LOCUS       pSBK.136                4574 bp ds-DNA     circular SYN 04-JUN-2021
DEFINITION  Sal->A + aTc->B_ncRNA_ribozyme-free. pSol backbone.
ACCESSION
VERSION     .
KEYWORDS    pSOL His tsf.
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct.
REFERENCE   1  (bases 1 to 3145)
  AUTHORS   Ron
  TITLE     Direct Submission
  JOURNAL   Exported Monday, Oct 12, 2015 from SnapGene 2.8.2
            http://www.snapgene.com
FEATURES             Location/Qualifiers
     misc_feature    1..66
                     /Original_Bases
                     /label
                     /note="Geneious type: Editing History Insertion"
     primer_bind     1..17
                     /Mismatches=0
                     /%GC="70.6"
                     /Hairpin_Tm="36.3"
                     /Self_Dimer_Tm="None"
                     /Tm="62.5"
                     /annotation_group="6059265472472008fa37be2-6973-4c7d-9ee9-
                     ccd7780c41a1"
                     /Sequence="CGCGTCCGAACCCAGCT"
                     /created_by="primer3"
                     /label="SBK_23"
     terminator      21..66
                     /created_by="santi.bhattaraikline"
                     /biology="LuxIA"
                     /iGEM_Part="BBa_B0011"
                     /direction="Bidirectional"
                     /modified_by="santi.bhattaraikline"
                     /label="LuxIA Terminator"
     primer_bind     29..86
                     /modified_by="santi.bhattaraikline"
                     /Mismatches=0
                     /%GC="34.5"
                     /Hairpin_Tm="57.9"
                     /Self_Dimer_Tm="40.9"
                     /Tm="66.0"
                     /annotation_group="11535106886812008ba7e358-5411-4221-80c9
                     -03081d055312"
                     /Sequence="TAAAAAGCCAGATTATTAATCCGGCTTTTTTATTATTTCGCTTAACG
                     ATCGTTGGCTG"
                     /created_by="primer3"
                     /label="dualProm_bb_fow"
     primer_bind     29..86
                     /modified_by="santi.bhattaraikline"
                     /Mismatches=0
                     /%GC="34.5"
                     /Hairpin_Tm="57.9"
                     /Self_Dimer_Tm="40.9"
                     /Tm="66.0"
                     /annotation_group="284610742080300a1b2de3e-de52-49ac-879d-
                     f12736df630a"
                     /Sequence="TAAAAAGCCAGATTATTAATCCGGCTTTTTTATTATTTCGCTTAACG
                     ATCGTTGGCTG"
                     /created_by="primer3"
                     /label="dualProm_bb_fow"
     primer_bind     complement(65..86)
                     /Mismatches=0
                     /%GC="50.0"
                     /Hairpin_Tm="53.5"
                     /Self_Dimer_Tm="32.3"
                     /Tm="61.8"
                     /annotation_group="6059267433431008ef9acf5-f56b-4f37-a88d-
                     fe73a5d11ced"
                     /Sequence="CAGCCAACGATCGTTAAGCGAA"
                     /created_by="primer3"
                     /label="SBK_24"
     source          join(205..>1729,<67..204)
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     terminator      67
                     /label="L3S3P21"
     misc_feature    68..1727
                     /Original_Bases="ACCACAATTCAGCAAATTGTGAACATCATCACGTTCATCTT
                     TCCCTGGTTGCCAATGGCCCATTTTCCTGTCAGTAACGAGAAGGTCGCGAATTCAGGC
                     GCTTTTTAGACTGGTCGTAGGGAGACCACAACGGTTTCCCTCTAGAAATAATTTTGTT
                     TAACTATAAGAAGGAGATATACATATGCATCATCACCACCATCACATGGCAGAAATTA
                     CGGCTTCCCTGGTGAAAGAATTGCGCGAACGTACGGGTGCAGGCATGATGGACTGCAA
                     GAAGGCGCTGACCGAAGCCAATGGCGACATCGAGCTGGCGATCGAGAACATGCGTAAA
                     TCGGGTGCAATCAAAGCGGCCAAAAAGGCGGGTAATGTTGCGGCAGACGGCGTCATTA
                     AAACCAAGATCGATGGTAACTATGGCATTATTCTGGAAGTCAACTGTCAGACGGATTT
                     CGTGGCGAAGGACGCGGGTTTTCAAGCCTTCGCCGACAAGGTCCTGGACGCAGCGGTG
                     GCTGGTAAAATCACGGATGTCGAAGTGTTGAAAGCACAGTTTGAAGAAGAGCGTGTCG
                     CTCTGGTGGCAAAGATCGGCGAGAATATCAATATCCGCCGTGTTGCGGCACTGGAAGG
                     TGATGTTCTGGGCAGCTACCAGCACGGTGCGCGTATTGGCGTTCTGGTGGCTGCCAAG
                     GGTGCGGACGAAGAGCTGGTGAAACATATCGCGATGCATGTCGCCGCGAGCAAACCAG
                     AGTTTATCAAACCGGAAGATGTTAGCGCCGAGGTCGTGGAAAAAGAGTATCAGGTACA
                     GCTGGACATTGCGATGCAGTCCGGCAAACCTAAAGAGATCGCAGAGAAGATGGTCGAG
                     GGCCGCATGAAGAAGTTTACCGGTGAGGTTAGCCTGACTGGTCAACCGTTTGTTATGG
                     AGCCGAGCAAAACGGTTGGTCAATTGCTGAAAGAGCACAATGCGGAAGTGACCGGTTT
                     CATTCGTTTTGAGGTGGGTGAAGGCATTGAGAAAGTGGAAACCGATTTCGCGGCAGAA
                     GTTGCGGCTATGAGCAAGCAATCTGAGAATCTGTACTTCCAGGGTTAATAGAGCGGCC
                     GCCACCGCTGAGCAATAACTAGCATAACCCCTTGGGGCCTCTAAACGGGTCTTGAGGG
                     GTTTTT"
                     /label="ACCACAATTCAGCAAATTGTGAACATCATCACGTTCATCTTTCCCTGGTT
                     GCCAATGGCCCATTTTCCTGTCAGTAACGAGAAGGTCGCGAATTCAGGCGCTTTTTAG
                     ACTGGTCGTAGGGAGACCACAACGGTTTCCCTCTAGAAATAATTTTGTTTAACTATAA
                     GAAGGAGATATACATATGCATCATCACCACCATCACATGGCAGAAATTACGGCTTCCC
                     TGGTGAAAGAATTGCGCGAACGTACGGGTGCAGGCATGATGGACTGCAAGAAGGCGCT
                     GACCGAAGCCAATGGCGACATCGAGCTGGCGATCGAGAACATGCGTAAATCGGGTGCA
                     ATCAAAGCGGCCAAAAAGGCGGGTAATGTTGCGGCAGACGGCGTCATTAAAACCAAGA
                     TCGATGGTAACTATGGCATTATTCTGGAAGTCAACTGTCAGACGGATTTCGTGGCGAA
                     GGACGCGGGTTTTCAAGCCTTCGCCGACAAGGTCCTGGACGCAGCGGTGGCTGGTAAA
                     ATCACGGATGTCGAAGTGTTGAAAGCACAGTTTGAAGAAGAGCGTGTCGCTCTGGTGG
                     CAAAGATCGGCGAGAATATCAATATCCGCCGTGTTGCGGCACTGGAAGGTGATGTTCT
                     GGGCAGCTACCAGCACGGTGCGCGTATTGGCGTTCTGGTGGCTGCCAAGGGTGCGGAC
                     GAAGAGCTGGTGAAACATATCGCGATGCATGTCGCCGCGAGCAAACCAGAGTTTATCA
                     AACCGGAAGATGTTAGCGCCGAGGTCGTGGAAAAAGAGTATCAGGTACAGCTGGACAT
                     TGCGATGCAGTCCGGCAAACCTAAAGAGATCGCAGAGAAGATGGTCGAGGGCCGCATG
                     AAGAAGTTTACCGGTGAGGTTAGCCTGACTGGTCAACCGTTTGTTATGGAGCCGAGCA
                     AAACGGTTGGTCAATTGCTGAAAGAGCACAATGCGGAAGTGACCGGTTTCATTCGTTT
                     TGAGGTGGGTGAAGGCATTGAGAAAGTGGAAACCGATTTCGCGGCAGAAGTTGCGGCT
                     ATGAGCAAGCAATCTGAGAATCTGTACTTCCAGGGTTAATAGAGCGGCCGCCACCGCT
                     GAGCAATAACTAGCATAACCCCTTGGGGCCTCTAAACGGGTCTTGAGGGGTTTTT"
                     /note="Geneious type: Editing History Replacement"
     misc_feature    74..79
                     /Recognition_pattern="CGAT^CG"
                     /Methylation="Dam:cut, CpG:blocked"
                     /annotation_group="CGATCG"
                     /label="PvuI"
                     /note="Geneious type: restriction site"
     promoter        87..202
                     /label="pSalTTC"
     operator        124..154
                     /label="NahO"
     promoter        168..173
                     /label="-35"
     promoter        190..195
                     /label="-10"
     misc_RNA        205..>225
                     /note="hammerhead ribozyme from the tobacco ringspot virus
                      satellite RNA (Khvorova et al., 2003)"
