LOCUS       pTE5639-SecY(T166L)EG        6013 bp    DNA     circular SYN 19-NOV-2025
DEFINITION  pSecY(T166L)EG - superactive inserter mutant of SecY.
ACCESSION
VERSION     .
KEYWORDS    .
SOURCE      .
  ORGANISM  .
COMMENT     Annotated with pLannotate v1.2.1.
FEATURES             Location/Qualifiers
     rep_origin      1..429
                     /note="pLannotate"
                     /database="snapgene"
                     /identity="100.0"
                     /match_length="100.0"
                     /fragment="False"
                     /other="rep_origin"
                     /label="f1 ori"
     CDS             complement(548..1363)
                     /note="pLannotate"
                     /database="snapgene"
                     /identity="99.8"
                     /match_length="100.0"
                     /fragment="False"
                     /other="CDS"
                     /label="KanR"
     rep_origin      1485..2073
                     /note="pLannotate"
                     /database="snapgene"
                     /identity="100.0"
                     /match_length="100.0"
                     /fragment="False"
                     /other="rep_origin"
                     /label="ori"
     CDS             complement(2500..2691)
                     /note="pLannotate"
                     /database="snapgene"
                     /identity="100.0"
                     /match_length="100.0"
                     /fragment="False"
                     /other="CDS"
                     /label="rop"
     promoter        3633..3651
                     /note="pLannotate"
                     /database="snapgene"
                     /identity="100.0"
                     /match_length="100.0"
                     /fragment="False"
                     /other="promoter"
                     /label="T7 promoter"
     misc_feature    3633..6002
                     /created_by="MeierM"
                     /modified_by="MeierM"
                     /label="secYEG_operon"
     misc_feature    3653..3676
                     /ApEinfo_revcolor="#c6c9d1"
                     /ApEinfo_fwdcolor="#c6c9d1"
                     /label="5' UTR spacer"
     primer_bind     3653..3701
                     /%GC="26.5"
                     /Hairpin_Tm="27.2"
                     /Self_Dimer_Tm="4.9"
                     /Tm="58.6"
                     /annotation_group="1738837564600dca72e93-7842-487d-9764-6f
                     90e13d0285"
                     /Sequence="CGAGTAGAAATAATTTTGTTTAAGATTAAAGAGGAGAAATTACATAT
                     GG"
                     /Mismatches=0
                     /#_Local_Off-target_Sites=1
                     /Local_Off-target="SAS385-T7-space-YEG<br>"
                     /Local_Off-target_#1="<html>CGAGTAGAAATAATTTTGTTTAAGATTAAA
                     GAGGAGAAATTACATATGG</html>"
                     /Location_#1="<html>504 -&gt; 552
                     (SAS385-T7-space-YEG)</html>"
                     /label="p260_GCT7space_F"
     misc_feature    3677..3697
                     /ApEinfo_revcolor="#75c6a9"
                     /ApEinfo_fwdcolor="#75c6a9"
                     /label="B0030 RBS"
