLE     Engineering variants of the I-SceI homing endonuclease with 
            strand-specific and site-specific DNA-nicking activity.
  JOURNAL   J Mol Biol. 2008 Sep 26;382(1):188-202. doi: 
            10.1016/j.jmb.2008.07.010. Epub 2008 Jul 11.
  PUBMED    18644379
REFERENCE   2  (bases 1 to 6371)
  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: 
            "10.1016/j.jmb.2008.07.010"; journalName: "J Mol Biol"; date: 
            "2008-09-26- 26"; volume: "382"; issue: "1"; pages: "188-202"
COMMENT     Sequence Label: pET15b I-SceI (Ctermhis -MH)
FEATURES             Location/Qualifiers
     source          1..6371
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     948..967
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(987..1064)
                     /gene="lacI"
                     /label=lacI promoter
     primer_bind     1270..1289
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     primer_bind     1373..1392
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        1373..1391
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    1392..1416
                     /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)."
     RBS             1431..1453
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             2174..2191
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(2245..2263)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      2259..2306
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     promoter        complement(join(2481..2486,2487..2503,2504..2509))
                     /label=tet promoter
                     /note="E. coli promoter for tetracycline efflux protein 
                     gene"
     primer_bind     complement(2536..2554)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        2622..2726
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(2727..2795,2796..3587)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRVDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPAAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     complement(2945..2964)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      3758..4346
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4247..4266
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     4500..4517
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    4532..4671
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(4597..4616)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     4757..4779
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(4773..4964)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
ORIGIN
        1 gtggtttttc ttttcaccag tgagacgggc aacagctgat tgcccttcac cgcctggccc
       61 tgagagagtt gcagcaagcg gtccacgctg gtttgcccca gcaggcgaaa atcctgtttg
      121 atggtggtta acggcgggat ataacatgag ctgtcttcgg tatcgtcgta tcccactacc
      181 gagatatccg caccaacgcg cagcccggac tcggtaatgg cgcgcattgc gcccagcgcc
      241 atctgatcgt tggcaaccag catcgcagtg ggaacgatgc cctcattcag catttgcatg
      301 gtttgttgaa aaccggacat ggcactccag tcgccttccc gttccgctat cggctgaatt
      361 tgattgcgag tgagatattt atgccagcca gccagacgca gacgcgccga gacagaactt
      421 aatgggcccg ctaacagcgc gatttgctgg tgacccaatg cgaccagatg ctccacgccc
      481 agtcgcgtac cgtcttcatg ggagaaaata atactgttga tgggtgtctg gtcagagaca
      541 tcaagaaata acgccggaac attagtgcag gcagcttcca cagcaatggc atcctggtca
      601 tccagcggat agttaatgat cagcccactg acgcgttgcg cgagaagatt gtgcaccgcc
      661 gctttacagg cttcgacgcc gcttcgttct accatcgaca ccaccacgct ggcacccagt
      721 tgatcggcgc gagatttaat cgccgcgaca atttgcgacg gcgcgtgcag ggccagactg
