LOCUS       sequence-366335-        4348 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  This plasmid carries Yn situ preamplifier A1-20B1 sequence and is 
            used for nickase based synthesis..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4348)
  AUTHORS   Wu Y, Xu W, Ma L, Yu Z, Wang Y, Yu CR
  TITLE     Robust and sensitive in situ RNA detection using Yn-situ.
  JOURNAL   Cell Rep Methods. 2022 Apr 18;2(4):100201. doi: 
            10.1016/j.crmeth.2022.100201. eCollection 2022 Apr 25.
  PUBMED    35497500
REFERENCE   2  (bases 1 to 4348)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Dec 15, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Cell Rep 
            Methods."; date: "2022-04-18"; volume: "2(4):100201. doi"; pages: " 
            10.1016/j.crmeth.2022.100201. eCollection 2022 Apr 25"
COMMENT     Sequence Label: A1-20B1(P004)
FEATURES             Location/Qualifiers
     source          1..4348
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      1..589
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     490..509
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     743..760
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    877..898
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(913..918,919..936,937..943)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    951..967
                     /label=lac operator
                     /bound_moiety="lac repressor encoded by lacI"
                     /note="The lac repressor binds to the lac operator to 
                     inhibit transcription in E. coli. This inhibition can be 
                     relieved by adding lactose or 
                     isopropyl-beta-D-thiogalactopyranoside (IPTG)."
     primer_bind     956..978
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     975..991
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     975..991
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     promoter        1009..1027
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     1009..1026
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     primer_bind     complement(2006..2025)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(2007..2025)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(2032..2049)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(2032..2048)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2041..2063)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     CDS             2186..2488
                     /codon_start=1
                     /gene="ccdB"
                     /product="CcdB, a bacterial toxin that poisons DNA gyrase"
                     /label=ccdB
                     /note="Plasmids containing the ccdB gene cannot be 
                     propagated in standard E. coli strains."
                     /translation="QFKVYTYKRESRYRLFVDVQSDIIDTPGRRMVIPLASARLLSDKV
                     SRELYPVVHIGDESWRMMTTDMASVPVSVIGEEVADLSHRENDIKNAINLMFWGI"
     primer_bind     2367..2386
                     /label=ccdB-fwd
                     /note="ccdB gene, forward primer"
     CDS             2837..3631
                     /codon_start=1
                     /product="aminoglycoside phosphotransferase"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     CDS             2837..3631