     exon            3698..5029
                     /ApEinfo_revcolor="#c7b0e3"
                     /ApEinfo_fwdcolor="#c7b0e3"
                     /Color="#aa00aa"
                     /modified_by="MeierM"
                     /label="secY"
     misc_feature    4193..4195
                     /created_by="MeierM"
                     /label="T166L"
     primer_bind     complement(4331..4352)
                     /%GC="63.6"
                     /Hairpin_Tm="41.3"
                     /Self_Dimer_Tm="None"
                     /Tm="65.9"
                     /annotation_group="17391108476004e1e691e-e73e-47a4-ac2c-86
                     75b16e352d"
                     /created_by="primer3"
                     /Sequence="CGAGGAAGTGCAGGTCGCCTTG"
                     /Mismatches=0
                     /#_Local_Off-target_Sites=1
                     /Local_Off-target="SAS385-T7-space-YEG<br>"
                     /Local_Off-target_#1="<html>CGAGGAAGTGCAGGTCGCCTTG</html>"
                     /Location_#1="<html>1,182 &lt;- 1,203
                     (SAS385-T7-space-YEG)</html>"
                     /label="p163_secY_seq_R"
     misc_feature    5030..5055
                     /ApEinfo_revcolor="#b7e6d7"
                     /ApEinfo_fwdcolor="#b7e6d7"
                     /modified_by="MeierM"
                     /label="synthetic RBS 1"
     exon            5056..5439
                     /ApEinfo_revcolor="#9eafd2"
                     /ApEinfo_fwdcolor="#9eafd2"
                     /Color="#0000ee"
                     /modified_by="MeierM"
                     /label="secE"
     primer_bind     5124..5145
                     /%GC="59.1"
                     /Hairpin_Tm="31.0"
                     /Self_Dimer_Tm="8.8"
                     /Tm="66.0"
                     /annotation_group="2218319241610075495a05-014c-4ef3-97a1-e
                     860d0793835"
                     /created_by="primer3"
                     /Sequence="GGCATTGCTCCTGGTGGCGATT"
                     /Mismatches=0
                     /#_Off-target_Sites=0
                     /Off-target_Template="SAS385-T7-space-YEG"
                     /label="p50_secEseq_F"
     primer_bind     5124..5145
                     /%GC="59.1"
                     /Hairpin_Tm="31.0"
                     /Self_Dimer_Tm="8.8"
                     /Tm="66.0"
                     /annotation_group="263998626463008bf6f89a-fda3-434b-9174-d
                     ee3f0d0aa36"
                     /created_by="primer3"
                     /Sequence="GGCATTGCTCCTGGTGGCGATT"
                     /Mismatches=0
                     /#_Local_Off-target_Sites=0
                     /Local_Off-target="SAS385-T7-space-YEG"
                     /label="p50_secEseq_F"
     primer_bind     complement(5221..5242)
                     /%GC="59.1"
                     /Hairpin_Tm="None"
                     /Self_Dimer_Tm="12.1"
                     /Tm="65.8"
                     /annotation_group="22183792486300447f2990-69d0-4fe0-916e-2
                     0d9c0427773"
                     /created_by="primer3"
                     /Sequence="TCGTTAACAGCGCGACACCACC"
