LOCUS       pAL-B10                 3041 bp ds-DNA     circular     07-MAR-2025
DEFINITION  .
KEYWORDS    "accession:addgene_31027_14207"
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
COMMENT     Imported from database: AddGene Entry: pFUS_B10 URL:
            https://www.addgene.org/browse/sequence/14207/ GENERAL Depositing
            Labs: Adam Bogdanove, Daniel Voytas Publication: Cermak et al
            Nucleic Acids Res. 2011 Apr 14. ():. BACKBONE Backbone
            manufacturer: Invitrogen Backbone size w/o insert (bp): 2447 Vector
            backbone: pCR8 CLONING INFORMATION 3′ cloning site: XbaI (not
            destroyed) 3′ sequencing primer: pCR8_R1 (cgaaccgaacaggcttatgt) 5′
            cloning site: AflII (not destroyed) 5′ sequencing primer: pCR8_F1
            (ttgatgcctggcagttccct) Cloning method: Restriction Enzyme GROWTH IN
            BACTERIA Bacterial Resistance(s): Spectinomycin Copy number: High
            Copy Growth Strain(s): DH5alpha Growth Temperature: 37°C Growth
            instructions: DH10B/LB+antibiotics GENE/INSERT Gene/Insert name:
            LacZ + BsaI restriction sites Insert Size (bp): 459 TERMS AND
            LICENSES Academic/Nonprofit Terms:  Industry Terms:
FEATURES             Location/Qualifiers
     rep_origin      1..589
                     /label="ori"
                     /ApEinfo_revcolor="#d59687"
                     /ApEinfo_fwdcolor="#d59687"
                     /note="direction: RIGHT note: high-copy-number ColE1/pMB1/pBR322/pUC origin of replication"
     primer_bind     490..509
                     /label="pBR322ori-F"
                     /ApEinfo_revcolor="#84b0dc"
                     /ApEinfo_fwdcolor="#84b0dc"
                     /note="note: pBR322 origin, forward primer"
     primer_bind     743..760
                     /label="L4440"
                     /ApEinfo_revcolor="#ffef86"
                     /ApEinfo_fwdcolor="#ffef86"
                     /note="note: L4440 vector, forward primer"
     terminator      919..946
                     /label="rrnB T2 terminator"
                     /ApEinfo_revcolor="#c6c9d1"
                     /ApEinfo_fwdcolor="#c6c9d1"
                     /note="note: transcription terminator T2 from the E. coli rrnB gene"
     terminator      1038..1124
                     /label="rrnB T1 terminator"
                     /ApEinfo_revcolor="#9eafd2"
                     /ApEinfo_fwdcolor="#9eafd2"
                     /note="gene: Escherichia coli rrnB note: transcription terminator T1 from the E. coli rrnB gene"
     CDS             complement(1045..1551)
                     /label="lacZ-alpha"
                     /ApEinfo_revcolor="#b4abac"
                     /ApEinfo_fwdcolor="#b4abac"
                     /note="gene: lacZ fragment product: LacZ-alpha fragment of beta-galactosidase translation: MTMITPSAQLTLTKGNKSWRTSRGGPVPNSPYSESYYARSLAVVLQRRDWENPGVTQLNRLAAHPPFASWRNSEEARTDRPSQQLRSLNGEWDAPCSGAGLDSHHLVRGQAMRRLRLAVVLQRRDWENIHASVNARVGNCQASNKTKGSVGRLGLSFYLLFVGERSPE"
     CDS             complement(1045..1551)
                     /ApEinfo_revcolor="#84b0dc"
                     /ApEinfo_fwdcolor="#84b0dc"
                     /translation="MTMITPSAQLTLTKGNKSWRTSRGGPVPNSPYSESYYARSLAVVLQRRDWENPGVTQLNRLAAHPPFASWRNSEEARTDRPSQQLRSLNGEWDAPCSGAGLVPHHLVRGQAMRRLRLAVVLQRRDWENIHASVNARVGNCQASNKTKGSVGRLGLSFYLLFVGERSPE*"
     primer_bind     1188..1204
                     /label="M13 fwd"
                     /ApEinfo_revcolor="#c6c9d1"
                     /ApEinfo_fwdcolor="#c6c9d1"
                     /note="note: common sequencing primer, one of multiple similar variants"
     misc_feature    1244..1247
                     /label="BsaI site"
                     /ApEinfo_revcolor="#b1ff67"
                     /ApEinfo_fwdcolor="#b1ff67"
     primer_bind     1416..1432
                     /label="M13 fwd"