                     /codon_start=1
                     /gene="aph(3')-II (or nptII)"
                     /product="aminoglycoside phosphotransferase from Tn5"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(2891..2910)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     3501..3520
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     regulatory      3831..3840
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             3837..4211
                     /codon_start=1
                     /gene="Sh ble from Streptoalloteichus hindustanus"
                     /product="antibiotic-binding protein"
                     /label=BleoR
                     /note="confers resistance to bleomycin, phleomycin, and 
                     Zeocin(TM)"
                     /translation="MAKLTSAVPVLTARDVAGAVEFWTDRLGFSRDFVEDDFAGVVRDD
                     VTLFISAVQDQVVPDNTLAWVWVRGLDELYAEWSEVVSTNFRDASGPAMTEIGEQPWGR
                     EFALRDPAGNCVHFVAEEQD"
ORIGIN
        1 ttgagatcct ttttttctgc gcgtaatctg ctgcttgcaa acaaaaaaac caccgctacc
       61 agcggtggtt tgtttgccgg atcaagagct accaactctt tttccgaagg taactggctt
      121 cagcagagcg cagataccaa atactgttct tctagtgtag ccgtagttag gccaccactt
      181 caagaactct gtagcaccgc ctacatacct cgctctgcta atcctgttac cagtggctgc
      241 tgccagtggc gataagtcgt gtcttaccgg gttggactca agacgatagt taccggataa
      301 ggcgcagcgg tcgggctgaa cggggggttc gtgcacacag cccagcttgg agcgaacgac
      361 ctacaccgaa ctgagatacc tacagcgtga gctatgagaa agcgccacgc ttcccgaagg
      421 gagaaaggcg gacaggtatc cggtaagcgg cagggtcgga acaggagagc gcacgaggga
      481 gcttccaggg ggaaacgcct ggtatcttta tagtcctgtc gggtttcgcc acctctgact
      541 tgagcgtcga tttttgtgat gctcgtcagg ggggcggagc ctatggaaaa acgccagcaa
      601 cgcggccttt ttacggttcc tggccttttg ctggcctttt gctcacatgt tctttcctgc
      661 gttatcccct gattctgtgg ataaccgtat taccgccttt gagtgagctg ataccgctcg
      721 ccgcagccga acgaccgagc gcagcgagtc agtgagcgag gaagcggaag agcgcccaat
      781 acgcaaaccg cctctccccg cgcgttggcc gattcattaa tgcagctggc acgacaggtt
      841 tcccgactgg aaagcgggca gtgagcgcaa cgcaattaat gtgagttagc tcactcatta
      901 ggcaccccag gctttacact ttatgcttcc ggctcgtatg ttgtgtggaa ttgtgagcgg
      961 ataacaattt cacacaggaa acagctatga ccatgattac gccaagctat ttaggtgaca
     1021 ctatagaata ctcaagctat gcatcaagct tggtaccgag ctcggatcca ctagtaacgg
     1081 ccgccagtgt gctggaattc gcccttgctc ttcgcaatgg cttgtcgttg acgtgacgta
     1141 tgattctcta agaggagggc agcaaacggg aagagtcttc ctttacgaag aggagggcag
     1201 caaacgggaa gagtcttcct ttacgaagag gagggcagca aacgggaaga gtcttccttt
     1261 acgaagagga gggcagcaaa cgggaagagt cttcctttac gaagaggagg gcagcaaacg
     1321 ggaagagtct tcctttacga agaggagggc agcaaacggg aagagtcttc ctttacgaag
     1381 aggagggcag caaacgggaa gagtcttcct ttacgaagag gagggcagca aacgggaaga
     1441 gtcttccttt acgaagagga gggcagcaaa cgggaagagt cttcctttac gaagaggagg
     1501 gcagcaaacg ggaagagtct tcctttacga agaggagggc agcaaacggg aagagtcttc
     1561 ctttacgaag aggagggcag caaacgggaa gagtcttcct ttacgaagag gagggcagca
     1621 aacgggaaga gtcttccttt acgaagagga gggcagcaaa cgggaagagt cttcctttac
     1681 gaagaggagg gcagcaaacg ggaagagtct tcctttacga agaggagggc agcaaacggg