      781 gaggtggcaa cgccaatcag caacgactgt ttgcccgcca gttgttgtgc cacgcggttg
      841 ggaatgtaat tcagctccgc catcgccgct tccacttttt cccgcgtttt cgcagaaacg
      901 tggctggcct ggttcaccac gcgggaaacg gtctgataag agacaccggc atactctgcg
      961 acatcgtata acgttactgg tttcacattc accaccctga attgactctc ttccgggcgc
     1021 tatcatgcca taccgcgaaa ggttttgcgc cattcgatgg tgtccgggat ctcgacgctc
     1081 tcccttatgc gactcctgca ttaggaagca gcccagtagt aggttgaggc cgttgagcac
     1141 cgccgccgca aggaatggtg catgcaagga gatggcgccc aacagtcccc cggccacggg
     1201 gcctgccacc atacccacgc cgaaacaagc gctcatgagc ccgaagtggc gagcccgatc
     1261 ttccccatcg gtgatgtcgg cgatataggc gccagcaacc gcacctgtgg cgccggtgat
     1321 gccggccacg atgcgtccgg cgtagaggat cgagatctcg atcccgcgaa attaatacga
     1381 ctcactatag gggaattgtg agcggataac aattcccctc tagaaataat tttgtttaac
     1441 tttaagaagg agatatacca tccatatgaa aaacatcaaa aaaaaccagg taatgaacct
     1501 gggtccgaac tctaaactgc tgaaagaata caaatcccag ctgatcgaac tgaacatcga
     1561 acagttcgaa gcaggtatcg gtctgatcct gggtgatgct tacatccgtt ctcgtgatga
     1621 aggtaaaacc tactgtatgc agttcgagtg gaaaaacaaa gcatacatgg accacgtatg
     1681 tctgctgtac gatcagtggg tactgtcccc gccgcacaaa aaagaacgtg ttaaccacct
     1741 gggtaacctg gtaatcacct ggggcgccca gactttcaaa caccaagctt tcaacaaact
     1801 ggctaacctg ttcatcgtta acaacaaaaa aaccatcccg aacaacctgg ttgaaaacta
     1861 cctgaccccg atgtctctgg catactggtt catggatgat ggtggtaaat gggattacaa
     1921 caaaaactct accaacaaat cgatcgtact gaacacccag tctttcactt tcgaagaagt
     1981 agaatacctg gttaagggtc tgcgtaacaa attccaactg aactgttacg taaaaatcaa
     2041 caaaaacaaa ccgatcatct acatcgattc tatgtcttac ctgatcttct acaacctgat
     2101 caaaccgtac ctgatcccgc agatgatgta caaactgccg aacactatct cctccgaaac
     2161 tttcctgaaa tctcaccacc accaccacca ctaaggatcc ggctgctaac aaagcccgaa
     2221 aggaagctga gttggctgct gccaccgctg agcaataact agcataaccc cttggggcct
     2281 ctaaacgggt cttgaggggt tttttgctga aaggaggaac tatatccgga tatcccgcaa
     2341 gaggcccggc agtaccggca taaccaagcc tatgcctaca gcatccaggg tgacggtgcc
     2401 gaggatgacg atgagcgcat tgttagattt catacacggt gcctgactgc gttagcaatt
     2461 taactgtgat aaactaccgc attaaagctt atcgatgata agctgtcaaa catgagaatt
     2521 cttgaagacg aaagggcctc gtgatacgcc tatttttata ggttaatgtc atgataataa
     2581 tggtttctta gacgtcaggt ggcacttttc ggggaaatgt gcgcggaacc cctatttgtt
     2641 tatttttcta aatacattca aatatgtatc cgctcatgag acaataaccc tgataaatgc
     2701 ttcaataata ttgaaaaagg aagagtatga gtattcaaca tttccgtgtc gcccttattc
     2761 ccttttttgc ggcattttgc cttcctgttt ttgctcaccc agaaacgctg gtgaaagtaa
     2821 aagatgctga agatcagttg ggtgcacgag tgggttacat cgaactggat ctcaacagcg
     2881 gtaagatcct tgagagtttt cgccccgaag aacgttttcc aatgatgagc acttttaaag
     2941 ttctgctatg tggcgcggta ttatcccgtg ttgacgccgg gcaagagcaa ctcggtcgcc
     3001 gcatacacta ttctcagaat gacttggttg agtactcacc agtcacagaa aagcatctta
     3061 cggatggcat gacagtaaga gaattatgca gtgctgccat aaccatgagt gataacactg
     3121 cggccaactt acttctgaca acgatcggag gaccgaagga gctaaccgct tttttgcaca
     3181 acatggggga tcatgtaact cgccttgatc gttgggaacc ggagctgaat gaagccatac
     3241 caaacgacga gcgtgacacc acgatgcctg cagcaatggc aacaacgttg cgcaaactat
     3301 taactggcga actacttact ctagcttccc ggcaacaatt aatagactgg atggaggcgg
     3361 ataaagttgc aggaccactt ctgcgctcgg cccttccggc tggctggttt attgctgata
     3421 aatctggagc cggtgagcgt gggtctcgcg gtatcattgc agcactgggg ccagatggta
     3481 agccctcccg tatcgtagtt atctacacga cggggagtca ggcaactatg gatgaacgaa