                     /label="sTRSV HHRz"
     primer_bind     complement(205..232)
                     /Mismatches=0
                     /%GC="50.0"
                     /Hairpin_Tm="51.3"
                     /Self_Dimer_Tm="None"
                     /Tm="65.5"
                     /annotation_group="1962266928034004e594139-f923-413b-82fb-
                     8d86bdf52cc0"
                     /Sequence="CTTATCAGGAAAGCACATCCGGTGACAG"
                     /created_by="primer3"
                     /label="pSBK_106_4kld_rev"
     primer_bind     complement(205..232)
                     /Mismatches=0
                     /%GC="50.0"
                     /Hairpin_Tm="51.3"
                     /Self_Dimer_Tm="None"
                     /Tm="65.5"
                     /annotation_group="2846136483079005f63de8b-635a-4941-b083-
                     60c83af4b328"
                     /Sequence="CTTATCAGGAAAGCACATCCGGTGACAG"
                     /created_by="primer3"
                     /label="106_4kld_rev"
     misc_signal     <226..239
                     /created_by="User"
                     /Transferred_From="template"
                     /Transferred_Similarity="100.00%"
                     /label="T7 to msr"
     primer_bind     233..259
                     /Mismatches=0
                     /%GC="55.6"
                     /Hairpin_Tm="57.4"
                     /Self_Dimer_Tm="14.9"
                     /Tm="68.4"
                     /annotation_group="1962264997445003409ea02-ebca-4b8e-ba7d-
                     948603512ddb"
                     /Sequence="ATTCCGTATGCGCACCCTTAGCGAGAG"
                     /created_by="primer3"
                     /label="pSBK_106_4kld_fow"
     primer_bind     233..259
                     /Mismatches=0
                     /%GC="55.6"
                     /Hairpin_Tm="57.4"
                     /Self_Dimer_Tm="14.9"
                     /Tm="68.4"
                     /annotation_group="28461518588110047ab6449-571b-4b0a-b35c-
                     e3bcd4cf3f29"
                     /Sequence="ATTCCGTATGCGCACCCTTAGCGAGAG"
                     /created_by="primer3"
                     /label="106_4kld_fow"
     misc_rna        240..321
                     /created_by="User"
                     /Transferred_From="template"
                     /Transferred_Similarity="100.00%"
                     /label="msdRNA"
     misc_feature    241..252
                     /created_by="User"
                     /Transferred_From="template"
                     /Transferred_Similarity="100.00%"
                     /label="'primer'"
     primer_bind     268..293
                     /Mismatches=0
                     /%GC="38.5"
                     /Hairpin_Tm="48.0"
                     /Self_Dimer_Tm="14.8"
                     /Tm="60.9"
                     /annotation_group="165387026849300c49d1fc8-efe8-4c9d-94a1-
                     a5f631540b61"
                     /Sequence="TTAAGGTCAACCTCTGGATGTTGTTT"
                     /created_by="primer3"
                     /label="Uloop_detect_fow_wt"
     primer_bind     268..293
                     /Mismatches=0
                     /%GC="38.5"
                     /Hairpin_Tm="48.0"
                     /Self_Dimer_Tm="14.8"
                     /Tm="60.9"
                     /annotation_group="104686168500800fb95c72f-ed1f-4602-98bf-
                     04fce4c05e39"
                     /Sequence="TTAAGGTCAACCTCTGGATGTTGTTT"
                     /created_by="primer3"
                     /label="Uloop_detect_fow_wt"
     primer_bind     268..293
                     /Mismatches=0
                     /%GC="38.5"
                     /Hairpin_Tm="48.0"
                     /Self_Dimer_Tm="14.8"
                     /Tm="60.9"
                     /annotation_group="2846097175357005e88e825-8093-496f-80c3-
                     8da8d8547b7d"
                     /Sequence="TTAAGGTCAACCTCTGGATGTTGTTT"
                     /created_by="primer3"
                     /label="Uloop_detect_fow_wt"
     motif           283..290
                     /created_by="santi.bhattaraikline"
                     /label="Stem (RT Recognition)"
     motif           291..293
                     /created_by="santi.bhattaraikline"
                     /label="UUU Loop (RT Recognition)"
     primer          complement(294..>310)
                     /created_by="User"
                     /label="86_rev"
     motif           complement(294..301)
                     /created_by="santi.bhattaraikline"
                     /label="Stem (RT Recognition)"
     primer          complement(300..>310)
                     /modified_by="User"
                     /label="msd_Rev"
     misc_feature    complement(311..367)
                     /created_by="User"
                     /modified_by="Seth"
                     /label="msd"
     motif           311..327
                     /created_by="santi.bhattaraikline"
                     /label="Flanking"
     primer_bind     complement(323..345)
                     /modified_by="santi.bhattaraikline"
                     /Mismatches=0
                     /%GC="47.8"
                     /Hairpin_Tm="38.3"
                     /Self_Dimer_Tm="30.1"
                     /Tm="60.9"
                     /annotation_group="165388223594900d66123d0-4b94-4c29-a6b8-
                     c4b17b8da6f1"
                     /Sequence="GTTGCTGCAACCTCTCCATTTTC"
                     /created_by="primer3"
                     /label="SPCR_Ec86_rev"
     primer_bind     complement(323..345)
                     /modified_by="santi.bhattaraikline"
                     /Mismatches=0
                     /%GC="47.8"
                     /Hairpin_Tm="38.3"
                     /Self_Dimer_Tm="30.1"
                     /Tm="60.9"
                     /annotation_group="1046850431475008f846192-d87d-45c7-b0aa-
                     9e4360ff4811"
                     /Sequence="GTTGCTGCAACCTCTCCATTTTC"
                     /created_by="primer3"
                     /label="SPCR_Ec86_rev"
     primer_bind     complement(323..345)
                     /modified_by="santi.bhattaraikline"
                     /Mismatches=0
                     /%GC="47.8"
                     /Hairpin_Tm="38.3"
                     /Self_Dimer_Tm="30.1"
                     /Tm="60.9"
                     /annotation_group="2846113077188008e757e54-4017-4dcf-af15-
                     f15694a22c66"
                     /Sequence="GTTGCTGCAACCTCTCCATTTTC"
                     /created_by="primer3"
                     /label="SPCR_Ec86_rev"
     motif           328..338
                     /created_by="santi.bhattaraikline"
                     /label="Stem"
     motif           339..341
                     /created_by="santi.bhattaraikline"
                     /label="Loop"
     motif           complement(342..352)
                     /created_by="santi.bhattaraikline"
                     /label="Stem"
     motif           complement(353..367)
                     /created_by="santi.bhattaraikline"
                     /label="Flanking"
     primer_bind     complement(354..376)
                     /Mismatches=0
                     /Mismatches=0