                     /ApEinfo_revcolor="#f58a5e"
                     /ApEinfo_fwdcolor="#f58a5e"
                     /note="note: common sequencing primer, one of multiple similar variants"
     promoter        1442..1460
                     /label="T7 promoter"
                     /ApEinfo_revcolor="#f8d3a9"
                     /ApEinfo_fwdcolor="#f8d3a9"
                     /note="note: promoter for bacteriophage T7 RNA polymerase"
     primer_bind     1442..1461
                     /label="T7"
                     /ApEinfo_revcolor="#85dae9"
                     /ApEinfo_fwdcolor="#85dae9"
                     /note="note: T7 promoter, forward primer"
     promoter        complement(1508..1526)
                     /label="T3 promoter"
                     /ApEinfo_revcolor="#b4abac"
                     /ApEinfo_fwdcolor="#b4abac"
                     /note="note: promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(1508..1528)
                     /label="T3"
                     /ApEinfo_revcolor="#d59687"
                     /ApEinfo_fwdcolor="#d59687"
                     /note="note: T3 promoter, forward primer"
     primer_bind     complement(1547..1563)
                     /label="M13 Reverse"
                     /ApEinfo_revcolor="#faac61"
                     /ApEinfo_fwdcolor="#faac61"
                     /note="note: In lacZ gene. Also called M13-rev"
     primer_bind     complement(1547..1563)
                     /label="M13 rev"
                     /ApEinfo_revcolor="#c6c9d1"
                     /ApEinfo_fwdcolor="#c6c9d1"
                     /note="note: common sequencing primer, one of multiple similar variants"
     primer_bind     complement(1560..1582)
                     /label="M13/pUC Reverse"
                     /ApEinfo_revcolor="#b1ff67"
                     /ApEinfo_fwdcolor="#b1ff67"
                     /note="note: In lacZ gene"
     protein_bind    1571..1587
                     /label="lac operator"
                     /ApEinfo_revcolor="#b1ff67"
                     /ApEinfo_fwdcolor="#b1ff67"
                     /note="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(1595..1625)
                     /label="lac promoter"
                     /ApEinfo_revcolor="#b4abac"
                     /ApEinfo_fwdcolor="#b4abac"
                     /note="note: promoter for the E. coli lac operon"
     protein_bind    1640..1661
                     /label="CAP binding site"
                     /ApEinfo_revcolor="#faac61"
                     /ApEinfo_fwdcolor="#faac61"
                     /note="bound_moiety: E. coli catabolite activator protein note: CAP binding activates transcription in the presence of cAMP."
     misc_feature    complement(1703..1706)
                     /label="BsaI site"
                     /ApEinfo_revcolor="#b1ff67"
                     /ApEinfo_fwdcolor="#b1ff67"
     CDS             2158..2949
                     /label="SmR"
                     /ApEinfo_revcolor="#f58a5e"
                     /ApEinfo_fwdcolor="#f58a5e"
                     /note="gene: aadA product: aminoglycoside adenylyltransferase (Murphy, 1985) note: confers resistance to spectinomycin and streptomycin translation: MREAVIAEVSTQLSEVVGVIERHLEPTLLAVHLYGSAVDGGLKPHSDIDLLVTVTVRLDETTRRALINDLLETSASPGESEILRAVEVTIVVHDDIIPWRYPAKRELQFGEWQRNDILAGIFEPATIDIDLAILLTKAREHSVALVGPAAEELFDPVPEQDLFEALNETLTLWNSPPDWAGDERNVVLTLSRIWYSAVTGKIAPKDVAADWAMERLPAQYQPVILEARQAYLGQEEDRLASRADQLEEFVHYVKGEITKVVGK"
     CDS             2158..2949
                     /ApEinfo_revcolor="#84b0dc"
                     /ApEinfo_fwdcolor="#84b0dc"
                     /translation="MREAVIAEVSTQLSEVVGVIERHLEPTLLAVHLYGSAVDGGLKPHSDIDLLVTVTVRLDETTRRALINDLLETSASPGESEILRAVEVTIVVHDDIIPWRYPAKRELQFGEWQRNDILAGIFEPATIDIDLAILLTKAREHSVALVGPAAEELFDPVPEQDLFEALNETLTLWNSPPDWAGDERNVVLTLSRIWYSAVTGKIAPKDVAADWAMERLPAQYQPVILEARQAYLGQEEDRLASRADQLEEFVHYVKGEITKVVGK*"
