LOCUS       pEN-L4-AG-R1            4089 bp    DNA     circular     28-JUL-2022
DEFINITION  .
ACCESSION   p00046
KEYWORDS    .
SOURCE      
  ORGANISM  .
FEATURES             Location/Qualifiers
     attL4           1..97
                     /created_by="mamee"
                     /modified_by="mamee"
                     /note="Derived using Geneious Prime 2022.1.1 'Annotate
                     from Database' based on nucleotide similarity"
                     /Transferred_From="attL4"
                     /Transferred_Similarity="100.00%"
                     /Primary_Match="100.00%; Adjusted-Interval-0:0/0; <a
                     href=""urn:local:.:lr-eztz0t4"">attL4</a>"
     GG              complement(100..110)
                     /created_by="thjac"
                     /modified_by="thjac"
                     /note="Derived using Geneious Prime 2020.2.2 'Annotate
                     from Database' based on nucleotide similarity"
                     /Transferred_From="A"
                     /Transferred_Similarity="100.00%"
                     /Primary_Match="100.00%; Adjusted-Interval-0:0/0; <a
                     href=""urn:local:.:85-dgycryy"">A</a>"
                     /label="A"
     CDS             complement(117..422)
                     /Transferred_From="ccdB"
                     /Transferred_Similarity="99.67%"
                     /label="ccdB"
     misc_feature    204
                     /Original_Bases="G"
                     /label="G"
                     /note="Geneious type: Editing History Replacement"
     misc_feature    207
                     /Original_Bases="T"
                     /label="T"
                     /note="Geneious type: Editing History Replacement"
     primer_bind     377..397
                     /%GC="47.6"
                     /Hairpin_Tm="45.0"
                     /Self_Dimer_Tm="None"
                     /Tm="57.6"
                     /annotation_group="66881012735019009151d93c-20a9-405a-a2f2
                     -e5a52455d32a"
                     /created_by="primer3"
                     /Sequence="GACGATAACGGCTCTCTCTTT"
                     /Mismatches=0
                     /#_Off-target_Sites=0
                     /Off-target_Template="pEN-L4-AG-R1"
                     /label="P15"
     UTR             complement(423..741)
                     /Mismatches=1
                     /%_Identity="99.68652037617555"
                     /Motif="AAGCCAGATAACAGTATGCGTATTTGCGCGCTGATTTTTGCGGTATAAGA
                     ATATATACTGATATGTATACCCGAAGTATGTCAAAAGGAGGTATGCTATGAAGCAGCG
                     TATTACAGTGACAGTTGACAGCGACAGCTATCAGTTGCTCAAGGCATATATGATGTCA
                     ATATCTCCGGTCTGGTAAGCACAACCATGCAGAATGAAGCCCGTCGTCTGCGTGCCGA
                     ACGCTGGAAAGCGGAAAATCAGGAAGGGATGGCTGAGGTCGCCCGGTTTATTGAAATG
                     AACGGCTCTTTTGCTGACGAGAACAGGGGCTGGTGAA"
                     /note="Derived using Geneious Prime 2020.2.2 'Annotate
                     from Database' based on nucleotide similarity"
                     /Transferred_From="ccdB"
                     /Transferred_Similarity="99.37%"
                     /Primary_Match="99.37%; Adjusted-Interval-0:0/0; ccdB
                     5'UTR (<a href=""urn:local:.:8r-dgycryy"">ccdB</a>)"
                     /label="ccdB 5'UTR"
     misc_feature    431
                     /Original_Bases="T"
                     /label="T"
                     /note="Geneious type: Editing History Replacement"
     misc_feature    649
                     /Original_Bases="C"
                     /label="C"
                     /note="Geneious type: Editing History Replacement"
     misc_feature    complement(764..1423)
                     /Transferred_From="cmR"
                     /Transferred_Similarity="99.85%"
                     /modified_by="mamee"
                     /note="Derived using Geneious Prime 2022.1.1 'Annotate
                     from Database' based on nucleotide similarity"
                     /Transferred_From="CmR"
                     /Transferred_Similarity="100.00%"
                     /Primary_Match="100.00%; Adjusted-Interval-0:0/0; <a