                     /%GC="47.8"
                     /Hairpin_Tm="43.0"
                     /Self_Dimer_Tm="None"
                     /Tm="60.3"
                     /annotation_group="165387220752800f87128de-d812-4826-a0c5-
                     79ec029f3fb2"
                     /Sequence="CACCCTTACGTCAGAAAAAACGG"
                     /created_by="primer3"
                     /label="020_4kld_rev"
     primer_bind     complement(354..376)
                     /Mismatches=0
                     /Mismatches=0
                     /%GC="47.8"
                     /Hairpin_Tm="43.0"
                     /Self_Dimer_Tm="None"
                     /Tm="60.3"
                     /annotation_group="104685231276500ad3fdbe4-f078-4bd4-908d-
                     16d39be9d11a"
                     /Sequence="CACCCTTACGTCAGAAAAAACGG"
                     /created_by="primer3"
                     /label="020_4kld_rev"
     primer_bind     complement(354..376)
                     /Mismatches=0
                     /Mismatches=0
                     /%GC="47.8"
                     /Hairpin_Tm="43.0"
                     /Self_Dimer_Tm="None"
                     /Tm="60.3"
                     /annotation_group="284609340240500df32a754-190d-4159-840d-
                     bea48f78c9e9"
                     /Sequence="CACCCTTACGTCAGAAAAAACGG"
                     /created_by="primer3"
                     /label="020_4kld_rev"
     primer          <368..378
                     /created_by="User"
                     /label="86_fow"
     primer          <368..376
                     /modified_by="User"
                     /label="msd_Fow"
     misc_feature    369..395
                     /created_by="User"
                     /label="'primer'"
     primer_bind     complement(373..405)
                     /Mismatches=0
                     /%GC="51.5"
                     /Hairpin_Tm="36.7"
                     /Self_Dimer_Tm="14.9"
                     /Tm="70.6"
                     /annotation_group="104685606991900fc49279d-c126-499f-90d8-
                     6c65003d6157"
                     /Sequence="TGGTACCGAGTGATAAGATTCCGTATGCGCACC"
                     /created_by="primer3"
                     /label="057_ins_rev"
     primer_bind     complement(373..405)
                     /Mismatches=0
                     /%GC="51.5"
                     /Hairpin_Tm="36.7"
                     /Self_Dimer_Tm="14.9"
                     /Tm="70.6"
                     /annotation_group="284612493866100dd758574-13fd-4a37-b7b2-
                     cb4371a16a4b"
                     /Sequence="TGGTACCGAGTGATAAGATTCCGTATGCGCACC"
                     /created_by="primer3"
                     /label="057_ins_rev"
     primer_bind     386..419
                     /Mismatches=0
                     /%GC="35.3"
                     /Hairpin_Tm="None"
                     /Self_Dimer_Tm="13.5"
                     /Tm="64.6"
                     /annotation_group="10468466980390040ae308b-cd36-4932-872a-
                     b4fa5f7d843e"
                     /Sequence="AATCTTATCACTCGGTACCAAATTCCAGAAAAGA"
                     /created_by="primer3"
                     /label="057_bb_fow"
     primer_bind     386..419
                     /Mismatches=0
                     /%GC="35.3"
                     /Hairpin_Tm="None"
                     /Self_Dimer_Tm="13.5"
                     /Tm="64.6"
                     /annotation_group="284612680436800d7838df2-0a92-4a10-8bd8-
                     eb4bc2bd27a6"
                     /Sequence="AATCTTATCACTCGGTACCAAATTCCAGAAAAGA"
                     /created_by="primer3"
                     /label="057_bb_fow"
     terminator      396..455
                     /label="L3S2P21"
     promoter        460..513
                     /label="pLacIQ"
     promoter        478..483
                     /label="-35"
     promoter        501..506
                     /label="-10"
     RBS             514..542
                     /label="nah2"
     CDS             543..1445
                     /label="NahR-AM"
     misc_feature    597..602
                     /Recognition_pattern="G^TCGAC"
                     /Methylation="CpG:some blocked"
                     /annotation_group="GTCGAC"
                     /label="SalI"
                     /note="Geneious type: restriction site"
     misc_feature    597..602
                     /Recognition_pattern="GT^MKAC"
                     /Methylation="CpG:blocked"
                     /annotation_group="GTMKAC"
                     /label="AccI"
                     /note="Geneious type: restriction site"
     mutation        615..617
                     /label="I25V"
     primer_bind     complement(684..703)
                     /Mismatches=0
                     /%GC="55.0"
                     /Hairpin_Tm="None"
                     /Self_Dimer_Tm="None"
                     /Tm="58.5"
                     /annotation_group="285605932866700dd8b508c-f603-4c59-b732-
                     7c923876124f"
                     /Sequence="ACGAAGAGTGGGTCCTGTAG"
                     /created_by="primer3"
                     /label="SBK_22_NahR-AM"
     mutation        756..758
                     /label="P72H"
     misc_feature    865..870
                     /Recognition_pattern="A^GATCT"
                     /Methylation="Dam:cut, EcoBI:some blocked"
                     /annotation_group="AGATCT"
                     /label="BglII"
                     /note="Geneious type: restriction site"
     mutation        894..896
                     /label="V118A"
     mutation        1284..1286
                     /label="R425C"
     misc_feature    1373..1378
                     /Recognition_pattern="GCATG^C"
                     /Methylation="CpG:cut, EcoBI:blocked"
                     /annotation_group="GCATGC"
                     /label="SphI"
                     /note="Geneious type: restriction site"
     misc_feature    1449..1454
                     /Recognition_pattern="G^GATCC"
                     /Methylation="Dam:cut, Dcm:cut, CpG:cut"
                     /annotation_group="GGATCC"
                     /label="BamHI"
                     /note="Geneious type: restriction site"
     terminator      1455..1709
                     /label="induction operon terminator region"
     primer_bind     complement(1460..1483)
                     /%GC="45.8"
                     /Mismatches=0
                     /Hairpin_Tm="None"
                     /Self_Dimer_Tm="5.1"
                     /Tm="60.2"
                     /annotation_group="284608007514700679ec834-7374-4dd1-9fd5-
                     eae3aac8104c;
                     284614432464800c70d9e0b-de34-4884-af1b-50e7840ca4da"
                     /Sequence="CCGTCAATTGTCTGATTCGTTACC"
                     /created_by="primer3"
                     /label="SBK_014_mar_term; mar_term_SBK_014"
     misc_feature    1484..1489
                     /Recognition_pattern="C^TCGAG"
                     /Methylation="CpG:impaired"
                     /annotation_group="CTCGAG"
                     /label="XhoI"
                     /note="Geneious type: restriction site"
     misc_feature    1484..1489
