LOCUS       pSL811_cr-ki_C_C        5604 bp    DNA     circular SYN 19-AUG-2024
DEFINITION  synthetic circular DNA
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5604)
  AUTHORS   MRC Lab University of Dundee Allison Bridges
  TITLE     Direct Submission
  JOURNAL   Exported Apr 9, 2025 from SnapGene 8.0.2
            https://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..5604
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      1..589
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(751..769)
                     /label=oSL802_p_rev
     CDS             772..864
                     /codon_start=1
                     /label=lacI (fragment)
                     /note="pLannotate"
                     /note="/database=snapgene"
                     /note="/fragment=True"
                     /note="/identity=100.0"
                     /note="/match_length=8.6"
                     /note="/other=CDS"
                     /translation="APNTQTASPRALADSLMQLARQVSRLESGQ"
     protein_bind    877..898
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(913..918,919..936,937..943)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    951..967
                     /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)."
     primer_bind     975..991
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     promoter        1012..1030
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     misc_feature    1019..1043
                     /label=Overlap_1
     primer_bind     complement(1019..1043)
                     /label=oSL802_p_rev_60
     misc_feature    1043
                     /label=MCS (fragment)
                     /note="pLannotate"
                     /note="/database=snapgene"
                     /note="/fragment=True"
                     /note="/identity=100.0"
                     /note="/match_length=27.8"
                     /note="/other=misc_feature"
     misc_feature    1044..1051
                     /standard_name="BspQI"
                     /label=BspQI
     CDS             1052..1115
                     /codon_start=1
                     /label=Exon13
                     /translation="KRAALREALAQKMKQNIMGAN"
     misc_feature    1052..1101
                     /label=C-term_CFAP36_50bp upstream
     misc_feature    join(1085..1101,1105..1107)
                     /label=crispr target GAAGATGAAGCAAAACATCATGG
     primer_bind     1088..1117
                     /label=oSL821_Ex13_fwd
     misc_feature    1101^1102
                     /label=DSB
     misc_feature    1102..1115
                     /label=C-term_CFAP36_50bp_downstream
     CDS             1116..1121
                     /codon_start=1
                     /label=Linker
                     /translation="GG"
     CDS             1122..1145
                     /codon_start=1
                     /product="recognition and cleavage site for human 
                     rhinovirus 3C and PreScission proteases"
                     /label=HRV 3C site
                     /translation="LEVLFQGP"
     primer_bind     complement(1136..1151)
                     /label=oSL86_mSG_rev_68
     CDS             1146..1154
                     /codon_start=1
                     /label=Linker
                     /translation="GGG"
     CDS             1155..1802
                     /codon_start=1
                     /label=mStayGold
                     /translation="ASTPFKFQLKGTINGKSFTVEGEGEGNSHEGSHKGKYVCTSGKLP
                     MSWAALGTSFGYGMKYYTKYPSGLKNWFHEVMPEGFTYDRHIQYKGDGSIHAKHQHFMK