ORIGIN
        1 ttgagatcct ttttttctgc gcgtaatctg ctgcttgcaa acaaaaaaac caccgctacc
       61 agcggtggtt tgtttgccgg atcaagagct accaactctt tttccgaagg taactggctt
      121 cagcagagcg cagataccaa atactgtcct tctagtgtag ccgtagttag gccaccactt
      181 caagaactct gtagcaccgc ctacatacct cgctctgcta atcctgttac cagtggctgc
      241 tgccagtggc gataagtcgt gtcttaccgg gttggactca agacgatagt taccggataa
      301 ggcgcagcgg tcgggctgaa cggggggttc gtgcacacag cccagcttgg agcgaacgac
      361 ctacaccgaa ctgagatacc tacagcgtga gcattgagaa agcgccacgc ttcccgaagg
      421 gagaaaggcg gacaggtatc cggtaagcgg cagggtcgga acaggagagc gcacgaggga
      481 gcttccaggg ggaaacgcct ggtatcttta tagtcctgtc gggtttcgcc acctctgact
      541 tgagcgtcga tttttgtgat gctcgtcagg ggggcggagc ctatggaaaa acgccagcaa
      601 cgcggccttt ttacggttcc tggccttttg ctggcctttt gctcacatgt tctttcctgc
      661 gttatcccct gattctgtgg ataaccgtat taccgccttt gagtgagctg ataccgctcg
      721 ccgcagccga acgaccgagc gcagcgagtc agtgagcgag gaagcggaag agcgcccaat
      781 acgcaaaccg cctctccccg cgcgttggcc gattcattaa tgcagctggc acgacaggtt
      841 tcccgactgg aaagcgggca gtgagcgcaa cgcaattaat acgcgtaccg ctagccagga
      901 agagtttgta gaaacgcaaa aaggccatcc gtcaggatgg ccttctgctt agtttgatgc
      961 ctggcagttt atggcgggcg tcctgcccgc caccctccgg gccgttgctt cacaacgttc
     1021 aaatccgctc ccggcggatt tgtcctactc aggagagcgt tcaccgacaa acaacagata
     1081 aaacgaaagg cccagtcttc cgactgagcc tttcgtttta tttgatgcct ggcagttccc
     1141 tactctcgcg ttaacgctag catggatgtt ttcccagtca cgacgttgta aaacgacggc
     1201 cagtcttaag cgtctcatgg cctgaccccg gaccaagtgg tggGGTAcga gaccggcgcc
     1261 gctacagggc gcgtcccatt cgccattcag gctgcgcaac tgttgggaag ggcgatcggt
     1321 gcgggcctct tcgctattac gccagctggc gaaaggggga tgtgctgcaa ggcgattaag
     1381 ttgggtaacg ccagggtttt cccagtcacg acgttgtaaa acgacggcca gtgagcgcgc
     1441 gtaatacgac tcactatagg gcgaattggg taccgggccc cccctcgagg tcctccagct
     1501 tttgttccct ttagtgaggg ttaattgcgc gcttggcgta atcatggtca tagctgtttc
     1561 ctgtgtgaaa ttgttatccg ctcacaattc cacacaacat acgagccgga agcataaagt
     1621 gtaaagcctg gggtgcctaa tgagtgagct aactcacatt aattgcgttg cgctcactgc
     1681 ccgctttcca ccggtggtct ctCGTAgcaa gcaagcgctc gaaacggtgc agcggctgtt
     1741 gccggtgctg tgccaggacc atggcctgac cccggaccaa gtggtggcta tcgagacgtc
     1801 tagaccagcc aggacagaaa tgcctcgact tcgctgctac ccaaggttgc cgggtgacgc
     1861 acaccgtgga aacggatgaa ggcacgaacc cagtggacat aagcctgttc ggttcgtaag
     1921 ctgtaatgca agtagcgtat gcgctcacgc aactggtcca gaaccttgac cgaacgcagc
     1981 ggtggtaacg gcgcagtggc ggttttcatg gcttgttatg actgtttttt tggggtacag
     2041 tctatgcctc gggcatccaa gcagcaagcg cgttacgccg tgggtcgatg tttgatgtta
     2101 tggagcagca acgatgttac gcagcagggc agtcgcccta aaacaaagtt aaacattatg
     2161 agggaagcgg tgatcgccga agtatcgact caactatcag aggtagttgg cgtcatcgag
     2221 cgccatctcg aaccgacgtt gctggccgta catttgtacg gctccgcagt ggatggcggc
     2281 ctgaagccac acagtgatat tgatttgctg gttacggtga ccgtaaggct tgatgaaaca
     2341 acgcggcgag ctttgatcaa cgaccttttg gaaacttcgg cttcccctgg agagagcgag
     2401 attctccgcg ctgtagaagt caccattgtt gtgcacgacg acatcattcc gtggcgttat
     2461 ccagctaagc gcgaactgca atttggagaa tggcagcgca atgacattct tgcaggtatc
     2521 ttcgagccag ccacgatcga cattgatctg gctatcttgc tgacaaaagc aagagaacat
     2581 agcgttgcct tggtaggtcc agcggcggag gaactctttg atccggttcc tgaacaggat
     2641 ctatttgagg cgctaaatga aaccttaacg ctatggaact cgccgcccga ctgggctggc
     2701 gatgagcgaa atgtagtgct tacgttgtcc cgcatttggt acagcgcagt aaccggcaaa
     2761 atcgcgccga aggatgtcgc tgccgactgg gcaatggagc gcctgccggc ccagtatcag
     2821 cccgtcatac ttgaagctag acaggcttat cttggacaag aagaagatcg cttggcctcg
     2881 cgcgcagatc agttggaaga atttgtccac tacgtgaaag gcgagatcac caaggtagtc
     2941 ggcaaataac cctcgagcca cccatgacca aaatccctta acgtgagtta cgcgtcgttc
     3001 cactgagcgt cagaccccgt agaaaagatc aaaggatctt c
//