     1741 aagagtcttc ctttacgaag aggagggcag caaacgggaa gagtcttcct ttacgaagag
     1801 gagggcagca aacgggaaga gtcttccttt acgaagagga gggcagcaaa cgggaagagt
     1861 cttcctttac gaagaggagg gcagcaaacg ggaagagtct tcctttacgt atagtgagtc
     1921 gtagctcttc gcaatgaagg gcgaattctg cagatatcca tcacactggc ggccgctcga
     1981 gcatgcatct agagggccca attcgcccta tagtgagtcg tattacaatt cactggccgt
     2041 cgttttacaa cgtcgtgact gggaaaaccc tggcgttacc caacttaatc gccttgcagc
     2101 acatccccct ttcgccagct ggcgtaatag cgaagaggcc cgcaccgatc gcccttccca
     2161 acagttgcgc agcctatacg tacggcagtt taaggtttac acctataaaa gagagagccg
     2221 ttatcgtctg tttgtggatg tacagagtga tattattgac acgccggggc gacggatggt
     2281 gatccccctg gccagtgcac gtctgctgtc agataaagtc tcccgtgaac tttacccggt
     2341 ggtgcatatc ggggatgaaa gctggcgcat gatgaccacc gatatggcca gtgtgccggt
     2401 ctccgttatc ggggaagaag tggctgatct cagccaccgc gaaaatgaca tcaaaaacgc
     2461 cattaacctg atgttctggg gaatataaat gtcaggcatg agattatcaa aaaggatctt
     2521 cacctagatc cttttcacgt agaaagccag tccgcagaaa cggtgctgac cccggatgaa
     2581 tgtcagctac tgggctatct ggacaaggga aaacgcaagc gcaaagagaa agcaggtagc
     2641 ttgcagtggg cttacatggc gatagctaga ctgggcggtt ttatggacag caagcgaacc
     2701 ggaattgcca gctggggcgc cctctggtaa ggttgggaag ccctgcaaag taaactggat
     2761 ggctttcttg ccgccaagga tctgatggcg caggggatca agctctgatc aagagacagg
     2821 atgaggatcg tttcgcatga ttgaacaaga tggattgcac gcaggttctc cggccgcttg
     2881 ggtggagagg ctattcggct atgactgggc acaacagaca atcggctgct ctgatgccgc
     2941 cgtgttccgg ctgtcagcgc aggggcgccc ggttcttttt gtcaagaccg acctgtccgg
     3001 tgccctgaat gaactgcaag acgaggcagc gcggctatcg tggctggcca cgacgggcgt
     3061 tccttgcgca gctgtgctcg acgttgtcac tgaagcggga agggactggc tgctattggg
     3121 cgaagtgccg gggcaggatc tcctgtcatc tcaccttgct cctgccgaga aagtatccat
     3181 catggctgat gcaatgcggc ggctgcatac gcttgatccg gctacctgcc cattcgacca
     3241 ccaagcgaaa catcgcatcg agcgagcacg tactcggatg gaagccggtc ttgtcgatca
     3301 ggatgatctg gacgaagagc atcaggggct cgcgccagcc gaactgttcg ccaggctcaa
     3361 ggcgagcatg cccgacggcg aggatctcgt cgtgacccat ggcgatgcct gcttgccgaa
     3421 tatcatggtg gaaaatggcc gcttttctgg attcatcgac tgtggccggc tgggtgtggc
     3481 ggaccgctat caggacatag cgttggctac ccgtgatatt gctgaagagc ttggcggcga
     3541 atgggctgac cgcttcctcg tgctttacgg tatcgccgct cccgattcgc agcgcatcgc
     3601 cttctatcgc cttcttgacg agttcttctg aattattaac gcttacaatt tcctgatgcg
     3661 gtattttctc cttacgcatc tgtgcggtat ttcacaccgc atcaggtggc acttttcggg
     3721 gaaatgtgcg cggaacccct atttgtttat ttttctaaat acattcaaat atgtatccgc
     3781 tcatgagaca ataaccctga taaatgcttc aataatagca cgtgaggagg gccaccatgg
     3841 ccaagttgac cagtgccgtt ccggtgctca ccgcgcgcga cgtcgccgga gcggtcgagt
     3901 tctggaccga ccggctcggg ttctcccggg acttcgtgga ggacgacttc gccggtgtgg
     3961 tccgggacga cgtgaccctg ttcatcagcg cggtccagga ccaggtggtg ccggacaaca
     4021 ccctggcctg ggtgtgggtg cgcggcctgg acgagctgta cgccgagtgg tcggaggtcg
     4081 tgtccacgaa cttccgggac gcctccgggc cggccatgac cgagatcggc gagcagccgt
     4141 gggggcggga gttcgccctg cgcgacccgg ccggcaactg cgtgcacttc gtggccgagg
     4201 agcaggactg acacgtgcta aaacttcatt tttaatttaa aaggatctag gtgaagatcc
     4261 tttttgataa tctcatgacc aaaatccctt aacgtgagtt ttcgttccac tgagcgtcag
     4321 accccgtaga aaagatcaaa ggatcttc
//