                     NGTYHNIVEFTGQDFKENSPVLTGDMNVSLPNDVQHIPRDDGVECPVTLLYPLLSDKSK
                     CVEAHQNTICKPLHNQPAPDVPYHWIRKQYTQSKDDTEERDHICQSETLEAHL"
     primer_bind     complement(1186..1204)
                     /label=mSG_N_rev
     primer_bind     1742..1758
                     /label=mSG_C_fwd
     primer_bind     1803..1835
                     /label=oSL807_2Strep_fwd
     CDS             1803..1814
                     /codon_start=1
                     /label=Linker
                     /translation="GSGS"
     CDS             join(1815..1838,1839..1874,1875..1898)
                     /codon_start=1
                     /product="two Strep-Tag(R)II moieties connected by a linker
                     for enhanced binding to Strep-Tactin(R), an engineered form
                     of streptavidin (Schmidt et al., 2013)"
                     /label=Twin-Strep-tag
                     /translation="WSHPQFEKGGGSGGGSGGSAWSHPQFEK"
     3'UTR           1908..2413
                     /label=3'UTR CFAP36
     primer_bind     complement(2437..2489)
                     /label=pSL810_p2_rev_65
     misc_feature    2437..2465
                     /label=Overlap_2
     promoter        2443..2465
                     /label=beta-tubulin promoter
     gene            2443..2465
                     /label=hyg
     primer_bind     2466..2489
                     /label=crBSDr_fwd_64
     promoter        2486..2701
                     /standard_name="rbcS2-P"
                     /label=rbcS2-P promoter
     intron          2710..2854
                     /locus_tag="rbcS2 intron1"
                     /standard_name="rbcS2-i1"
                     /label=rbcS2-i1
     CDS             2874..3296
                     /standard_name="Blasticidin S Deaminase"
                     /label=Blasticidin S Deaminase
     3'UTR           3312..3545
                     /locus_tag="rbcS2 3' UTR"
                     /standard_name="rbcS2-T"
                     /label=rbcS2-T
     misc_feature    3524..3550
                     /label=Overlap_3
     primer_bind     complement(3524..3545)
                     /label=crBSDr_rev_65
     primer_bind     3524..3545
                     /label=pSL810_p1_fwd_65
     misc_feature    3551..3600
                     /label=C-term_CFAP36_50bp_downstream
     CDS             3551..3567
                     /codon_start=1
                     /label=Exon13
                     /translation="MGAN"
     misc_feature    3554..3556
                     /label=crispr target GAAGATGAAGCAAAACATCATGG
     3'UTR           3568..3600
                     /label=3'UTR CFAP36
     misc_feature    complement(3601..3607)
                     /standard_name="BspQI"
                     /label=BspQI
     misc_feature    3637..3661
                     /standard_name="Ligation"
                     /label=Ligation
                     /note="Geneious type: ligation"
     misc_feature    3697..3796
                     /gene="aacC1"
                     /product="gentamycin acetyltransferase"
                     /label=filler_100bp
                     /note="confers resistance to gentamycin"
     misc_feature    3797..3823
                     /label=Overlap_4
     primer_bind     3797..3822
                     /label=oSL802_p_fwd
     promoter        complement(3802..3820)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             3828..3983
                     /codon_start=1
                     /label=lacZ-alpha (fragment)