                     /Recognition_pattern="C^YCGRG"
                     /Methylation="CpG:blocked"
                     /annotation_group="CYCGRG"
                     /label="AvaI"
                     /note="Geneious type: restriction site"
     primer_bind     complement(1636..1655)
                     /Mismatches=0
                     /%GC="55.0"
                     /Hairpin_Tm="None"
                     /Self_Dimer_Tm="19.2"
                     /Tm="58.4"
                     /annotation_group="1046869165674003220b6ff-db9d-46d2-b48f-
                     d04aef6df6d0"
                     /Sequence="GTGATGTCGGCGATATAGGC"
                     /created_by="primer3"
                     /label="SLS_138"
     primer_bind     complement(1636..1655)
                     /Mismatches=0
                     /%GC="55.0"
                     /Hairpin_Tm="None"
                     /Self_Dimer_Tm="19.2"
                     /Tm="58.4"
                     /annotation_group="28461325352980001c69ab8-0e1b-4075-b75d-
                     d4ba062c3581"
                     /Sequence="GTGATGTCGGCGATATAGGC"
                     /created_by="primer3"
                     /label="SLS_138"
     primer_bind     complement(1710..1739)
                     /Mismatches=0
                     /%GC="56.7"
                     /Hairpin_Tm="51.5"
                     /Self_Dimer_Tm="2.1"
                     /Tm="71.2"
                     /annotation_group="1064202264959000308e07c-709a-4831-883d-
                     593435cf2047"
                     /Sequence="CTCCTTTCAGCAGTGAGCGAGGAAGCACCT"
                     /created_by="primer3"
                     /label="081_ins_rev"
     primer_bind     complement(1710..1739)
                     /Mismatches=0
                     /%GC="56.7"
                     /Hairpin_Tm="51.5"
                     /Self_Dimer_Tm="2.1"
                     /Tm="71.2"
                     /annotation_group="284608387202500f8434896-7ff5-4feb-aa8b-
                     0692eb0b8e14"
                     /Sequence="CTCCTTTCAGCAGTGAGCGAGGAAGCACCT"
                     /created_by="primer3"
                     /label="081_ins_rev"
     primer_bind     complement(1710..1729)
                     /Mismatches=0
                     /%GC="60.0"
                     /Hairpin_Tm="53.9"
                     /Self_Dimer_Tm="2.1"
                     /Tm="62.2"
                     /annotation_group="10468598117210042249baf-42e8-40e3-ab35-
                     12dd0cd76fd3"
                     /Sequence="CAGTGAGCGAGGAAGCACCT"
                     /created_by="primer3"
                     /label="mari_cassette_rev"
     primer_bind     complement(1710..1729)
                     /Mismatches=0
                     /%GC="60.0"
                     /Hairpin_Tm="53.9"
                     /Self_Dimer_Tm="2.1"
                     /Tm="62.2"
                     /annotation_group="284608575181500b2ea7e6c-0818-44f2-9827-
                     8f7f9a60dadc"
                     /Sequence="CAGTGAGCGAGGAAGCACCT"
                     /created_by="primer3"
                     /label="mari_cassette_rev"
     primer_bind     1720..1752
                     /Mismatches=0
                     /%GC="51.5"
                     /Hairpin_Tm="58.9"
                     /Self_Dimer_Tm="None"
                     /Tm="70.1"
                     /annotation_group="106419012123700b8fa9736-7c9e-4e97-a06d-
                     b5f2d635b845"
                     /Sequence="TCGCTCACTGCTGAAAGGAGGAACTATATCCGG"
                     /created_by="primer3"
                     /label="081_bb_fow"
     primer_bind     1720..1752
                     /Mismatches=0
                     /%GC="51.5"
                     /Hairpin_Tm="58.9"
                     /Self_Dimer_Tm="None"
                     /Tm="70.1"
                     /annotation_group="2846140569546004226636c-c85d-4c6b-93d4-
                     867caa2814be"
                     /Sequence="TCGCTCACTGCTGAAAGGAGGAACTATATCCGG"
                     /created_by="primer3"
                     /label="081_bb_fow"
     primer_bind     1730..1752
                     /Sequence="CTGAAAGGAGGAACTATATCCGG"
                     /Hairpin_Tm="44.6"
                     /Tm="57.9"
                     /Self_Dimer_Tm="None"
                     /%GC="47.8"
                     /created_by="primer3"
                     /label="pSol_bb_fow"
     misc_feature    1758..1763
                     /Recognition_pattern="G^AATTC"
                     /Methylation="CpG:variable, EcoBI:variable"
                     /annotation_group="GAATTC"
                     /label="EcoRI"
                     /note="Geneious type: restriction site"
     misc_feature    1770..1775
                     /Recognition_pattern="GAT^ATC"
                     /Methylation="CpG:some impaired, EcoBI:some blocked"
                     /annotation_group="GATATC"
                     /label="EcoRV"
                     /note="Geneious type: restriction site"
     terminator      1827..1856
                     /note="T3Te terminator"
                     /note="phage T3 early transcription terminator"
     primer_bind     complement(1827..1844)
                     /%GC="66.7"
                     /Mismatches=0
                     /Hairpin_Tm="57.6"
                     /Self_Dimer_Tm="4.7"
                     /Tm="60.4"
                     /annotation_group="2846142457628009a03c3df-ad9d-4980-81b2-
                     50e12dd66ce8;
                     284614996806700331d4031-e6f8-4f42-9ffe-f3e868d42832"
                     /Sequence="CACCCGTGAAGGTGAGCC"
                     /created_by="primer3"
                     /label="SBK_012_T3Te_term; T3Te_term_SBK_012"
     rep_origin      1878..2465
                     /direction="RIGHT"
                     /note="ori"
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     misc_feature    2207..2212
                     /Recognition_pattern="G^TGCAC"
                     /Methylation="CpG:blocked, EcoKI:variable"
                     /annotation_group="GTGCAC"
                     /label="ApaLI"
                     /note="Geneious type: restriction site"
     primer_bind     2427..2446
                     /Mismatches=0
                     /%GC="45.0"
                     /Hairpin_Tm="None"
                     /Self_Dimer_Tm="None"
                     /Tm="56.7"
                     /annotation_group="284608767823600e3cee0e8-f7af-4560-b390-
                     06c1876cb7c2"
                     /Sequence="TTTTTGTGATGCTCGTCAGG"
                     /created_by="primer3"
                     /label="SLS_152"
     terminator      2477..2504
                     /note="T7Te terminator"
                     /note="phage T7 early transcription terminator"
     CDS             complement(2528..2719)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low
                     copy number"
                     /note="rop"
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDH
                     ADELYRSCLARFGDDGENL"
     primer_bind     complement(2696..2729)
                     /Sequence="CGGGGCGTAAATGACCAAACAGGAAAAAACCGCC"
                     /Hairpin_Tm="46.4"
                     /Tm="72.7"