                     /Mismatches=0
                     /#_Off-target_Sites=0
                     /Off-target_Template="SAS385-T7-space-YEG"
                     /label="p51_secEseq_R"
     primer_bind     complement(5221..5242)
                     /%GC="59.1"
                     /Hairpin_Tm="None"
                     /Self_Dimer_Tm="12.1"
                     /Tm="65.8"
                     /annotation_group="26400104724400fefd8207-7765-48a1-9c53-7
                     b92fadba1a4"
                     /created_by="primer3"
                     /Sequence="TCGTTAACAGCGCGACACCACC"
                     /Mismatches=0
                     /#_Local_Off-target_Sites=0
                     /Local_Off-target="SAS385-T7-space-YEG"
                     /label="p51_secEseq_R"
     misc_feature    5440..5465
                     /ApEinfo_revcolor="#b7e6d7"
                     /ApEinfo_fwdcolor="#b7e6d7"
                     /modified_by="MeierM"
                     /label="synthetic RBS 2"
     exon            5466..5798
                     /ApEinfo_revcolor="#84b0dc"
                     /ApEinfo_fwdcolor="#84b0dc"
                     /Color="#008811"
                     /modified_by="MeierM"
                     /label="secG"
     terminator      5799..5860
                     /ApEinfo_revcolor="#ffef86"
                     /ApEinfo_fwdcolor="#ffef86"
                     /note="Derived using Geneious Prime 2024.0.7 'Annotate
                     from Database' based on nucleotide similarity"
                     /Transferred_From="pTE5604-SAS358-T7-space-Y(P276R)EG"
                     /Transferred_Similarity="100.00%"
                     /Primary_Match="100.00%; Adjusted-Interval-0:0/0; L3S2P21
                     (<a
                     href=""urn:local:MeierM:1d-isvrgiv"">pTE5604-SAS358-T7-spa
                     ce-Y(P276R)EG</a>)"
                     /Alternative_Match="100.00%; Adjusted-Interval-0:0/0;
                     L3S2P21 (<a
                     href=""urn:local:MeierM:1e-isvrgiv"">pTE5607-SAS385-T7-spa
                     ce-YEG</a>)"
                     /Alternative_Match="100.00%; Adjusted-Interval-0:0/0;
                     L3S2P21 (<a
                     href=""urn:local:MeierM:1f-isvrgiv"">pTE5608-SAS435-secA_T
                     7space</a>)"
                     /Alternative_Match="100.00%; Adjusted-Interval-0:0/0;
                     L3S2P21 (<a
                     href=""urn:local:MeierM:p-isvre86"">pTE5602-pMM-020-T7-pro
                     OmpA-pep99</a>)"
                     /Alternative_Match="100.00%; Adjusted-Interval-0:0/0;
                     L3S2P21 (<a
                     href=""urn:local:MeierM:q-isvre87"">pTE5603-pMM-031-T7-PR(
                     -1tm)-pep104</a>)"
                     /Alternative_Match="100.00%; Adjusted-Interval-0:0/0;
                     L3S2P21 (<a
                     href=""urn:local:MeierM:s-isvre87"">pTE5605-pMM-090-T7-spa
                     ce-secY(P84L)EG_lib#22</a>)"
                     /label="L3S2P21"
     terminator      5874..6002
                     /ApEinfo_revcolor="#85dae9"
                     /ApEinfo_fwdcolor="#85dae9"