                     href=""urn:local:.:29-f0o4v7s"">CmR</a>"
                     /label="CmR"
                     /note="Geneious type: Bacterial Selection"
     misc_feature    839
                     /Original_Bases="G"
                     /label="G"
                     /note="Geneious type: Editing History Replacement"
     primer_bind     1174..1193
                     /%GC="55.0"
                     /Hairpin_Tm="None"
                     /Self_Dimer_Tm="None"
                     /Tm="60.0"
                     /annotation_group="6688101329925400586640ae-62f7-4e6b-a47e
                     -ebaac140519e"
                     /created_by="primer3"
                     /Sequence="TCACCAGCTCACCGTCTTTC"
                     /Mismatches=0
                     /#_Off-target_Sites=0
                     /Off-target_Template="pEN-L4-AG-R1"
                     /label="P16"
     misc_feature    1453
                     /Original_Bases="C"
                     /label="C"
                     /note="Geneious type: Editing History Replacement"
     promoter        complement(1477..1507)
                     /note="Lac promoter of Escherichia coli containing 2 base
                     pair mutations"
                     /regulatory_class="promoter"
                     /note="Derived using Geneious Prime 2020.2.4 'Annotate
                     from Database' based on nucleotide similarity"
                     /X-Transferred_From="lacUV5"
                     /X-Transferred_Similarity="100.00%"
                     /X-Primary_Match="100.00%; Adjusted-Interval-0:0/0; <a
                     href=""urn:local:.:1g0-c949y1m"">lacUV5</a>"
                     /note="Derived using Geneious Prime 2022.1.1 'Annotate
                     from Database' based on nucleotide similarity"
                     /Transferred_From="CmR"
                     /Transferred_Similarity="100.00%"
                     /Primary_Match="100.00%; Adjusted-Interval-0:0/0; lacUV5
                     (<a href=""urn:local:.:29-f0o4v7s"">CmR</a>)"
                     /label="lacUV5"
     GG              1532..1542
                     /%_Identity=100
                     /Motif="GGTCTCAGTAT"
                     /note="Derived using Geneious Prime 2020.2.2 'Annotate
                     from Database' based on nucleotide similarity"
                     /Transferred_From="G"
                     /Transferred_Similarity="100.00%"
                     /Primary_Match="100.00%; Adjusted-Interval-0:0/0; G
                     overlap (<a href=""urn:local:.:86-dgycryy"">G</a>)"
                     /modified_by="mamee"
                     /label="G"
     attR1           1544..1700
                     /created_by="mamee"
                     /Color="#00cccc"
                     /modified_by="mamee"
                     /note="Derived using Geneious Prime 2022.1.1 'Annotate
                     from Database' based on nucleotide similarity"
                     /Transferred_From="attR1"
                     /Transferred_Similarity="99.36%"
                     /Primary_Match="99.36%; Adjusted-Interval-0:0/0; <a
                     href=""urn:local:.:y-f0o11e8"">attR1</a>"
     misc_feature    1670
                     /Original_Bases="T"
                     /label="T"
                     /note="Geneious type: Editing History Replacement"
     promoter        complement(1741..1758)
                     /created_by="mamee"
                     /note="Derived using Geneious Prime 2022.1.1 'Annotate
                     from Database' based on nucleotide similarity"
                     /Transferred_From="T7 promoter"
                     /Transferred_Similarity="100.00%"
                     /Primary_Match="100.00%; Adjusted-Interval-0:0/0; <a
                     href=""urn:local:.:yz-f3tbldq"">T7 promoter</a>"
                     /label="T7 promoter"
     primer          complement(1763..1779)
                     /invitrogen=2010000
                     /note="Derived using Geneious Prime 2020.2.2 'Annotate
                     from Database' based on nucleotide similarity"
                     /Transferred_From="pDONRP4-P3 - M13 reverse primer"