                     /Self_Dimer_Tm="None"
                     /%GC="52.9"
                     /created_by="primer3"
                     /label="15chlor_bb_rev"
     primer_bind     2710..2735
                     /Sequence="GTTTGGTCATTTACGCCCCGCCCTGC"
                     /Hairpin_Tm="None"
                     /Tm="70.4"
                     /Self_Dimer_Tm="None"
                     /%GC="61.5"
                     /created_by="primer3"
                     /label="15chlor_ins_fow"
     CDS             complement(2720..3379)
                     /codon_start=1
                     /gene="cat"
                     /product="chloramphenicol acetyltransferase"
                     /note="confers resistance to chloramphenicol"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAF
                     LKTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETF
                     SSLWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNV
                     ANMDNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGG
                     A"
                     /label="CmR"
     source          <2720..>3379
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     misc_feature    2723..3376
                     /Original_Bases="GAAAAACTCATCGAGCATCAAGTGAAACTGCAATTTATTCA
                     TATCAGGATTATCAATACCATATTTTTGAAAAAGCCGTTTCTGTAATGAAGGAGAAAA
                     CTCACCGAGGCAGTTCCATAGGATGGCAAGATCCTGGTATCGGTCTGCGATTCCGACT
                     CGTCCAACATCAATACAACCTATTAATTTCCCCTCGTCAAAAATAAGGTTATCAAGTG
                     AGAAATCACCATGAGTGACGACTGAATCCGGTGAGAATGGCAAAAGCTTATGCATTTC
                     TTTCCAGACTTGTTCAACAGGCCAGCCATTACGCTCGTCATCAAAATCACTCGCACCA
                     ACCAAACCGTTATTCATTCGTGATTGCGCCTGAGCGAGACGAAATACGCGATCGCCGT
                     TAAAAGGACAATTACAAACAGGAATCGAATGCAACCGGCGCAGGAACACTGCCAGCGC
                     ATCAACAATATTTTCACCTGAATCAGGATATTCTTCTAATACCTGGAATGCTGTTTTC
                     CCTGGGATCGCAGTGGTGAGTAACCATGCATCATCAGGAGTACGGATAAAATGCTTGA
                     TGGTCGGAAGAGGCATAAATTCCGTCAGCCAGTTTAGCCTGACCATCTCATCTGTAAC
                     ATCATTGGCAACGCTACCTTTGCCATGTTTCAGAAACAACTCTGGCGCATCGGGCTTC
                     CCATACAATCGATAGATTGTCGCACCTGATTGCCCGACATTATCGCGAGCCCATTTAT
                     ACCCATATAAATCAGCATCCATGTTGGAATTTAATCGCGGCCTCGAGCAAGACGTTTC
                     CCGTTGAATATGGCT"
                     /label="GAAAAACTCATCGAGCATCAAGTGAAACTGCAATTTATTCATATCAGGAT
                     TATCAATACCATATTTTTGAAAAAGCCGTTTCTGTAATGAAGGAGAAAACTCACCGAG
                     GCAGTTCCATAGGATGGCAAGATCCTGGTATCGGTCTGCGATTCCGACTCGTCCAACA
                     TCAATACAACCTATTAATTTCCCCTCGTCAAAAATAAGGTTATCAAGTGAGAAATCAC
                     CATGAGTGACGACTGAATCCGGTGAGAATGGCAAAAGCTTATGCATTTCTTTCCAGAC
                     TTGTTCAACAGGCCAGCCATTACGCTCGTCATCAAAATCACTCGCACCAACCAAACCG
                     TTATTCATTCGTGATTGCGCCTGAGCGAGACGAAATACGCGATCGCCGTTAAAAGGAC
                     AATTACAAACAGGAATCGAATGCAACCGGCGCAGGAACACTGCCAGCGCATCAACAAT
                     ATTTTCACCTGAATCAGGATATTCTTCTAATACCTGGAATGCTGTTTTCCCTGGGATC
                     GCAGTGGTGAGTAACCATGCATCATCAGGAGTACGGATAAAATGCTTGATGGTCGGAA
                     GAGGCATAAATTCCGTCAGCCAGTTTAGCCTGACCATCTCATCTGTAACATCATTGGC
                     AACGCTACCTTTGCCATGTTTCAGAAACAACTCTGGCGCATCGGGCTTCCCATACAAT
                     CGATAGATTGTCGCACCTGATTGCCCGACATTATCGCGAGCCCATTTATACCCATATA
                     AATCAGCATCCATGTTGGAATTTAATCGCGGCCTCGAGCAAGACGTTTCCCGTTGAAT
                     ATGGCT"
                     /note="Geneious type: Editing History Replacement"
     misc_feature    2746..2751
                     /Recognition_pattern="AGT^ACT"
                     /Methylation="EcoBI:blocked"
                     /annotation_group="AGTACT"
                     /label="ScaI"
                     /note="Geneious type: restriction site"
     misc_feature    2860..2865
                     /Recognition_pattern="C^CATGG"
                     /annotation_group="CCATGG"
                     /label="NcoI"
                     /note="Geneious type: restriction site"
     primer_bind     3290..3310
                     /%GC="42.9"
                     /Mismatches=0
                     /Hairpin_Tm="37.3"
                     /Self_Dimer_Tm="None"
                     /Tm="57.4"
                     /annotation_group="284609529004800df9f42da-f7d5-4345-afc4-
                     bc5e431afd2a;
                     284613060424300aa9deaef-f3a6-4b7a-a5f2-fe8c5ad60fd3"
                     /Sequence="TTGAGCAACTGACTGAAATGC"
                     /created_by="primer3"
                     /label="SBK_16_CmR; SBK_16_CmR"
     primer_bind     complement(3352..3389)
                     /Sequence="TTCAGGAGCTATGGAGAAAAAAATCACTGGATATACCA"
                     /Hairpin_Tm="45.7"
                     /Tm="60.5"
                     /Self_Dimer_Tm="None"
                     /%GC="36.8"
                     /created_by="primer3"
                     /label="15chlor_ins_rev"
     primer_bind     3370..3405
                     /Sequence="TTTTCTCCATAGCTCCTGAAAATCTCGATAACTCAA"
                     /Hairpin_Tm="31.3"
                     /Tm="65.8"
                     /Self_Dimer_Tm="None"
                     /%GC="36.1"
                     /created_by="primer3"
                     /label="15chlor_bb_fow"
     promoter        complement(3380..3470)
                     /note="cat promoter"
                     /note="promoter of the E. coli cat gene encoding 
                     chloramphenicol acetyltransferase"
     primer_bind     3444..3465
                     /%GC="50.0"
                     /Mismatches=0
                     /Hairpin_Tm="None"
                     /Self_Dimer_Tm="None"
                     /Tm="60.8"
                     /annotation_group="284610929636700bb793016-17cb-4ad3-bfa0-
                     25f53c6eb0a5;
                     28461462108890047adc909-4b38-46a0-8cd0-f7512c3e10f8"
                     /Sequence="AAGTTGGAACCTCTTACGTGCC"
                     /created_by="primer3"
                     /label="cat_promoter_rev_SBK_015; SBK_015_cat_prom_rev"
     terminator      3471..3502
                     /note="tonB terminator"
                     /note="bidirectional E. coli tonB-P14 transcription 
                     terminator"
     primer_bind     3476..3493
                     /%GC="66.7"
                     /Mismatches=0
                     /Hairpin_Tm="67.2"
                     /Self_Dimer_Tm="39.5"
                     /Tm="61.4"
                     /annotation_group="28460819875670001fc826b-54dc-4891-8cef-
                     83ea0d486bc6;
                     284613845270600579d5680-0ee3-4988-967d-7e00e732900c"
                     /Sequence="AAAGCCTCCGGTCGGAGG"
                     /created_by="primer3"
                     /label="SBK_013_tonB_term; tonB_term_SBK_013"
     primer_bind     complement(3508..3544)
                     /Mismatches=0
                     /%GC="43.2"
                     /Hairpin_Tm="29.6"
                     /Self_Dimer_Tm="14.2"
                     /Tm="61.8"
                     /annotation_group="8905124240520006522981-5d77-4e37-9cb4-e
                     527176f3356"