                     /note="Derived using Geneious Prime 2024.0.7 'Annotate
                     from Database' based on nucleotide similarity"
                     /Transferred_From="pTE5604-SAS358-T7-space-Y(P276R)EG"
                     /Transferred_Similarity="100.00%"
                     /Primary_Match="100.00%; Adjusted-Interval-0:0/0;
                     T7_terminator (<a
                     href=""urn:local:MeierM:1d-isvrgiv"">pTE5604-SAS358-T7-spa
                     ce-Y(P276R)EG</a>)"
                     /Alternative_Match="100.00%; Adjusted-Interval-0:0/0;
                     T7_terminator (<a
                     href=""urn:local:MeierM:1e-isvrgiv"">pTE5607-SAS385-T7-spa
                     ce-YEG</a>)"
                     /Alternative_Match="100.00%; Adjusted-Interval-0:0/0;
                     T7_terminator (<a
                     href=""urn:local:MeierM:1f-isvrgiv"">pTE5608-SAS435-secA_T
                     7space</a>)"
                     /Alternative_Match="100.00%; Adjusted-Interval-0:0/0;
                     T7_terminator (<a
                     href=""urn:local:MeierM:p-isvre86"">pTE5602-pMM-020-T7-pro
                     OmpA-pep99</a>)"
                     /Alternative_Match="100.00%; Adjusted-Interval-0:0/0;
                     T7_terminator (<a
                     href=""urn:local:MeierM:q-isvre87"">pTE5603-pMM-031-T7-PR(
                     -1tm)-pep104</a>)"
                     /Alternative_Match="100.00%; Adjusted-Interval-0:0/0;
                     T7_terminator (<a
                     href=""urn:local:MeierM:s-isvre87"">pTE5605-pMM-090-T7-spa
                     ce-secY(P84L)EG_lib#22</a>)"
                     /Alternative_Match="100.00%; Adjusted-Interval-0:56/-25;
                     T7 terminator (<a
                     href=""urn:local:MeierM:j-jwuyh4e"">251113ONT-SAS453-scott
                     </a>)"
                     /label="T7_terminator"
ORIGIN      
        1 acgcgccctg tagcggcgca ttaagcgcgg cgggtgtggt ggttacgcgc agcgtgaccg
       61 ctacacttgc cagcgcccta gcgcccgctc ctttcgcttt cttcccttcc tttctcgcca
      121 cgttcgccgg ctttccccgt caagctctaa atcgggggct ccctttaggg ttccgattta
      181 gtgctttacg gcacctcgac cccaaaaaac ttgattaggg tgatggttca cgtagtgggc
      241 catcgccctg atagacggtt tttcgccctt tgacgttgga gtccacgttc tttaatagtg
      301 gactcttgtt ccaaactgga acaacactca accctatctc ggtctattct tttgatttat
      361 aagggatttt gccgatttcg gcctattggt taaaaaatga gctgatttaa caaaaattta
      421 acgcgaattt taacaaaata ttaacgctta caatttaggt ggcacttttc ggggaaatgt
      481 gcgcggaacc cctatttgtt tatttttcta aatacattca aatatgtatc cgctcatgaa
      541 ttaattctta gaaaaactca tcgagcatca aatgaaactg caatttattc atatcaggat
      601 tatcaatacc atatttttga aaaagccgtt tctgtaatga aggagaaaac tcaccgaggc
      661 agttccatag gatggcaaga tcctggtatc ggtctgcgat tccgactcgt ccaacatcaa
      721 tacaacctat taatttcccc tcgtcaaaaa taaggttatc aagtgagaaa tcaccatgag
      781 tgacgactga atccggtgag aatggcaaaa gtttatgcat ttctttccag acttgttcaa
      841 caggccagcc attacgctcg tcatcaaaat cactcgcatc aaccaaaccg ttattcattc
      901 gtgattgcgc ctgagcgaga cgaaatacgc gatcgctgtt aaaaggacaa ttacaaacag