     3541 atagacagat cgctgagata ggtgcctcac tgattaagca ttggtaactg tcagaccaag
     3601 tttactcata tatactttag attgatttaa aacttcattt ttaatttaaa aggatctagg
     3661 tgaagatcct ttttgataat ctcatgacca aaatccctta acgtgagttt tcgttccact
     3721 gagcgtcaga ccccgtagaa aagatcaaag gatcttcttg agatcctttt tttctgcgcg
     3781 taatctgctg cttgcaaaca aaaaaaccac cgctaccagc ggtggtttgt ttgccggatc
     3841 aagagctacc aactcttttt ccgaaggtaa ctggcttcag cagagcgcag ataccaaata
     3901 ctgtccttct agtgtagccg tagttaggcc accacttcaa gaactctgta gcaccgccta
     3961 catacctcgc tctgctaatc ctgttaccag tggctgctgc cagtggcgat aagtcgtgtc
     4021 ttaccgggtt ggactcaaga cgatagttac cggataaggc gcagcggtcg ggctgaacgg
     4081 ggggttcgtg cacacagccc agcttggagc gaacgaccta caccgaactg agatacctac
     4141 agcgtgagct atgagaaagc gccacgcttc ccgaagggag aaaggcggac aggtatccgg
     4201 taagcggcag ggtcggaaca ggagagcgca cgagggagct tccaggggga aacgcctggt
     4261 atctttatag tcctgtcggg tttcgccacc tctgacttga gcgtcgattt ttgtgatgct
     4321 cgtcaggggg gcggagccta tggaaaaacg ccagcaacgc ggccttttta cggttcctgg
     4381 ccttttgctg gccttttgct cacatgttct ttcctgcgtt atcccctgat tctgtggata
     4441 accgtattac cgcctttgag tgagctgata ccgctcgccg cagccgaacg accgagcgca
     4501 gcgagtcagt gagcgaggaa gcggaagagc gcctgatgcg gtattttctc cttacgcatc
     4561 tgtgcggtat ttcacaccgc aatggtgcac tctcagtaca atctgctctg atgccgcata
     4621 gttaagccag tatacactcc gctatcgcta cgtgactggg tcatggctgc gccccgacac
     4681 ccgccaacac ccgctgacgc gccctgacgg gcttgtctgc tcccggcatc cgcttacaga
     4741 caagctgtga ccgtctccgg gagctgcatg tgtcagaggt tttcaccgtc atcaccgaaa
     4801 cgcgcgaggc agctgcggta aagctcatca gcgtggtcgt gaagcgattc acagatgtct
     4861 gcctgttcat ccgcgtccag ctcgttgagt ttctccagaa gcgttaatgt ctggcttctg
     4921 ataaagcggg ccatgttaag ggcggttttt tcctgtttgg tcactgatgc ctccgtgtaa
     4981 gggggatttc tgttcatggg ggtaatgata ccgatgaaac gagagaggat gctcacgata
     5041 cgggttactg atgatgaaca tgcccggtta ctggaacgtt gtgagggtaa acaactggcg
     5101 gtatggatgc ggcgggacca gagaaaaatc actcagggtc aatgccagcg cttcgttaat
     5161 acagatgtag gtgttccaca gggtagccag cagcatcctg cgatgcagat ccggaacata
     5221 atggtgcagg gcgctgactt ccgcgtttcc agactttacg aaacacggaa accgaagacc
     5281 attcatgttg ttgctcaggt cgcagacgtt ttgcagcagc agtcgcttca cgttcgctcg
     5341 cgtatcggtg attcattctg ctaaccagta aggcaacccc gccagcctag ccgggtcctc
     5401 aacgacagga gcacgatcat gcgcacccgt ggccaggacc caacgctgcc cgagatgcgc
     5461 cgcgtgcggc tgctggagat ggcggacgcg atggatatgt tctgccaagg gttggtttgc
     5521 gcattcacag ttctccgcaa gaattgattg gctccaattc ttggagtggt gaatccgtta
     5581 gcgaggtgcc gccggcttcc attcaggtcg aggtggcccg gctccatgca ccgcgacgca
     5641 acgcggggag gcagacaagg tatagggcgg cgcctacaat ccatgccaac ccgttccatg
     5701 tgctcgccga ggcggcataa atcgccgtga cgatcagcgg tccaatgatc gaagttaggc
     5761 tggtaagagc cgcgagcgat ccttgaagct gtccctgatg gtcgtcatct acctgcctgg
     5821 acagcatggc ctgcaacgcg ggcatcccga tgccgccgga agcgagaaga atcataatgg
     5881 ggaaggccat ccagcctcgc gtcgcgaacg ccagcaagac gtagcccagc gcgtcggccg
     5941 ccatgccggc gataatggcc tgcttctcgc cgaaacgttt ggtggcggga ccagtgacga
     6001 aggcttgagc gagggcgtgc aagattccga ataccgcaag cgacaggccg atcatcgtcg
     6061 cgctccagcg aaagcggtcc tcgccgaaaa tgacccagag cgctgccggc acctgtccta
     6121 cgagttgcat gataaagaag acagtcataa gtgcggcgac gatagtcatg ccccgcgccc
     6181 accggaagga gctgactggg ttgaaggctc tcaagggcat cggtcgagat cccggtgcct
     6241 aatgagtgag ctaacttaca ttaattgcgt tgcgctcact gcccgctttc cagtcgggaa
     6301 actgtcgtgc cagctgcatt aatgaatcgg ccaacgcgcg gggagaggcg gtttgcgtat
     6361 tgggcgccag g
//
