LOCUS       sequence-257171-        4440 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Cas9 R promoter targeting sgRNA3 expression cassette for Zea mays.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4440)
  AUTHORS   Young JK, Gasior SL, Jones S, Wang L, Navarro P, Vickroy B, 
            Barrangou R
  TITLE     The repurposing of type I-E CRISPR-Cascade for gene activation in 
            plants.
  JOURNAL   Commun Biol. 2019 Oct 18;2:383. doi: 10.1038/s42003-019-0637-6. 
            eCollection 2019.
  PUBMED    31646186
REFERENCE   2  (bases 1 to 4440)
  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: "Commun 
            Biol."; date: "2019-10-18"; volume: "2:383. doi"; pages: " 
            10.1038/s42003-019-0637-6. eCollection 2019"
COMMENT     Sequence Label: PV528
FEATURES             Location/Qualifiers
     source          1..4440
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     123..142
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     376..393
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    510..531
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(546..551,552..569,570..576)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    584..600
                     /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     589..611
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     608..624
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     608..624
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     promoter        645..663
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     misc_RNA        complement(1128..1203)
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
     primer_bind     complement(2232..2248)
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2233..2249)
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     primer_bind     complement(2273..2292)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(2274..2292)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(2299..2316)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(2299..2315)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2308..2330)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      2457..2912
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2544..2563)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2754..2775
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        2938..3042
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(3043..3111,3112..3903)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     complement(3261..3280)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      join(4074..4440,1..222)
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
ORIGIN
        1 gaactgagat acctacagcg tgagctatga gaaagcgcca cgcttcccga agggagaaag
       61 gcggacaggt atccggtaag cggcagggtc ggaacaggag agcgcacgag ggagcttcca
      121 gggggaaacg cctggtatct ttatagtcct gtcgggtttc gccacctctg acttgagcgt
      181 cgatttttgt gatgctcgtc aggggggcgg agcctatgga aaaacgccag caacgcggcc
      241 tttttacggt tcctggcctt ttgctggcct tttgctcaca tgttctttcc tgcgttatcc
      301 cctgattctg tggataaccg tattaccgcc tttgagtgag ctgataccgc tcgccgcagc
      361 cgaacgaccg agcgcagcga gtcagtgagc gaggaagcgg aagagcgccc aatacgcaaa
      421 ccgcctctcc ccgcgcgttg gccgattcat taatgcagct ggcacgacag gtttcccgac
      481 tggaaagcgg gcagtgagcg caacgcaatt aatgtgagtt agctcactca ttaggcaccc
      541 caggctttac actttatgct tccggctcgt atgttgtgtg gaattgtgag cggataacaa
      601 tttcacacag gaaacagcta tgaccatgat tacgccaagc tcgaaattaa ccctcactaa
      661 agggaacaaa agctggagct ccaccgcggt ggcggccgct ctagaactag tcagaacaag
      721 tacctcggcg tgatcatcca gtgcctcgtg accgagacca agacctccgt gtccaggcac
      781 atctacttct tctccgctcg cggcaggatc gaccccctcg tgtacctcga cgagttcctc
      841 aggaactcag agcccgtgct caagagggtg aacaggaccg gcaactcctc ctccaacaag
      901 caggagtacc agctcctcaa ggacaacctc gtgaggtcct acaacaaggc cctcaagaag
      961 aacgccccct actccatctt cgccatcaag aacggcccca agtcccacat cggtaggcac
     1021 ctcatgacct ccttcctctc aatgaagggc ctcaccgagc tcaccaacgt ggtgggcaac
     1081 tggtccgaca agagggccct gcaggaattc gcggccgcaa aaaaaaagca ccgactcggt
