LOCUS       Exported                2254 bp ds-DNA     circular SYN 21-FEB-2022
DEFINITION  .
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 2254)
  AUTHORS   Carolyn Huang
  TITLE     Direct Submission
  JOURNAL   Exported Tuesday, Feb 22, 2022 from SnapGene 5.0.8
            https://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..2254
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     terminator      2..19
                     /label=L3S2P55 Terminator (Voigt)
                     /note="A strong synthetic promoter from Voigt's Nature 
                     Methods paper
                     
                     Also used by Alex Nielsen as a dSpyCas9 sgRNA terminator in
                     his CRISPRi logic gates paper
                     "
                     /note="color: #ffffff; direction: RIGHT"
     protein_bind    20..23
                     /label=BsaI
                     /note="color: #00ff2c"
     misc_feature    24..48
                     /label=ConE scar
                     /note="color: #fffd6c"
     protein_bind    24..27
                     /label=BsmBI
                     /note="color: #00ff15"
     terminator      complement(68..124)
                     /label=ECK120033737 Terminator (Voigt)
                     /note="rho-independent terminator of thrLABC operon"
                     /note="color: #ffffff; direction: LEFT"
     misc_feature    complement(125..233)
                     /label=CamR Terminator
                     /note="color: #66ccff; direction: LEFT"
     CDS             complement(234..893)
                     /label=CamR
                     /note="color: #0000ff; direction: LEFT"
     promoter        complement(894..998)
                     /label=CamR Promoter
                     /note="color: #66ccff; direction: LEFT"
     rep_origin      complement(1266..1810)
                     /direction=LEFT
                     /label=p15A ori
                     /note="Plasmids containing the medium-copy-number p15A 
                     origin of replication can be propagated in E. coli cells 
                     that contain a second plasmid with the ColE1 origin.
                     "
                     /note="color: #ffff00"
     terminator      1911..2000
                     /label=ECK120029600 Terminator (Voigt)
                     /note="rho-independent terminator of spheroplast protein Y 
                     transcript"
                     /note="color: #ffffff; direction: RIGHT"
     misc_feature    2020..2044
                     /label=ConS scar
                     /note="color: #fffd6c"
     protein_bind    2020..2023
                     /label=BsmBI
                     /note="color: #00ff15"
     misc_feature    2045..2048
                     /label=Pt2/2a BsaI Junction Scar
                     /note="color: #800080"
     promoter        2049..2146
                     /label=proD Promoter (strong constitutive)
                     /note="from ""Design, construction and characterization of 
                     a set of insulated bacterial promoters""
                     
                     The paper says this is synthetic. In other words, I assumed
                     this was from a ProD gene, but it does not appear to be the
                     case; specifically, the proline pathway in E. coli has 
                     genes proABC but not proD...
                     "
                     /note="This forward directional feature has 6 segments:
                        1: 2049 .. 2110 / #339966
                        2: 2111 .. 2116 / #00ff00 / -35
                        3: 2117 .. 2133 / #339966
                        4: 2134 .. 2138 / #00ff00 / -10
                        5: 2139 .. 2145 / #339966
                        6: 2146 .. 2146 / #00ff00"
     misc_feature    2149..2212
                     /label=Transcript
                     /note="color: #a6acb3; direction: RIGHT"
     misc_feature    2149..2184
                     /label=LbCpf1 DR
                     /note="color: #000000; direction: BOTH"
     misc_feature    2165..2174
                     /label=Conserved across As/Lb/Fn/Mb/Pm
                     /note="color: #33cccc"
     misc_feature    2185..2212
                     /label=TTTV PAM targeting Qi GFP
                     /label=sfGFP
                     /note="ApEinfo_revcolor=green
                     ApEinfo_fwdcolor=#00f700
                     ApEinfo_graphicformat=arrow_data {{0 1 2 0 0 -1} {} 0} 
                     width 5 offset 0"
                     /note="color: #00f700"
     misc_feature    complement(2185..2186)
                     /label=SauCas9 PAM (less stringent)
                     /note="color: #3366ff; direction: LEFT"
     misc_feature    2186..2191
                     /label=SauCas9 PAM (less stringent)
                     /note="color: #3366ff; direction: RIGHT"
     misc_feature    2187..2192
                     /label=SauCas9 PAM
                     /note="color: #0000ff; direction: RIGHT"
     misc_feature    complement(2193..2212)
                     /label=Strutt SauCas9 noPAM GFP2
                     /note="This reverse directional feature has 2 segments:
                        1: 2193 .. 2198 / #008000
                        2: 2199 .. 2212 / #339966"
     misc_feature    complement(2194..2199)
                     /label=SauCas9 PAM (less stringent)
                     /note="color: #3366ff; direction: LEFT"
     misc_feature    complement(2195..2200)
                     /label=SauCas9 PAM (less stringent)
                     /note="color: #3366ff; direction: LEFT"
     misc_feature    complement(2196..2212)
                     /label=NGG PAM SpyCas9 GFP1 (Best Qi)
                     /note="This reverse directional feature has 2 segments:
                        1: 2196 .. 2198 / #ff0000 / PAM
                        2: 2199 .. 2212 / #00ff00 / Spacer"
     misc_feature    complement(2200..2205)
                     /label=SauCas9 PAM
                     /note="color: #0000ff; direction: LEFT"
     misc_feature    2207..2212
                     /label=SauCas9 PAM
                     /note="color: #0000ff; direction: RIGHT"
     terminator      join(2213..2254,1..19)
                     /label=L3S2P21 Terminator (Voigt)
                     /note="Strongest synthetic promoter from Voigt's Nature 
                     Methods paper
                     
