LOCUS       sequence-444249-        6098 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  For expression of the type I CBASS system encoded by Methylosarcina 
            fibrata.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6098)
  AUTHORS   Richmond-Buccola D, Hobbs SJ, Garcia JM, Toyoda H, Gao J, Shao S, 
            Lee ASY, Kranzusch PJ
  TITLE     A large-scale type I CBASS antiphage screen identifies the phage 
            prohead protease as a key determinant of immune activation and 
            evasion.
  JOURNAL   Cell Host Microbe. 2024 Jul 10;32(7):1074-1088.e5. doi: 
            10.1016/j.chom.2024.05.021. Epub 2024 Jun 24.
  PUBMED    38917809
REFERENCE   2  (bases 1 to 6098)
  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"; doi: 
            "10.1016/j.chom.2024.05.021"; journalName: "Cell Host Microbe"; 
            date: "2024-07-10- 10"; volume: "32"; issue: "7"; pages: 
            "1074-1088.e5"
COMMENT     Sequence Label: pBAD02_CBASS_Methylosarcina_fibrata
FEATURES             Location/Qualifiers
     source          1..6098
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     140..162
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(318..1196)
                     /codon_start=1
                     /gene="araC"
                     /product="L-arabinose regulatory protein"
                     /label=araC
                     /translation="MAEAQNDPLLPGYSFNAHLVAGLTPIEANGYLDFFIDRPLGMKGY
                     ILNLTIRGQGVVKNQGREFVCRPGDILLFPPGEIHHYGRHPEAREWYHQWVYFRPRAYW
                     HEWLNWPSIFANTGFFRPDEAHQPHFSDLFGQIINAGQGEGRYSELLAINLLEQLLLRR
                     MEAINESLHPPMDNRVREACQYISDHLADSNFDIASVAQHVCLSPSRLSHLFRQQLGIS
                     VLSWREDQRISQAKLLLSTTRMPIATVGRNVGFDDQLYFSRVFKKCTGASPSEFRAGCE
                     EKVNDVAVKLS"
     promoter        1223..1507
                     /gene="araBAD"
                     /label=araBAD promoter
                     /note="promoter of the L-arabinose operon of E. coli; the 
                     araC regulatory gene is transcribed in the opposite 
                     direction (Guzman et al., 1995)"
     primer_bind     1430..1449
                     /label=pBAD Forward
                     /note="For vectors with E. coli araBAD promoter, forward 
                     primer"
     terminator      3937..4008
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     terminator      4069..4096
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB 
                     gene"
     promoter        4115..4206
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(4207..4275,4276..5067)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRVDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     complement(4425..4444)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      5238..5826
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5727..5746
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     5980..5997
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    join(6012..6098,1..54)
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(6078..6097)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
ORIGIN
        1 atagttaagc cagtatacac tccgctatcg ctacgtgact gggtcatggc tgcgccccga
       61 cacccgccaa cacccgctga cgcgccctga cgggcttgtc tgctcccggc atccgcttac
      121 agacaagctg tgaccgtctc cgggagctgc atgtgtcaga ggttttcacc gtcatcaccg
      181 aaacgcgcga ggcagcagat caattcgcgc gcgaaggcga agcggcatgc ataatgtgcc
      241 tgtcaaatgg acgaagcagg gattctgcaa accctatgct actccgtcaa gccgtcaatt
      301 gtctgattcg ttaccaatta tgacaacttg acggctacat cattcacttt ttcttcacaa