                     /Transferred_Similarity="100.00%"
                     /Primary_Match="100.00%; Adjusted-Interval-0:0/0; M13
                     reverse primer (pDONRP4-P3 - M13 reverse primer)"
                     /label="M13 reverse primer"
     promoter        1841..1891
                     /created_by="chgai"
                     /X-Transferred_From="pCG034_pKan_Kan-CBE"
                     /X-Transferred_Similarity="100.00%"
                     /X-Primary_Match="100.00%; Adjusted-Interval-0:0/0;
                     Kan_promoter (<a
                     href=""urn:local:chgai:ll-ch8iv1b"">pCG034_pKan_Kan-CBE</a
                     >)"
                     /X-Alternative_Match="100.00%; Adjusted-Interval-0:0/0;
                     Kan_promoter (<a
                     href=""urn:local:chgai:lo-ch8iv1b"">pCG040_ABE-Nuc-NLS_Kan
                     -reporter_control</a>)"
                     /X-Alternative_Match="100.00%; Adjusted-Interval-0:0/0;
                     Kan_promoter (<a
                     href=""urn:local:chgai:lr-ch8iv1b"">pCG043_Kan-ABE-Dest_co
                     ntrol_ccdB</a>)"
                     /X-Alternative_Match="100.00%; Adjusted-Interval-0:0/0;
                     Kan_promoter (<a
                     href=""urn:local:chgai:lu-ch8iv1c"">pCG046_ABE-LbCas12a-NL
                     S_Kan_control</a>)"
                     /X-Alternative_Match="100.00%; Adjusted-Interval-0:0/0;
                     Kan_promoter (<a
                     href=""urn:local:chgai:kx-ch8iv19"">pCG005_pGGD-pKan-KanR-
                     E</a>)"
                     /X-Alternative_Match="100.00%; Adjusted-Interval-0:0/0;
                     Kan_promoter (<a
                     href=""urn:local:chgai:ky-ch8iv19"">pCG006_pGGA-Kan_promot
                     er-B</a>)"
                     /X-Alternative_Match="100.00%; Adjusted-Interval-0:0/0;
                     Kan_promoter (<a
                     href=""urn:local:chgai:lm-ch8iv1b"">pCG035_pKan_Kan</a>)"
                     /X-Alternative_Match="100.00%; Adjusted-Interval-0:0/0;
                     Kan_promoter (<a
                     href=""urn:local:chgai:ln-ch8iv1b"">pCG039_ABE-N7-NLS_Kan-
                     reporter_control</a>)"
                     /X-Alternative_Match="100.00%; Adjusted-Interval-0:0/0;
                     Kan_promoter (<a
                     href=""urn:local:chgai:lp-ch8iv1b"">pCG041_ABE-SV40-NLS_Ka
                     n-reporter_control</a>)"
                     /X-Alternative_Match="100.00%; Adjusted-Interval-0:0/0;
                     Kan_promoter (<a
                     href=""urn:local:chgai:lw-ch8iv1c"">pCG054_Kan_CBE_control
                     </a>)"
                     /modified_by="mamee"
                     /note="Derived using Geneious Prime 2022.1.1 'Annotate
                     from Database' based on nucleotide similarity"
                     /Transferred_From="KanR"
                     /Transferred_Similarity="100.00%"
                     /Primary_Match="100.00%; Adjusted-Interval-0:0/0; kan (<a
                     href=""urn:local:.:8j-dgycryy"">KanR</a>)"
                     /label="kan"
     misc_feature    1892..2701
                     /modified_by="wacae"
                     /Transferred_From="KanR"
                     /Transferred_Similarity="100.00%"
                     /label="KanR"
                     /note="Geneious type: Bacterial Selection"
     primer_bind     1917..1936
                     /%GC="55.0"
                     /Hairpin_Tm="None"
                     /Self_Dimer_Tm="25.9"
                     /Tm="59.8"
                     /annotation_group="668810138659200023b9f5eb-3831-4881-b3d8
                     -2b357ae1f14a"
                     /created_by="primer3"
                     /Sequence="CGAGGCCGCGATTAAATTCC"
                     /Mismatches=0
                     /#_Off-target_Sites=0
                     /Off-target_Template="pEN-L4-AG-R1"
                     /label="P44"
     primer_bind     2551..2570
                     /%GC="60.0"
                     /Hairpin_Tm="40.0"
                     /Self_Dimer_Tm="None"