                     /Sequence="TGGGTCCTGACATTTAAATCATACCTGACCTCCATAG"
                     /created_by="primer3"
                     /label="dualProm_bb_rev_ins117"
     primer_bind     complement(3508..3544)
                     /Mismatches=0
                     /%GC="43.2"
                     /Hairpin_Tm="29.6"
                     /Self_Dimer_Tm="14.2"
                     /Tm="61.8"
                     /annotation_group="284612117768200d64849d9-efe7-4c7b-a7ad-
                     103ca6b5f23d"
                     /Sequence="TGGGTCCTGACATTTAAATCATACCTGACCTCCATAG"
                     /created_by="primer3"
                     /label="dualProm_bb_rev_ins117"
     primer_bind     complement(3508..3534)
                     /Mismatches=0
                     /%GC="37.0"
                     /Hairpin_Tm="33.2"
                     /Self_Dimer_Tm="None"
                     /Tm="57.8"
                     /annotation_group="115351088134910001111a19-7466-4c4e-9309
                     -dc5c6533375d"
                     /Sequence="CATTTAAATCATACCTGACCTCCATAG"
                     /created_by="primer3"
                     /label="dualProm_bb_rev"
     primer_bind     complement(3508..3534)
                     /Mismatches=0
                     /%GC="37.0"
                     /Hairpin_Tm="33.2"
                     /Self_Dimer_Tm="None"
                     /Tm="57.8"
                     /annotation_group="2846155611050008601f33c-5c03-4fda-8544-
                     3830205b34c7"
                     /Sequence="CATTTAAATCATACCTGACCTCCATAG"
                     /created_by="primer3"
                     /label="dualProm_bb_rev"
     primer_bind     3525..3560
                     /Mismatches=0
                     /%GC="33.3"
                     /Hairpin_Tm="46.1"
                     /Self_Dimer_Tm="18.6"
                     /Tm="64.6"
                     /annotation_group="115403040513610099b64131-2554-4f9e-9fbd
                     -0367a50af4bc"
                     /Sequence="GATTTAAATGTCAGGACCCACTTTCACATTTAAGTT"
                     /created_by="primer3"
                     /label="frag117_ins_fow"
     primer_bind     3525..3560
                     /Mismatches=0
                     /%GC="33.3"
                     /Hairpin_Tm="46.1"
                     /Self_Dimer_Tm="18.6"
                     /Tm="64.6"
                     /annotation_group="2846089631194005313b578-c4d9-4f43-8906-
                     21826b164259"
                     /Sequence="GATTTAAATGTCAGGACCCACTTTCACATTTAAGTT"
                     /created_by="primer3"
                     /label="frag117_ins_fow"
     misc_feature    3535..4574
                     /Original_Bases="GTCAGTATTGAGCGATATCTAGAGAATTCGTC"
                     /label="GTCAGTATTGAGCGATATCTAGAGAATTCGTC"
                     /note="Geneious type: Editing History Replacement"
     CDS             complement(3535..4158)
                     /label="TetR"
     source          join(complement(<3535..4496),complement(4497..>4574))
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     misc_feature    3575..3580
                     /Recognition_pattern="CA^TATG"
                     /Methylation="EcoBI:cut"
                     /annotation_group="CATATG"
                     /label="NdeI"
                     /note="Geneious type: restriction site"
     primer_bind     3867..3886
                     /#_Off-target_Sites=0
                     /Off-target_Template="pSBK.136 (not cloned)"
                     /Sequence="TTGCTCCATCGCGATGACTT"
                     /Hairpin_Tm="41.2"
                     /Tm="60.1"
                     /Self_Dimer_Tm="42.4"
                     /%GC="50.0"
                     /modified_by="santi.bhattaraikline"
                     /label="SBK_21_TetR"
     misc_feature    3875..3880
                     /Recognition_pattern="TCG^CGA"
                     /Methylation="Dam:blocked, CpG:blocked"
                     /annotation_group="TCGCGA"
                     /label="NruI"
                     /note="Geneious type: restriction site"
     RBS             complement(4159..4186)
                     /label="tet1"
     promoter        complement(4187..4240)
                     /label="pLacI"
     promoter        complement(4194..4199)
                     /label="-10"
     primer_bind     4201..4221
                     /%GC="52.4"
                     /Mismatches=0
                     /Hairpin_Tm="57.9"
                     /Self_Dimer_Tm="24.1"
                     /Tm="61.0"
                     /annotation_group="284611119131100dbc70368-d9b9-4e09-9220-
                     3350ec59cdbc;
                     284614810147000a4ef2f22-fcd7-498d-bf88-c5bdca226ba4"
                     /Sequence="CATACCGCGAAAGGTTTTGCG"
                     /created_by="primer3"
                     /label="SBK_19; SBK_19"
     promoter        complement(4217..4222)
                     /label="-35"
     terminator      complement(4245..4305)
                     /label="L3S2P21"
     primer_bind     complement(4282..4315)
                     /Mismatches=0
                     /%GC="35.3"
                     /Hairpin_Tm="None"
                     /Self_Dimer_Tm="13.5"
                     /Tm="64.6"
                     /annotation_group="1046738310769001cc628ef-d56a-4904-a5bc-
                     f6e220dbdcf5"
                     /Sequence="AATCTTATCACTCGGTACCAAATTCCAGAAAAGA"
                     /created_by="primer3"
                     /label="057_bb_fow"
     primer_bind     complement(4282..4315)
                     /Mismatches=0
                     /%GC="35.3"
                     /Hairpin_Tm="None"
                     /Self_Dimer_Tm="13.5"
                     /Tm="64.6"
                     /annotation_group="2846128708820007ca1546c-b534-4b25-b571-
                     0772ed7792fb"
                     /Sequence="AATCTTATCACTCGGTACCAAATTCCAGAAAAGA"
                     /created_by="primer3"
                     /label="057_bb_fow"
     primer_bind     4296..4328
                     /Mismatches=0
                     /%GC="51.5"
                     /Hairpin_Tm="36.7"
                     /Self_Dimer_Tm="14.9"
                     /Tm="70.6"
                     /annotation_group="10467476893350072e34779-e262-45a8-9e22-
                     09ffb8fb62b1"
                     /Sequence="TGGTACCGAGTGATAAGATTCCGTATGCGCACC"
                     /created_by="primer3"
                     /label="057_ins_rev"
     primer_bind     4296..4328
                     /Mismatches=0
                     /%GC="51.5"
                     /Hairpin_Tm="36.7"
                     /Self_Dimer_Tm="14.9"
                     /Tm="70.6"
                     /annotation_group="2846123053628002ec7c412-4059-401d-be69-
                     b8d1e21ccdb1"
                     /Sequence="TGGTACCGAGTGATAAGATTCCGTATGCGCACC"
                     /created_by="primer3"
                     /label="057_ins_rev"
     misc_feature    4306..4332
                     /created_by="User"
                     /label="'primer'"
     primer          complement(4323..>4333)
                     /created_by="User"
                     /label="86_fow"
     primer          complement(4325..>4333)
                     /modified_by="User"