                     /note="pLannotate"
                     /note="/database=snapgene"
                     /note="/fragment=True"
                     /note="/identity=100.0"
                     /note="/match_length=89.7"
                     /note="/other=CDS"
                     /translation="SLAVVLQRRDWENPGVTQLNRLAAHPPFASWRNSEEARTDRPSQQ
                     LRSLNGE"
     primer_bind     complement(3830..3846)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     rep_origin      3988..4443
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     promoter        4469..4573
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(4574..4642,4643..5434)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
ORIGIN
        1 ttgagatcct ttttttctgc gcgtaatctg ctgcttgcaa acaaaaaaac caccgctacc
       61 agcggtggtt tgtttgccgg atcaagagct accaactctt tttccgaagg taactggctt
      121 cagcagagcg cagataccaa atactgttct 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 gctatgagaa 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 gtgagttagc tcactcatta
      901 ggcaccccag gctttacact ttatgcttcc ggctcgtatg ttgtgtggaa ttgtgagcgg
      961 ataacaattt cacacaggaa acagctatga ccatgattac gccaagcgcg caattaaccc
     1021 tcactaaagg gaacaaaagc tgggctcttc tcaagcgcgc tgcgctgcgt gaggccctgg
     1081 cgcagaagat gaagcaaaac atcatgggag caaacggtgg cctggaagtt ctgttccagg
     1141 ggcccggcgg cggtgccagc acccccttca agttccagct gaagggcacc atcaacggca
     1201 agagcttcac cgtggagggc gagggcgagg gcaacagcca cgagggcagc cacaagggca
     1261 agtacgtgtg caccagcggc aagctgccca tgagctgggc cgccctgggc accagcttcg
     1321 gctacggcat gaagtactac accaagtacc ccagcggcct gaagaactgg ttccacgagg
     1381 tgatgcccga gggcttcacc tacgacagac acatccagta caagggcgac ggcagcatcc
     1441 acgccaagca ccagcacttc atgaagaacg gcacctacca caacatcgtg gagttcaccg
     1501 gccaggactt caaggagaac agccccgtgc tgaccggcga catgaacgtg agcctgccca
     1561 acgacgtgca gcacatcccc agagacgacg gcgtggagtg ccccgtgacc ctgctgtacc
     1621 ccctgctgag cgacaagagc aagtgcgtgg aggcccacca gaacaccatc tgcaagcccc
     1681 tgcacaacca gcccgccccc gacgtgccct accactggat cagaaagcag tacacccaga
     1741 gcaaggacga caccgaggag agagaccaca tctgccagag cgagaccctg gaggcccacc
     1801 tgggttctgg ttcttggagt catccacagt tcgagaaggg cgggggctcc ggaggtggat
     1861 caggaggttc cgcctggtca caccctcaat tcgagaaggg tgaatgagga gctgggagca
     1921 tcgagagcac gggcgggggt gcgcgggggc agagcgagat gctgacgttg ccagaggtcc
     1981 ggtttggcag gtggtgactg atgtgagggg tgagcgcagg tggtgccgct ggaccgaaat
     2041 attgtagagg aggcatgcag gcgagcgatt gcagtgtgcg gacgtgaagt gcaataggtg
     2101 ctgcaggtgt tttgggttgc cgtgattcgc gatagcagga ctgagtttgt gcagaggtgg
     2161 tttgcaagcg aaatgggttt ggcttattat tggcttgtgc cttccggcgt tgttctggcg
     2221 acctttgcca aagtgccttt ggccagtgcc atgccgcgcg catgtgtagt catcttcaac
     2281 gggcattgtc atggcattgt cactcagcta atggtatgtc aaacaccgct gagtagttcg
     2341 aggagaagag gatgtcaaac gccgctgagt atgcacgccc cagcctcggg cgcacagtgt
     2401 cacaatcagc actacgctgc agctcttagt actgagggta ccctttcttg cgctatgaca
     2461 cttccgatcc catcaagctt gcatgccggg cgcgccagaa ggagcgcagc caaaccagga
     2521 tgatgtttga tggggtattt gagcacttgc aacccttatc cggaagcccc ctggcccaca
     2581 aaggctaggc gccaatgcaa gcagttcgca tgcagcccct ggagcggtgc cctcctgata
     2641 aaccggccag ggggcctatg ttctttactt ttttacaaga gaagtcactc aacatcttaa
     2701 aatggccagg tgagtcgacg agcaagcccg gcggatcagg cagcgtgctt gcagatttga
     2761 cttgcaacgc ccgcattgtg tcgacgaagg cttttggctc ctctgtcgct gtctcaagca