      361 ccggcacgga actcgctcgg gctggccccg gtgcattttt taaatacccg cgagaaatag
      421 agttgatcgt caaaaccaac attgcgaccg acggtggcga taggcatccg ggtggtgctc
      481 aaaagcagct tcgcctggct gatacgttgg tcctcgcgcc agcttaagac gctaatccct
      541 aactgctggc ggaaaagatg tgacagacgc gacggcgaca agcaaacatg ctgtgcgacg
      601 ctggcgatat caaaattgct gtctgccagg tgatcgctga tgtactgaca agcctcgcgt
      661 acccgattat ccatcggtgg atggagcgac tcgttaatcg cttccatgcg ccgcagtaac
      721 aattgctcaa gcagatttat cgccagcagc tccgaatagc gcccttcccc ttgcccggcg
      781 ttaatgattt gcccaaacag gtcgctgaaa tgcggctggt gcgcttcatc cgggcgaaag
      841 aaccccgtat tggcaaatat tgacggccag ttaagccatt catgccagta ggcgcgcgga
      901 cgaaagtaaa cccactggtg ataccattcg cgagcctccg gatgacgacc gtagtgatga
      961 atctctcctg gcgggaacag caaaatatca cccggtcggc aaacaaattc tcgtccctga
     1021 tttttcacca ccccctgacc gcgaatggtg agattgagaa tataaccttt cattcccagc
     1081 ggtcggtcga taaaaaaatc gagataaccg ttggcctcaa tcggcgttaa acccgccacc
     1141 agatgggcat taaacgagta tcccggcagc aggggatcat tttgcgcttc agccatactt
     1201 ttcatactcc cgccattcag agaagaaacc aattgtccat attgcatcag acattgccgt
     1261 cactgcgtct tttactggct cttctcgcta accaaaccgg taaccccgct tattaaaagc
     1321 attctgtaac aaagcgggac caaagccatg acaaaaacgc gtaacaaaag tgtctataat
     1381 cacggcagaa aagtccacat tgattatttg cacggcgtca cactttgcta tgccatagca
     1441 tttttatcca taagattagc ggatcctacc tgacgctttt tatcgcaact ctctactgtt
     1501 tctccatacc cgttttttgg gctaacagga ggaattaacc atgaacgaaa tagaaaaaat
     1561 acagaatcaa cctgaaatga tcaggatagt tcgcgctttt atcgtgatat ataacaggtc
     1621 gaaacaatac caggcaatcg cattgctagt gacgatgctt ttgccaatac tcgggggtat
     1681 agcagcggtc tttaagcctg acctaaagcc ttttattgct attttcgcat tatccttcgg
     1741 atttatagac gctcttctca ttgaccggtc gcataaggtg cggatgaaaa ctgcggcaaa
     1801 actgcaagaa gaattcgact gccacgtcct atccctcgaa tggaataagt ttcttgtcgg
     1861 tagggcggtt gacccggagg atgtagcgcg ggaatcctcc tgcaagctaa gcgcctcaga
     1921 cgaaaggcgg ctaacagact ggtatccaac cgtagctaaa gaactgccta tccatttggc
     1981 aagaatcatt tgccaacgga caaacctatg gtacgacaga gagctaagaa caaggtacag
     2041 aaacatcata tcctggtcga ttgctcttat cgttatagct tcgacggtag tttcttcgac
     2101 cgtaggcttt accgtgactt ccttcgtaat caccattctt gtacccatat ccccggcgct
     2161 aaactgggca atacgagaat actatcgtca aaccgatgcg atgaagactt tggagaacct
     2221 gaaaagcgag atagagaagc tatgggacaa agcgctggga aatgctaccg tagaagaagt
     2281 gaccgaagct tccagggaac tacaggacgc gatctttgct caccgggcgt ccagcccctt
     2341 gatattcaac tggctttaca agcctctgcg gccaaagatg gaatcaaaca tgaacagcgg
     2401 cgctgagtat tgggtaaaac aggccaaaac ggcaaaaggt ataaattgag tctaggaccg
     2461 cattcgaggg caagtaatga agttgataac gcatttcgac gatttcttaa gggataccgt
     2521 caatctgaac caaactagag tgactcttct tgaggatagc atcgaagcaa taaagacatt
     2581 tatcaataat tcggattgga agccagagat tatagagttt aagccccaag ggtcatgggc
     2641 acataaaacc atcataaagc cgttgacgga ttacccattc gatgccgacc ttatcgttta
     2701 cgtcaagcct gtagacggat gggaggccaa ggattacgtc aacaacctct atacggagtt
     2761 ccaagggagc ggaatttata aggagaaaac acggaggtat tctcattgcg tgacaatcga
     2821 atatgcggga gaaaggaaaa tagatatagc cccgtgtgtc aaagacaggc tatattctgg
     2881 cacatacgag gtttgcaaca gaaattcaaa cgaatttgag agatccaagc cgggagaata
     2941 cacggactgg ctcatcgata aaaattctat atctgggaag aataacctca ggaaggtgac
     3001 gcgactttta aagtacatgc gcgacatcaa aacaaacttt atttgccagt cattcctcct
     3061 tacaactctg cttggggatc ggatttccta tgtagacagc atcaacaagc cgtttaccga
     3121 cgttcctacg agcctgaaaa tacttattgg caggttggac gactggttgc aagccaatcc
     3181 gaccaagcca acggtcagaa acccggtact gtacgaggaa atccagagca acgcatggga