                     /label="msd_Fow"
     primer_bind     4325..4347
                     /Mismatches=0
                     /Mismatches=0
                     /%GC="47.8"
                     /Hairpin_Tm="43.0"
                     /Self_Dimer_Tm="None"
                     /Tm="60.3"
                     /annotation_group="165377687888800f21d9da0-8e49-4780-b554-
                     feeb2e85ea00"
                     /Sequence="CACCCTTACGTCAGAAAAAACGG"
                     /created_by="primer3"
                     /label="020_4kld_rev"
     primer_bind     4325..4347
                     /Mismatches=0
                     /Mismatches=0
                     /%GC="47.8"
                     /Hairpin_Tm="43.0"
                     /Self_Dimer_Tm="None"
                     /Tm="60.3"
                     /annotation_group="10467439459930065f96e8e-e864-43dd-9624-
                     fec679ccd6d7"
                     /Sequence="CACCCTTACGTCAGAAAAAACGG"
                     /created_by="primer3"
                     /label="020_4kld_rev"
     primer_bind     4325..4347
                     /Mismatches=0
                     /Mismatches=0
                     /%GC="47.8"
                     /Hairpin_Tm="43.0"
                     /Self_Dimer_Tm="None"
                     /Tm="60.3"
                     /annotation_group="284609149801800ac0b08df-4da9-49dc-a205-
                     9be6e8c7f9f3"
                     /Sequence="CACCCTTACGTCAGAAAAAACGG"
                     /created_by="primer3"
                     /label="020_4kld_rev"
     misc_feature    4334..4390
                     /created_by="User"
                     /modified_by="Seth"
                     /label="msd"
     motif           4334..4348
                     /created_by="santi.bhattaraikline"
                     /label="Flanking"
     source          complement(<4349..>4373)
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     motif           4349..4358
                     /created_by="santi.bhattaraikline"
                     /label="Stem"
     motif           4359..4363
                     /created_by="santi.bhattaraikline"
                     /label="Loop"
     motif           complement(4364..4373)
                     /created_by="santi.bhattaraikline"
                     /label="Stem"
     motif           complement(4374..4390)
                     /created_by="santi.bhattaraikline"
                     /label="Flanking"
     misc_rna        complement(4380..4461)
                     /created_by="User"
                     /Transferred_From="template"
                     /Transferred_Similarity="100.00%"
                     /label="msdRNA"
     primer          <4391..4407
                     /created_by="User"
                     /label="86_rev"
     primer          <4391..4401
                     /modified_by="User"
                     /label="msd_Rev"
     motif           4400..4407
                     /created_by="santi.bhattaraikline"
                     /label="Stem (RT Recognition)"
     motif           4408..4410
                     /created_by="santi.bhattaraikline"
                     /label="UUU Loop (RT Recognition)"
     primer_bind     complement(4408..4433)
                     /Mismatches=0
                     /%GC="38.5"
                     /Hairpin_Tm="48.0"
                     /Self_Dimer_Tm="14.8"
                     /Tm="60.9"
                     /annotation_group="165377492396500db0f4f82-183f-4e01-b63f-
                     ecc181db7fa8"
                     /Sequence="TTAAGGTCAACCTCTGGATGTTGTTT"
                     /created_by="primer3"
                     /label="Uloop_detect_fow_wt"
     primer_bind     complement(4408..4433)
                     /Mismatches=0
                     /%GC="38.5"
                     /Hairpin_Tm="48.0"
                     /Self_Dimer_Tm="14.8"
                     /Tm="60.9"
                     /annotation_group="104675332340500b6b4c216-dd69-42a3-86af-
                     7cb84c005db4"
                     /Sequence="TTAAGGTCAACCTCTGGATGTTGTTT"
                     /created_by="primer3"
                     /label="Uloop_detect_fow_wt"
     primer_bind     complement(4408..4433)
                     /Mismatches=0
                     /%GC="38.5"
                     /Hairpin_Tm="48.0"
                     /Self_Dimer_Tm="14.8"
                     /Tm="60.9"
                     /annotation_group="28460990539680066a9f9fe-faf9-4d9f-aa59-
                     7a81144c0066"
                     /Sequence="TTAAGGTCAACCTCTGGATGTTGTTT"
                     /created_by="primer3"
                     /label="Uloop_detect_fow_wt"
     motif           complement(4411..4418)
                     /created_by="santi.bhattaraikline"
                     /label="Stem (RT Recognition)"
     primer_bind     complement(4442..4468)
                     /Mismatches=0
                     /%GC="55.6"
                     /Hairpin_Tm="57.4"
                     /Self_Dimer_Tm="14.9"
                     /Tm="68.4"
                     /annotation_group="19622856696510097a7a7d5-5d46-40f7-a146-
                     0baca6068f1f"
                     /Sequence="ATTCCGTATGCGCACCCTTAGCGAGAG"
                     /created_by="primer3"
                     /label="pSBK_106_4kld_fow"
     primer_bind     complement(4442..4468)
                     /Mismatches=0
                     /%GC="55.6"
                     /Hairpin_Tm="57.4"
                     /Self_Dimer_Tm="14.9"
                     /Tm="68.4"
                     /annotation_group="284615372842900162405eb-659b-4195-85b7-
                     4e3d3c0f1555"
                     /Sequence="ATTCCGTATGCGCACCCTTAGCGAGAG"
                     /created_by="primer3"
                     /label="106_4kld_fow"
     misc_feature    4449..4460
                     /created_by="User"
                     /Transferred_From="template"
                     /Transferred_Similarity="100.00%"
                     /label="'primer'"
     misc_signal     4462..>4475
                     /created_by="User"
                     /Transferred_From="template"
                     /Transferred_Similarity="100.00%"
                     /label="T7 to msr"
     source          complement(<4462..>4475)
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     primer_bind     4469..4496
                     /Mismatches=0
                     /%GC="50.0"
                     /Hairpin_Tm="51.3"
                     /Self_Dimer_Tm="None"
                     /Tm="65.5"
                     /annotation_group="1962287576682000334322d-31ce-4e91-8341-
                     d6d9e87a5a66"
                     /Sequence="CTTATCAGGAAAGCACATCCGGTGACAG"
                     /created_by="primer3"
                     /label="pSBK_106_4kld_rev"
     primer_bind     4469..4496
                     /Mismatches=0
                     /%GC="50.0"
                     /Hairpin_Tm="51.3"
                     /Self_Dimer_Tm="None"
                     /Tm="65.5"
                     /annotation_group="284613461240400cb00745a-1aec-4798-9bcd-
                     f8ee49bdd57d"
                     /Sequence="CTTATCAGGAAAGCACATCCGGTGACAG"
                     /created_by="primer3"