                     Also used by Alex Nielsen as a dSpyCas9 sgRNA terminator in
                     his CRISPRi logic gates paper
                     "
                     /note="color: #ffffff; direction: RIGHT"
ORIGIN
        1 ccttttttcg ttttggtccg ctgagcagtt acagagatgt tacgaaccac tagtgcactg
       61 cagtacacct ttggtcgaaa aaaaaagccc gcactgtcag gtgcgggctt ttttctgtgt
      121 ttccaccaat aaaaaacgcc cggcggcaac cgagcgttct gaacaaatcc agatggagtt
      181 ctgaggtcat tactggatct atcaacagga gtccaagcga gctcgatatc aaattacgcc
      241 ccgccctgcc actcatcgca gtactgttgt aattcattaa gcattctgcc gacatggaag
      301 ccatcacaaa cggcatgatg aacctgaatc gccagcggca tcagcacctt gtcgccttgc
      361 gtataatatt tgcccatggt gaaaacgggg gcgaagaagt tgtccatatt ggccacgttt
      421 aaatcaaaac tggtgaaact cacccaggga ttggctgaaa cgaaaaacat attctcaata
      481 aaccctttag ggaaataggc caggttttca ccgtaacacg ccacatcttg cgaatatatg
      541 tgtagaaact gccggaaatc gtcgtggtat tcactccaga gcgatgaaaa cgtttcagtt
      601 tgctcatgga aaacggtgta acaagggtga acactatccc atatcaccag ctcaccgtct
      661 ttcattgcca tacgaaattc cggatgagca ttcatcaggc gggcaagaat gtgaataaag
      721 gccggataaa acttgtgctt atttttcttt acggtcttta aaaaggccgt aatatccagc
      781 tgaacggtct ggttataggt acattgagca actgactgaa atgcctcaaa atgttcttta
      841 cgatgccatt gggatatatc aacggtggta tatccagtga tttttttctc cattttagct
      901 tccttagctc ctgaaaatct cgataactca aaaaatacgc ccggtagtga tcttatttca
      961 ttatggtgaa agttggaacc tcttacgtgc ccgatcaagc gctagcggag tgtatactgg
     1021 cttactatgt tggcactgat gagggtgtca gtgaagtgct tcatgtggca ggagaaaaaa
     1081 ggctgcaccg gtgcgtcagc agaatatgtg atacaggata tattccgctt cctcgctcac
     1141 tgactcgcta cgctcggtcg ttcgactgcg gcgagcggaa atggcttacg aacggggcgg
     1201 agatttcctg gaagatgcca ggaagatact taacagggaa gtgagagggc cgcggcaaag
     1261 ccgtttttcc ataggctccg cccccctgac aagcatcacg aaatctgacg ctcaaatcag
     1321 tggtggcgaa acccgacagg actataaaga taccaggcgt ttcccctggc ggctccctcg
     1381 tgcgctctcc tgttcctgcc tttcggttta ccggtgtcat tccgctgtta tggccgcgtt
     1441 tgtctcattc cacgcctgac actcagttcc gggtaggcag ttcgctccaa gctggactgt
     1501 atgcacgaac cccccgttca gtccgaccgc tgcgccttat ccggtaacta tcgtcttgag
     1561 tccaacccgg aaagacatgc aaaagcacca ctggcagcag ccactggtaa ttgatttaga
     1621 ggagttagtc ttgaagtcat gcgccggtta aggctaaact gaaaggacaa gttttggtga
     1681 ctgcgctcct ccaagccagt tacctcggtt caaagagttg gtagctcaga gaaccttcga
     1741 aaaaccgccc tgcaaggcgg ttttttcgtt ttcagagcaa gagattacgc gcagaccaaa
     1801 acgatctcaa gaagatcatc ttattaatca gataaaatat ttctagattt cagtgcaatt
     1861 tatctcttca aatgtagcac ctgaagtcag ccccatacga tataagttgt ttcagccaaa
     1921 aaacttaaga ccgccggtct tgtccactac cttgcagtaa tgcggtggac aggatcggcg
     1981 gttttctttt ctcttctcaa ccctgaattc gcatctagac tgaactggcc gataattgca
     2041 gacgaacgca cagctaacac cacgtcgtcc ctatctgctg ccctaggtct atgagtggtt
     2101 gctggataac tttacgggca tgcataaggc tcgtataata tattcagggt ttcaaagatt
     2161 aaataatttc tactaagtgt agatactgga gttgtcccaa ttcttgttga atctcggtac
     2221 caaattccag aaaagaggcc tcccgaaagg gggg
//