     3241 tgaaaccaaa tacagcaatt tcagaaataa gatcaaccta tacagagggt ggatcgatga
     3301 cgcctacgac gaatctgaca aggaagagag cattggaaaa tggaggcgcg tgtttgggga
     3361 tgacttcgca gcggaagaga cggttgctaa agcggcatgt gttagtgaat cggctgccct
     3421 agaattgagt ggcaggtcac tcgcatcggc tggttccgac cttgtcacct tagtgagaag
     3481 tttaggttcc atagctttgc caaagggatt taatcggctt ccacatatgc accgtcccaa
     3541 atggcgtgca agttctagtg ccggagtcgc ggtgaaagtt tctgcgtatt tatatacggg
     3601 tagatatgga cagccaataa ggcagataac aagtttagag ccattgaaaa gtggttactg
     3661 gattaaattt agtgcttcaa gtggttcggg tttgccattt cccaatgagt attatgttat
     3721 gtggagggtg acaaatacgg ataaagcggc ccaaaaggca caggcattga gaggagagtt
     3781 ttacgaaagc gacagcaaca attcgcgttg ggagggttta cagtatcgcg gggtacatat
     3841 tgtggaggct tttctcatta ggaaagcaac tgaggttatg caggggaaaa gcgatccttt
     3901 ttatgtagta attgaataat aagaattccc aggcatcaaa taaaacgaaa ggctcagtcg
     3961 aaagactggg cctttcgttt tatctgttgt ttgtcggtga acgctctcca gctggtacca
     4021 tatggcgggc aggacgcccg ccataaactg ccaggcatca aattaagcag aaggccatcc
     4081 tgacggatgg cctttttgcg tttctacaaa ctcttttgtt tatttttcta aatacattca
     4141 aatatgtatc cgctcatgag acaataaccc tgataaatgc ttcaataata ttgaaaaagg
     4201 aagagtatga gtattcaaca tttccgtgtc gcccttattc ccttttttgc ggcattttgc
     4261 cttcctgttt ttgctcaccc agaaacgctg gtgaaagtaa aagatgctga agatcagttg
     4321 ggtgcacgag tgggttacat cgaactggat ctcaacagcg gtaagatcct tgagagtttt
     4381 cgccccgaag aacgttttcc aatgatgagc acttttaaag ttctgctatg tggcgcggta
     4441 ttatcccgtg ttgacgccgg gcaagagcaa ctcggtcgcc gcatacacta ttctcagaat
     4501 gacttggttg agtactcacc agtcacagaa aagcatctta cggatggcat gacagtaaga
     4561 gaattatgca gtgctgccat aaccatgagt gataacactg cggccaactt acttctgaca
     4621 acgatcggag gaccgaagga gctaaccgct tttttgcaca acatggggga tcatgtaact
     4681 cgccttgatc gttgggaacc ggagctgaat gaagccatac caaacgacga gcgtgacacc
     4741 acgatgcctg tagcaatggc aacaacgttg cgcaaactat taactggcga actacttact
     4801 ctagcttccc ggcaacaatt aatagactgg atggaggcgg ataaagttgc aggaccactt
     4861 ctgcgctcgg cccttccggc tggctggttt attgctgata aatctggagc cggtgagcgt
     4921 gggtctcgcg gtatcattgc agcactgggg ccagatggta agccctcccg tatcgtagtt
     4981 atctacacga cggggagtca ggcaactatg gatgaacgaa atagacagat cgctgagata
     5041 ggtgcctcac tgattaagca ttggtaactg tcagaccaag tttactcata tatactttag
     5101 attgatttaa aacttcattt ttaatttaaa aggatctagg tgaagatcct ttttgataat
     5161 ctcatgacca aaatccctta acgtgagttt tcgttccact gagcgtcaga ccccgtagaa
     5221 aagatcaaag gatcttcttg agatcctttt tttctgcgcg taatctgctg cttgcaaaca
     5281 aaaaaaccac cgctaccagc ggtggtttgt ttgccggatc aagagctacc aactcttttt
     5341 ccgaaggtaa ctggcttcag cagagcgcag ataccaaata ctgtccttct agtgtagccg
     5401 tagttaggcc accacttcaa gaactctgta gcaccgccta catacctcgc tctgctaatc
     5461 ctgttaccag tggctgctgc cagtggcgat aagtcgtgtc ttaccgggtt ggactcaaga
     5521 cgatagttac cggataaggc gcagcggtcg ggctgaacgg ggggttcgtg cacacagccc
     5581 agcttggagc gaacgaccta caccgaactg agatacctac agcgtgagct atgagaaagc
     5641 gccacgcttc ccgaagggag aaaggcggac aggtatccgg taagcggcag ggtcggaaca
     5701 ggagagcgca cgagggagct tccaggggga aacgcctggt atctttatag tcctgtcggg
     5761 tttcgccacc tctgacttga gcgtcgattt ttgtgatgct cgtcaggggg gcggagccta
     5821 tggaaaaacg ccagcaacgc ggccttttta cggttcctgg ccttttgctg gccttttgct
     5881 cacatgttct ttcctgcgtt atcccctgat tctgtggata accgtattac cgcctttgag
     5941 tgagctgata ccgctcgccg cagccgaacg accgagcgca gcgagtcagt gagcgaggaa
     6001 gcggaagagc gcctgatgcg gtattttctc cttacgcatc tgtgcggtat ttcacaccgc
     6061 atatggtgca ctctcagtac aatctgctct gatgccgc
//