                     /Tm="60.0"
                     /annotation_group="668810161745040018b90adf-38f8-4190-9e17
                     -abbd27b74b4f"
                     /created_by="primer3"
                     /Sequence="CGCAGACCGATACCAGGATC"
                     /Mismatches=0
                     /#_Off-target_Sites=0
                     /Off-target_Template="pEN-L4-AG-R1"
                     /label="P42"
     rep_origin      2848..3436
                     /created_by="thjac"
                     /modified_by="mamee"
                     /note="Derived using Geneious Prime 2020.2.2 'Annotate
                     from Database' based on nucleotide similarity"
                     /Transferred_From="high-copy-number ColE1/pMB1/pBR322/pUC
                     origin of  replication"
                     /Transferred_Similarity="99.49%"
                     /Primary_Match="99.49%; Adjusted-Interval-0:0/0;
                     ColE1/pMB1/pBR322/pUC Ori (<a
                     href=""urn:local:.:8d-dk4mzop"">high-copy-number
                     ColE1/pMB1/pBR322/pUC origin of  replication</a>)"
                     /label="ColE1/pMB1/pBR322/pUC Ori"
     misc_feature    2995
                     /Original_Bases="C"
                     /label="C"
                     /note="Geneious type: Editing History Replacement"
     misc_feature    3239^3240
                     /Original_Bases="A"
                     /label="A"
                     /note="Geneious type: Editing History Deletion"
     misc_feature    3241
                     /Original_Bases
                     /label
                     /note="Geneious type: Editing History Insertion"
     primer_bind     3336..3355
                     /%GC="55.0"
                     /Hairpin_Tm="37.3"
                     /Self_Dimer_Tm="None"
                     /Tm="59.7"
                     /annotation_group="6688101445398700e009c757-1abc-43ef-9930
                     -a2632257dec3"
                     /created_by="primer3"
                     /Sequence="GGGGAAACGCCTGGTATCTT"
                     /Mismatches=0
                     /#_Off-target_Sites=0
                     /Off-target_Template="pEN-L4-AG-R1"
                     /label="P45"
     primer_bind     3944..3963
                     /%GC="55.0"
                     /Hairpin_Tm="None"
                     /Self_Dimer_Tm="None"
                     /Tm="60.0"
                     /annotation_group="6688101504100200cebea443-fcb0-4391-afba
                     -24260f7a8a6d"
                     /created_by="primer3"
                     /Sequence="CTTCCGACTGAGCCTTTCGT"
                     /Mismatches=0
                     /#_Off-target_Sites=0
                     /Off-target_Template="pEN-L4-AG-R1"
                     /label="P46"
     Primer          4035..4055
                     /created_by="mamee"
                     /modified_by="mamee"
                     /label="GWM13F"
     misc_feature    4068
                     /Original_Bases="C"
                     /label="C"
                     /note="Geneious type: Editing History Replacement"
ORIGIN      
        1 caaataatga ttttattttg actgatagtg acctgttcgt tgcaacaaat tgataagcaa
       61 tgctttttta taatgccaac tttgtataga aaagttgcta ccttgagacc gtcgacttat
      121 attccccaga acatcaggtt aatggcgttt ttgatgtcat tttcgcggtg gctgagatca
      181 gccacttctt ccccgataac ggaaaccggc acactggcca tatcggtggt catcatgcgc
      241 cagctttcat ccccgatatg caccaccggg taaagttcac gggagacttt atctgacagc
      301 agacgtgcac tggccagggg gatcaccatc cgtcgcccgg gcgtgtcaat aatatcactc
      361 tgtacatcca caaacagacg ataacggctc tctcttttat aggtgtaaac cttaaactgc
      421 atttcaccag cccctgttct cgtcagcaaa agagccgttc atttcaataa accgggcgac
      481 ctcagccatc ccttcctgat tttccgcttt ccagcgttcg gcacgcagac gacgggcttc
      541 attctgcatg gttgtgctta ccagaccgga gatattgaca tcatatatgc cttgagcaac
      601 tgatagctgt cgctgtcaac tgtcactgta atacgctgct tcatagcata cctctttttg
      661 acatacttcg ggtatacata tcagtatata ttcttatacc gcaaaaatca gcgcgcaaat
      721 acgcatactg ttatctggct tttagtaagc cggatccacg cgtttacgcc ccgccctgcc