                     /label="106_4kld_rev"
     misc_RNA        complement(<4476..4496)
                     /note="hammerhead ribozyme from the tobacco ringspot virus
                      satellite RNA (Khvorova et al., 2003)"
                     /label="sTRSV HHRz"
     promoter        complement(4499..4574)
                     /label="pTet*"
     promoter        complement(4505..4510)
                     /label="-10"
     operator        complement(4509..4527)
                     /label="tetO2"
     promoter        complement(4528..4533)
                     /label="-35"
     operator        complement(4534..4552)
                     /label="TetO2"
     primer_bind     complement(4554..48)
                     /modified_by="santi.bhattaraikline"
                     /Sequence="ATTAATAATCTGGCTTTTTATATTCTCTTAGAGCTGGGTTCGGACGC
                     GTTTTCAGCAGGACGCACTGAC"
                     /Mismatches=0
                     /annotation_group="9104e95b-fc31-41f3-a4fe-e7017d0d3a60"
                     /label="frag117_ins_rev_v2"
     primer_bind     complement(4554..48)
                     /modified_by="santi.bhattaraikline"
                     /Sequence="ATTAATAATCTGGCTTTTTATATTCTCTTAGAGCTGGGTTCGGACGC
                     GTTTTCAGCAGGACGCACTGAC"
                     /Mismatches=0
                     /annotation_group="058f7975-2e0c-443f-b824-4d409ae695cb"
                     /label="frag117_ins_rev_v2"
ORIGIN      
        1 cgcgtccgaa cccagctcta agagaatata aaaagccaga ttattaatcc ggctttttta
       61 ttatttcgct taacgatcgt tggctggggg cctcgcttgg gttattgctg gtgcccggcc
      121 gggcgcaata ttcatgttga tgatttatta tatatcgagt ggtgtattta tttatattgt
      181 ttgctccgtt accgttatta acagctgtca ccggatgtgc tttcctgata agattccgta
      241 tgcgcaccct tagcgagagg tttatcatta aggtcaacct ctggatgttg tttcggcatc
      301 ctgcattgaa tctgagttac aagaaaatgg agaggttgca gcaacctctc cacccgtttt
      361 ttctgacgta agggtgcgca tacggaatct tatcactcgg taccaaattc cagaaaagag
      421 gcctcccgaa aggggggcct ttttcgtttt ggtccaatgg cggcgcgcca tcgaatggtg
      481 caaaaccttt cgcggtatgg catgatagcg cccggaagag agtcaattca tgggggttga
      541 atatggaact gcgtgacctg gatttaaacc tgctggtggt gttcaaccag ttgctggtcg
      601 acagacgcgt ctctgtcact gcggagaacc tgggcctgac ccagcctgcc gtgagcaatg
      661 cgctgaaacg cctgcgcacc tcgctacagg acccactctt cgtgcgcaca catcagggaa
      721 tggaacccac accctatgcc gcgcatctgg ccgagcacgt cacttcggcc atgcacgcac
      781 tgcgcaacgc cctacagcac catgaaagct tcgatccgct gaccagcgag cgtaccttca
      841 ccctggccat gaccgacatt ggcgagatct acttcatgcc gcggctgatg gatgcgctgg
      901 ctcaccaggc ccccaattgc gtgatcagta cggtgcgcga cagttcgatg agcctgatgc
      961 aggccttgca gaacggaacc gtggacttgg ccgtgggcct gcttcccaat ctgcaaactg
     1021 gcttctttca gcgccggctg ctccagaatc actacgtgtg cctatgtcgc aaggaccatc
     1081 cagtcacccg cgaacccctg actctggagc gcttctgttc ctacggccac gtgcgtgtca
     1141 tcgccgctgg caccggccac ggcgaggtgg acacgtacat gacacgggtc ggcatccggc
     1201 gcgacatccg tctggaagtg ccgcacttcg ccgccgttgg ccacatcctc cagcgcaccg
     1261 atctgctcgc cactgtgccg atatgtttag ccgactgctg cgtagagccc ttcggcctaa
     1321 gcgccttgcc gcacccagtc gtcttgcctg aaatagccat caacatgttc tggcatgcga
     1381 agtaccacaa ggacctagcc aatatttggt tgcggcaact gatgtttgac ctgtttacgg
     1441 attgataagg atcctaattg gtaacgaatc agacaattga cggctcgagg gagtagcata
     1501 gggtttgcag aatccctgct tcgtccattt gacaggcaca ttatgcatcg atgataagct
     1561 gtcaaacatg agcagatcct ctacgccgga cgcatcgtgg ccggcatcac cggcgccaca
     1621 ggtgcggttg ctggcgccta tatcgccgac atcaccgatg gggaagatcg ggctcgccac
     1681 ttcgggctca tgagcaaata ttttatctga ggtgcttcct cgctcactgc tgaaaggagg
     1741 aactatatcc gggtaacgaa ttcaagcttg atatcattca ggacgagcct cagactccag
     1801 cgtaactgga ctgcaatcaa ctcactggct caccttcacg ggtgggcctt tcttcggtag
     1861 aaaatcaaag gatcttcttg agatcctttt tttctgcgcg taatctgctg cttgcaaaca
     1921 aaaaaaccac cgctaccagc ggtggtttgt ttgccggatc aagagctacc aactcttttt
     1981 ccgaggtaac tggcttcagc agagcgcaga taccaaatac tgttcttcta gtgtagccgt
     2041 agttaggcca ccacttcaag aactctgtag caccgcctac atacctcgct ctgctaatcc
     2101 tgttaccagt ggctgctgcc agtggcgata agtcgtgtct taccgggttg gactcaagac
     2161 gatagttacc ggataaggcg cagcggtcgg gctgaacggg gggttcgtgc acacagccca
     2221 gcttggagcg aacgacctac accgaactga gatacctaca gcgtgagcta tgagaaagcg
     2281 ccacgcttcc cgaagggaga aaggcggaca ggtatccggt aagcggcagg gtcggaacag
     2341 gagagcgcac gagggagctt ccagggggaa acgcctggta tctttatagt cctgtcgggt
     2401 ttcgccacct ctgacttgag catcgatttt tgtgatgctc gtcagggggg cggagcctat
     2461 ggaaaaacgc cagcaacgca gaaaggccca cccgaaggtg agccaggtga ttacatttgg
     2521 gccctcatca gaggttttca ccgtcatcac cgaaacgcgc gaggcagctg cggtaaagct
     2581 catcagcgtg gtcgtgaagc gattcacaga tgtctgcctg ttcatccgcg tccagctcgt
     2641 tgagtttctc cagaagcgtt aatgtctggc ttctgataaa gcgggccatg ttaagggcgg
     2701 ttttttcctg tttggtcatt tacgccccgc cctgccactc atcgcagtac tgttgtaatt
     2761 cattaagcat tctgccgaca tggaagccat cacaaacggc atgatgaacc tgaatcgcca
     2821 gcggcatcag caccttgtcg ccttgcgtat aatatttgcc catggtgaaa acgggggcga
     2881 agaagttgtc catattggcc acgtttaaat caaaactggt gaaactcacc cagggattgg
     2941 ctgagacgaa aaacatattc tcaataaacc ctttagggaa ataggccagg ttttcaccgt
     3001 aacacgccac atcttgcgaa tatatgtgta gaaactgccg gaaatcgtcg tggtattcac
     3061 tccagagcga tgaaaacgtt tcagtttgct catggaaaac ggtgtaacaa gggtgaacac
     3121 tatcccatat caccagctca ccgtctttca ttgccatacg aaattccgga tgagcattca
     3181 tcaggcgggc aagaatgtga ataaaggccg gataaaactt gtgcttattt ttctttacgg
     3241 tctttaaaaa ggccgtaata tccagctgaa cggtctggtt ataggtacat tgagcaactg
     3301 actgaaatgc ctcaaaatgt tctttacgat gccattggga tatatcaacg gtggtatatc
     3361 cagtgatttt tttctccata gctcctgaaa atctcgataa ctcaaaaaat acgcccggta
     3421 gtgatcttat ttcattatgg tgaaagttgg aacctcttac gtgccgatca agtcaaaagc
     3481 ctccggtcgg aggcttttga ctttctgcta tggaggtcag gtatgattta aatgtcagga
     3541 cccactttca catttaagtt gtttttctaa tccgcatatg atcaattcaa ggccgaataa
     3601 gaaggctggc tctgcacctt ggtgatcaaa taattcgata gcttgtcgta ataatggcgg
     3661 catactatca gtagtaggtg tttccctttc ttctttagcg acttgatgct cttgatcttc
     3721 caatacgcaa cctaaagtaa aatgccccac agcgctgagt gcatataatg cattctctag
     3781 tgaaaaacct tgttggcata aaaaggctaa ttgattttcg agagtttcat actgtttttc
     3841 tgtaggccgt gtacctaaat gtacttttgc tccatcgcga tgacttagta aagcacatct
     3901 aaaactttta gcgttattac gtaaaaaatc ttgccagctt tccccttcta aagggcaaaa
     3961 gtgagtatgg tgcctatcta acatctcaat ggctaaggcg tcgagcaaag cccgcttatt
     4021 ttttacatgc caatacaatg taggctgctc tacacctagc ttctgggcga gtttacgggt
     4081 tgttaaacct tcgattccga cctcattaag cagctctaat gcgctgttaa tcactttact
     4141 tttatctaat ctggacatat tcaccaccct gaattgactc tcttccgggc gctatcatgc
     4201 cataccgcga aaggttttgc gccattcgat ggcgcgccgc cattggacca aaacgaaaaa
     4261 aggcccccct ttcgggaggc ctcttttctg gaatttggta ccgagtgata agattccgta
     4321 tgcgcaccct tacgtcagaa aaaacgggtg gagaggtcag tctgacctct ccattttctt
     4381 gtaactcaga ttcaatgcag gatgccgaaa caacatccag aggttgacct taatgataaa
     4441 cctctcgcta agggtgcgca tacggaatct tatcaggaaa gcacatccgg tgacagctgt
     4501 gctcagtatc tctatcactg atagggatgt caatctctat cactgatagg gaggtcagtg
     4561 cgtcctgctg aaaa
//