     2821 gcatctaacc ctgcgtcgcc gtttccattt gcagctcgag cgacgtcatg gccatgaaaa
     2881 ccttcaacat cagccagcag gacctggagc tggtggaggt ggccaccgag aagatcacca
     2941 tgctgtacga ggacaacaag caccacgtgg gcgccgccat ccgcaccaag accggcgaga
     3001 tcatcagcgc cgtgcacatc gaggcctaca tcggccgcgt gaccgtgtgc gccgaggcca
     3061 tcgccatcgg cagcgccgtg agcaacggcc agaaggactt cgacaccatc gtggccgtgc
     3121 gccaccccta cagcgacgag gtggaccgca gcatccgcgt ggtgagcccc tgcggcatgt
     3181 gccgcgagct gatcagcgac tacgcccccg actgcttcgt gctgatcgag atgaacggca
     3241 agctggtcaa gaccaccatc gaggagctga tccccctgaa gtacacccgc aactaagctt
     3301 tgagaccatc cccgctccgt gtaaatggag gcgctcgttg atctgagcct tgccccctga
     3361 cgaacggcgg tggatggaag atactgctct caagtgctga agcggtagct tagctccccg
     3421 tttcgtgctg atcagtcttt ttcaacacgt aaaaagcgga ggagttttgc aattttgttg
     3481 gttgtaacga tcctccgttg attttggcct ctttctccat gggcgggctg ggcgtatttg
     3541 aagcgggtac tcatgggagc aaactgagga gctgggagca tcgagagcac gggcgggggt
     3601 gaagagccaa acgagcactt atacatacgc gactattctg ccgctataca taaccactca
     3661 ggagctctat gtgacaatgc ttccgtgacg ccgcgaggta cttgggtcga tatcaaagtg
     3721 catcacttct tcccgtatgc ccaactttgt atagagagcc actgcgggat cgtcaccgta
     3781 atctgcttgc acgtagttcg ccctatagtg agtcgtatta cgcgcgctca ctggccgtcg
     3841 ttttacaacg tcgtgactgg gaaaaccctg gcgttaccca acttaatcgc cttgcagcac
     3901 atcccccttt cgccagctgg cgtaatagcg aagaggcccg caccgatcgc ccttcccaac
     3961 agttgcgcag cctgaatggc gaatgggacg cgccctgtag cggcgcatta agcgcggcgg
     4021 gtgtggtggt tacgcgcagc gtgaccgcta cacttgccag cgccctagcg cccgctcctt
     4081 tcgctttctt cccttccttt ctcgccacgt tcgccggctt tccccgtcaa gctctaaatc
     4141 gggggctccc tttagggttc cgatttagtg ctttacggca cctcgacccc aaaaaacttg
     4201 attagggtga tggttcacgt agtgggccat cgccctgata gacggttttt cgccctttga
     4261 cgttggagtc cacgttcttt aatagtggac tcttgttcca aactggaaca acactcaacc
     4321 ctatctcggt ctattctttt gatttataag ggattttgcc gatttcggcc tattggttaa
     4381 aaaatgagct gatttaacaa aaatttaacg cgaattttaa caaaatatta acgcttacaa
     4441 tttaggtggc acttttcggg gaaatgtgcg cggaacccct atttgtttat ttttctaaat
     4501 acattcaaat atgtatccgc tcatgagaca ataaccctga taaatgcttc aataatattg
     4561 aaaaaggaag agtatgagta ttcaacattt ccgtgtcgcc cttattccct tttttgcggc
     4621 attttgcctt cctgtttttg ctcacccaga aacgctggtg aaagtaaaag atgctgaaga
     4681 tcagttgggt gcacgagtgg gttacatcga actggatctc aacagcggta agatccttga
     4741 gagttttcgc cccgaagaac gttttccaat gatgagcact tttaaagttc tgctatgtgg
     4801 cgcggtatta tcccgtattg acgccgggca agagcaactc ggtcgccgca tacactattc
     4861 tcagaatgac ttggttgagt actcaccagt cacagaaaag catcttacgg atggcatgac
     4921 agtaagagaa ttatgcagtg ctgccataac catgagtgat aacactgcgg ccaacttact
     4981 tctgacaacg atcggaggac cgaaggagct aaccgctttt ttgcacaaca tgggggatca
     5041 tgtaactcgc cttgatcgtt gggaaccgga gctgaatgaa gccataccaa acgacgagcg
     5101 tgacaccacg atgcctgtag caatggcaac aacgttgcgc aaactattaa ctggcgaact
     5161 acttactcta gcttcccggc aacaattaat agactggatg gaggcggata aagttgcagg
     5221 accacttctg cgctcggccc ttccggctgg ctggtttatt gctgataaat ctggagccgg
     5281 tgagcgtggg tctcgcggta tcattgcagc actggggcca gatggtaagc cctcccgtat
     5341 cgtagttatc tacacgacgg ggagtcaggc aactatggat gaacgaaata gacagatcgc
     5401 tgagataggt gcctcactga ttaagcattg gtaactgtca gaccaagttt actcatatat
     5461 actttagatt gatttaaaac ttcattttta atttaaaagg atctaggtga agatcctttt
     5521 tgataatctc atgaccaaaa tcccttaacg tgagttttcg ttccactgag cgtcagaccc
     5581 cgtagaaaag atcaaaggat cttc
//