      781 actcatcgca gtactgttgt aattcattaa gcattctgcc gacatggaag ccatcacaaa
      841 cggcatgatg aacctgaatc gccagcggca tcagcacctt gtcgccttgc gtataatatt
      901 tgcccatggt gaaaacgggg gcgaagaagt tgtccatatt ggccacgttt aaatcaaaac
      961 tggtgaaact cacccaggga ttggctgaga cgaaaaacat attctcaata aaccctttag
     1021 ggaaataggc caggttttca ccgtaacacg ccacatcttg cgaatatatg tgtagaaact
     1081 gccggaaatc gtcgtggtat tcactccaga gcgatgaaaa cgtttcagtt tgctcatgga
     1141 aaacggtgta acaagggtga acactatccc atatcaccag ctcaccgtct ttcattgcca
     1201 tacggaattc cggatgagca ttcatcaggc gggcaagaat gtgaataaag gccggataaa
     1261 acttgtgctt atttttcttt acggtcttta aaaaggccgt aatatccagc tgaacggtct
     1321 ggttataggt acattgagca actgactgaa atgcctcaaa atgttcttta cgatgccatt
     1381 gggatatatc aacggtggta tatccagtga tttttttctc cattttagct tccttagctc
     1441 ctgaaaatct cgacggatcc taactcaaaa tccacacatt atacgagccg gaagcataaa
     1501 gtgtaaagcc tggggtgcct aatgcggccg cggtctcagt atgcaagttt gtacaaaaaa
     1561 gttgaacgag aaacgtaaaa tgatataaat atcaatatat taaattagat tttgcataaa
     1621 aaacagacta cataatactg taaaacacaa catatgcagt cactatgaac caactactta
     1681 gatggtatta gtgacctgta gaattcgagc tctagagctg cagggcggcc gcgatatccc
     1741 ctatagtgag tcgtattaca tggtcatagc tgtttcctgg cagctctggc ccgtgtctca
     1801 aaatctctga tgttacattg cacaagataa aaatatatca tcatgaacaa taaaactgtc
     1861 tgcttacata aacagtaata caaggggtgt tatgagccat attcaacggg aaacgtcgag
     1921 gccgcgatta aattccaaca tggatgctga tttatatggg tataaatggg ctcgcgataa
     1981 tgtcgggcaa tcaggtgcga caatctatcg cttgtatggg aagcccgatg cgccagagtt
     2041 gtttctgaaa catggcaaag gtagcgttgc caatgatgtt acagatgaga tggtcagact
     2101 aaactggctg acggaattta tgcctcttcc gaccatcaag cattttatcc gtactcctga
     2161 tgatgcatgg ttactcacca ctgcgatccc cggaaaaaca gcattccagg tattagaaga
     2221 atatcctgat tcaggtgaaa atattgttga tgcgctggca gtgttcctgc gccggttgca
     2281 ttcgattcct gtttgtaatt gtccttttaa cagcgatcgc gtatttcgtc tcgctcaggc
     2341 gcaatcacga atgaataacg gtttggttga tgcgagtgat tttgatgacg agcgtaatgg
     2401 ctggcctgtt gaacaagtct ggaaagaaat gcataaactt ttgccattct caccggattc
     2461 agtcgtcact catggtgatt tctcacttga taaccttatt tttgacgagg ggaaattaat
     2521 aggttgtatt gatgttggac gagtcggaat cgcagaccga taccaggatc ttgccatcct
     2581 atggaactgc ctcggtgagt tttctccttc attacagaaa cggctttttc aaaaatatgg
     2641 tattgataat cctgatatga ataaattgca gtttcatttg atgctcgatg agtttttcta
     2701 atcagaattg gttaattggt tgtaacactg gcagagcatt acgctgactt gacgggacgg
     2761 cgcaagctca tgaccaaaat cccttaacgt gagttacgcg tcgttccact gagcgtcaga
     2821 ccccgtagaa aagatcaaag gatcttcttg agatcctttt tttctgcgcg taatctgctg
     2881 cttgcaaaca aaaaaaccac cgctaccagc ggtggtttgt ttgccggatc aagagctacc
     2941 aactcttttt ccgaaggtaa ctggcttcag cagagcgcag ataccaaata ctgttcttct
     3001 agtgtagccg tagttaggcc accacttcaa gaactctgta gcaccgccta catacctcgc
     3061 tctgctaatc ctgttaccag tggctgctgc cagtggcgat aagtcgtgtc ttaccgggtt
     3121 ggactcaaga cgatagttac cggataaggc gcagcggtcg ggctgaacgg ggggttcgtg
     3181 cacacagccc agcttggagc gaacgaccta caccgaactg agatacctac agcgtgagct
     3241 atgagaaagc gccacgcttc ccgaagggag aaaggcggac aggtatccgg taagcggcag
     3301 ggtcggaaca ggagagcgca cgagggagct tccaggggga aacgcctggt atctttatag
     3361 tcctgtcggg tttcgccacc tctgacttga gcgtcgattt ttgtgatgct cgtcaggggg
     3421 gcggagccta tggaaaaacg ccagcaacgc ggccttttta cggttcctgg ccttttgctg
     3481 gccttttgct cacatgttct ttcctgcgtt atcccctgat tctgtggata accgtattac
     3541 cgcctttgag tgagctgata ccgctcgccg cagccgaacg accgagcgca gcgagtcagt
     3601 gagcgaggaa gcggaagagc gcccaatacg caaaccgcct ctccccgcgc gttggccgat
     3661 tcattaatgc agctggcacg acaggtttcc cgactggaaa gcgggcagtg agcgcaacgc
     3721 aattaatacg cgtaccgcta gccaggaaga gtttgtagaa acgcaaaaag gccatccgtc
     3781 aggatggcct tctgcttagt ttgatgcctg gcagtttatg gcgggcgtcc tgcccgccac
     3841 cctccgggcc gttgcttcac aacgttcaaa tccgctcccg gcggatttgt cctactcagg
     3901 agagcgttca ccgacaaaca acagataaaa cgaaaggccc agtcttccga ctgagccttt
     3961 cgttttattt gatgcctggc agttccctac tctcgcgtta acgctagcat ggatgttttc
     4021 ccagtcacga cgttgtaaaa cgacggccag tcttaagctc gggcccgagt taacgctacc
     4081 atggagctc
//