      961 gaatcgaatg caaccggcgc aggaacactg ccagcgcatc aacaatattt tcacctgaat
     1021 caggatattc ttctaatacc tggaatgctg ttttcccggg gatcgcagtg gtgagtaacc
     1081 atgcatcatc aggagtacgg ataaaatgct tgatggtcgg aagaggcata aattccgtca
     1141 gccagtttag tctgaccatc tcatctgtaa catcattggc aacgctacct ttgccatgtt
     1201 tcagaaacaa ctctggcgca tcgggcttcc catacaatcg atagattgtc gcacctgatt
     1261 gcccgacatt atcgcgagcc catttatacc catataaatc agcatccatg ttggaattta
     1321 atcgcggcct agagcaagac gtttcccgtt gaatatggct cataacaccc cttgtattac
     1381 tgtttatgta agcagacagt tttattgttc atgaccaaaa tcccttaacg tgagttttcg
     1441 ttccactgag cgtcagaccc cgtagaaaag atcaaaggat cttcttgaga tccttttttt
     1501 ctgcgcgtaa tctgctgctt gcaaacaaaa aaaccaccgc taccagcggt ggtttgtttg
     1561 ccggatcaag agctaccaac tctttttccg aaggtaactg gcttcagcag agcgcagata
     1621 ccaaatactg tccttctagt gtagccgtag ttaggccacc acttcaagaa ctctgtagca
     1681 ccgcctacat acctcgctct gctaatcctg ttaccagtgg ctgctgccag tggcgataag
     1741 tcgtgtctta ccgggttgga ctcaagacga tagttaccgg ataaggcgca gcggtcgggc
     1801 tgaacggggg gttcgtgcac acagcccagc ttggagcgaa cgacctacac cgaactgaga
     1861 tacctacagc gtgagctatg agaaagcgcc acgcttcccg aagggagaaa ggcggacagg
     1921 tatccggtaa gcggcagggt cggaacagga gagcgcacga gggagcttcc agggggaaac
     1981 gcctggtatc tttatagtcc tgtcgggttt cgccacctct gacttgagcg tcgatttttg
     2041 tgatgctcgt caggggggcg gagcctatgg aaaaacgcca gcaacgcggc ctttttacgg
     2101 ttcctggcct tttgctggcc ttttgctcac atgttctttc ctgcgttatc ccctgattct
     2161 gtggataacc gtattaccgc ctttgagtga gctgataccg ctcgccgcag ccgaacgacc
     2221 gagcgcagcg agtcagtgag cgaggaagcg gaagagcgcc tgatgcggta ttttctcctt
     2281 acgcatctgt gcggtatttc acaccgcaat ggtgcactct cagtacaatc tgctctgatg
     2341 ccgcatagtt aagccagtat acactccgct atcgctacgt gactgggtca tggctgcgcc
     2401 ccgacacccg ccaacacccg ctgacgcgcc ctgacgggct tgtctgctcc cggcatccgc
     2461 ttacagacaa gctgtgaccg tctccgggag ctgcatgtgt cagaggtttt caccgtcatc
     2521 accgaaacgc gcgaggcagc tgcggtaaag ctcatcagcg tggtcgtgaa gcgattcaca
     2581 gatgtctgcc tgttcatccg cgtccagctc gttgagtttc tccagaagcg ttaatgtctg
     2641 gcttctgata aagcgggcca tgttaagggc ggttttttcc tgtttggtca ctaatgcctc
     2701 cgtgtaaggg ggatttctgt tcatgggggt aatgataccg atgaaacgag agaggatgct
     2761 cacgatacgg gttactgatg atgaacatgc ccggttactg gaacgttgtg agggtaaaca
     2821 actggcggta tggatgcggc gggaccagag aaaaatcact cagggtcaat gccagcgctt
     2881 cgttaataca gatgtaggtg ttccacaggg tagccagcag catcctgcga tgcagatccg
     2941 gaacataatg gtgcagggcg ctgacttccg cgtttccaga ctttacgaaa cacggaaacc
     3001 gaagaccatt catgttgttg ctcaggtcgc agacgttttg cagcagcagt cgcttcacgt
     3061 tcgctcgcgt atcggtgatt cattctgcta accagtaagg caaccccgcc agcctagccg
     3121 ggtcctcaac gacaggagca cgatcatgcg cacccgtggg gccgccatgc cggcgataat
     3181 ggcctgcttc tcgccgaaac gtttggtggc gggaccagtg acgaaggctt gagcgagggc
     3241 gtgcaagatt ccgaataccg caagcgacag gccgatcatc gtcgcgctcc agcgaaagcg
     3301 gtcctcgccg aaaatgaccc agagcgctgc cggcacctgt cctacgagtt gcatgataaa
     3361 gaagacagtc ataagtgcgg cgacgatagt catgccccgc gcccaccgga aggagctgac
     3421 tgggttggtg catgcaagga gatggcgccc aacagtcccc cggccacggg gcctgccacc