     1141 gccacttttt caagttgata acggactagc cttattttaa cttgctattt ctagctctaa
     1201 aactcttgct cggtcacttg gacaattcgg tgcttgcggc tcggtttata aggtacgctc
     1261 cactcttgtg attagtcaag gtgggacaaa acgtttgcgt attacccagc gctttgctcc
     1321 acctacggac tgctgtttgg gcttcgtcgc ggtgcttcgt attgggccgt gggccatgtt
     1381 tcttatggcc ctctctgtga cgaagttttt ggcacatacg ccgctatgac tttctctttt
     1441 acttttttgc aaatattctc cgtcaacgcg gcgctgacac tgtgtcctac tgtacataga
     1501 aggaaaaaag gtatataaaa aaataatttc taaacacgac ttactgctag tagaaattcg
     1561 aatagcattt tagcttttac aactgggcct ggaaattctt tggacaccac tattttacga
     1621 cttatcatca tcgtgtgtta gagcacaagg aactatacaa gcgaggttgc ttatttaagc
     1681 gggatcatta tgtcaaggat tcgatggggc taacaaaaac agctggaaac acagccagat
     1741 ctttgatgcc ccctcgctca aaacttgcat tccttgtaca tgcgctaaca aacagaagaa
     1801 atcccttttt acaatacaat tttactttcc tattcaccct gccatatatc atacaaagtt
     1861 ggccaggaca tgcgcccatg gccaccatct ctaaccctaa tggtgccatc aaactggcac
     1921 catgatgccg gttatctcac taccttgctg caatactgac actctgacag catgccaatt
     1981 tagctcagcg attactggta tccttcaggg cctgttcggt tgtaccggaa taagccagga
     2041 ttcattccag ctcatcaaaa tctatataaa ttagagaagt aatccggcta ggaattaatc
     2101 cggggctcta atccggaaca accgaacagg gcctcaagca atcttgattt tctggatcat
     2161 agactgaggg tgcatttttc agactttggc attgattaat ctaggttcat cattgtactc
     2221 tcaaagctta tcgataccgt cgacctcgag ggggggcccg gtacccaatt cgccctatag
     2281 tgagtcgtat tacaattcac tggccgtcgt tttacaacgt cgtgactggg aaaaccctgg
     2341 cgttacccaa cttaatcgcc ttgcagcaca tccccctttc gccagctggc gtaatagcga
     2401 agaggcccgc accgatcgcc cttcccaaca gttgcgcagc ctgaatggcg aatgggacgc
     2461 gccctgtagc ggcgcattaa gcgcggcggg tgtggtggtt acgcgcagcg tgaccgctac
     2521 acttgccagc gccctagcgc ccgctccttt cgctttcttc ccttcctttc tcgccacgtt
     2581 cgccggcttt ccccgtcaag ctctaaatcg ggggctccct ttagggttcc gatttagtgc
     2641 tttacggcac ctcgacccca aaaaacttga ttagggtgat ggttcacgta gtgggccatc
     2701 gccctgatag acggtttttc gccctttgac gttggagtcc acgttcttta atagtggact
     2761 cttgttccaa actggaacaa cactcaaccc tatctcggtc tattcttttg atttataagg
     2821 gattttgccg atttcggcct attggttaaa aaatgagctg atttaacaaa aatttaacgc
     2881 gaattttaac aaaatattaa cgcttacaat ttaggtggca cttttcgggg aaatgtgcgc
     2941 ggaaccccta tttgtttatt tttctaaata cattcaaata tgtatccgct catgagacaa
     3001 taaccctgat aaatgcttca ataatattga aaaaggaaga gtatgagtat tcaacatttc
     3061 cgtgtcgccc ttattccctt ttttgcggca ttttgccttc ctgtttttgc tcacccagaa
     3121 acgctggtga aagtaaaaga tgctgaagat cagttgggtg cacgagtggg ttacatcgaa
     3181 ctggatctca acagcggtaa gatccttgag agttttcgcc ccgaagaacg ttttccaatg
     3241 atgagcactt ttaaagttct gctatgtggc gcggtattat cccgtattga cgccgggcaa
     3301 gagcaactcg gtcgccgcat acactattct cagaatgact tggttgagta ctcaccagtc
     3361 acagaaaagc atcttacgga tggcatgaca gtaagagaat tatgcagtgc tgccataacc
     3421 atgagtgata acactgcggc caacttactt ctgacaacga tcggaggacc gaaggagcta
     3481 accgcttttt tgcacaacat gggggatcat gtaactcgcc ttgatcgttg ggaaccggag
     3541 ctgaatgaag ccataccaaa cgacgagcgt gacaccacga tgcctgtagc aatggcaaca
     3601 acgttgcgca aactattaac tggcgaacta cttactctag cttcccggca acaattaata
     3661 gactggatgg aggcggataa agttgcagga ccacttctgc gctcggccct tccggctggc
     3721 tggtttattg ctgataaatc tggagccggt gagcgtgggt ctcgcggtat cattgcagca
     3781 ctggggccag atggtaagcc ctcccgtatc gtagttatct acacgacggg gagtcaggca
     3841 actatggatg aacgaaatag acagatcgct gagataggtg cctcactgat taagcattgg
     3901 taactgtcag accaagttta ctcatatata ctttagattg atttaaaact tcatttttaa
     3961 tttaaaagga tctaggtgaa gatccttttt gataatctca tgaccaaaat cccttaacgt
     4021 gagttttcgt tccactgagc gtcagacccc gtagaaaaga tcaaaggatc ttcttgagat
     4081 cctttttttc tgcgcgtaat ctgctgcttg caaacaaaaa aaccaccgct accagcggtg
     4141 gtttgtttgc cggatcaaga gctaccaact ctttttccga aggtaactgg cttcagcaga
     4201 gcgcagatac caaatactgt tcttctagtg tagccgtagt taggccacca cttcaagaac
     4261 tctgtagcac cgcctacata cctcgctctg ctaatcctgt taccagtggc tgctgccagt
     4321 ggcgataagt cgtgtcttac cgggttggac tcaagacgat agttaccgga taaggcgcag
     4381 cggtcgggct gaacgggggg ttcgtgcaca cagcccagct tggagcgaac gacctacacc
//