     3481 atacccacgc cgaaacaagc gctcatgagc ccgaagtggc gagcccgatc ttccccatcg
     3541 gtgatgtcgg cgatataggc gccagcaacc gcacctgtgg cgccggtgat gccggccacg
     3601 atgcgtccgg cgtagaggat cgagatctcg ggtaatacga ctcactatag ggcgagtaga
     3661 aataattttg tttaagatta aagaggagaa attacatatg gctaaacaac cgggattaga
     3721 ttttcaaagt gccaaaggtg gcttaggcga gctgaaacgc agactgctgt ttgttatcgg
     3781 tgcgctgatt gtgttccgta ttggctcttt tattccgatc cctggtattg atgccgctgt
     3841 acttgccaaa ctgcttgagc aacagcgagg caccatcatt gagatgttta acatgttctc
     3901 tggtggtgct ctcagccgtg cttctatctt tgctctgggg atcatgccgt atatttcggc
     3961 gtcgatcatt atccagctgc tgacggtggt tcacccaacg ttggcagaaa ttaagaaaga
     4021 aggggagtct ggtcgtcgta agatcagcca gtacacccgc tacggtactc tggtgctggc
     4081 aatattccag tcgatcggta ttgctaccgg tctgccgaat atgcctggta tgcaaggcct
     4141 ggtgattaac ccgggctttg cattctactt caccgctgtt gtaagtctgg tcctgggaac
     4201 catgttcctg atgtggttgg gcgaacagat tactgaacga ggtatcggca acggtatttc
     4261 aatcattatc ttcgccggta ttgtcgcggg actcccgcca gccattgccc atactatcga
     4321 gcaagcgcgt caaggcgacc tgcacttcct cgtgttgctg ttggttgcag tattagtatt
     4381 tgcagtgacg ttctttgttg tatttgttga gcgtggtcaa cgccgcattg tggtaaacta
     4441 cgcgaaacgt cagcaaggtc gtcgtgtcta tgctgcacag agcacacatt taccgctgaa
     4501 agtgaatatg gcgggggtaa tcccggcaat cttcgcttcc agtattattc tgttcccggc
     4561 gaccatcgcg tcatggttcg ggggcggtac tggttggaac tggctgacaa caatttcgct
     4621 gtatttgcag cctgggcaac cgctttatgt gttactctat gcgtctgcaa tcatcttctt
     4681 ctgtttcttc tacacggcgt tggttttcaa cccgcgtgaa acagcagata acctgaagaa
     4741 gtccggtgca tttgtaccag gaattcgtcc gggagagcaa acggcgaagt atatcgataa
     4801 agtaatgacc cgcctgaccc tggttggtgc gctgtatatt acctttatct gcctgatccc
     4861 ggagttcatg cgtgatgcaa tgaaagtacc gttctacttc ggtgggacct cactgcttat
     4921 cgttgttgtc gtgattatgg actttatggc tcaagtgcaa actctgatga tgtccagtca
     4981 gtatgagtct gcattgaaga aggcgaacct gaaaggctac ggccgataat agtttaatct
     5041 ataaggggta tatatatgag tgcgaatacc gaagctcaag gaagcgggcg cggcctggaa
     5101 gcgatgaagt gggtcgttgt ggtggcattg ctcctggtgg cgattgtcgg caactatctt
     5161 tatcgcgaca ttatgctgcc gctgcgtgcg ctggccgtag taattctgat tgctgcagcg
     5221 ggtggtgtcg cgctgttaac gacaaaaggt aaagctaccg ttgcttttgc ccgtgaagcg
     5281 cgtaccgaag tccgtaaggt catttggccg actcgccagg aaacattgca caccacgctg
     5341 attgtggctg cggttaccgc agtaatgtca ctgatcctgt ggggactgga tggtattctg
     5401 gttcgcctgg tatcctttat cactggcctg aggttctgag ttagaaataa ttaaggctgg
     5461 ttgttatgta tgaagctctt ttagtagttt tccttattgt ggcaattggc cttgttggtc
     5521 tgatcatgct gcagcaaggt aaaggcgctg atatgggagc ctccttcgga gcaggcgctt
     5581 ccgctacgct gtttggttca agtggttctg gtaacttcat gacccgcatg acggcgctgc
     5641 tggcaacgtt attcttcatc atcagtctgg tgctgggtaa catcaatagc aacaaaacca
     5701 ataaaggtag cgaatgggaa aatctgagtg caccggcgaa aaccgaacaa actcagccag
     5761 ctgctccggc taagccgacc agcgatatcc cgaactaacc tcggtaccaa attccagaaa
     5821 agaggcctcc cgaaaggggg gccttttttc gttttggtcc cactgagatc cggctgctaa
     5881 caaagcccga aaggaagctg agttggctgc tgccaccgct gagcaataac tagcataacc
     5941 ccttggggcc tctaaacggg tcttgagggg ttttttgctg aaaggaggaa ctatatccgg
     6001 attggcgaat ggg
//
