LOCUS       sequence-343842-        5695 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Plasmids for gene knockout were constructed based on suicide plasmid
            pHFX-B60.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5695)
  AUTHORS   Lin L, Chen J, Mitra R, Gao Q, Cheng F, Xu T, Zuo Z, Xiang H, Han J
  TITLE     Optimising PHBV biopolymer production in haloarchaea via 
            CRISPRi-mediated redirection of carbon flux.
  JOURNAL   Commun Biol. 2021 Aug 25;4(1):1007. doi: 10.1038/s42003-021-02541-z.
  PUBMED    34433872
REFERENCE   2  (bases 1 to 5695)
  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: "2021-08-25"; volume: "4(1):1007. doi"; pages: " 
            10.1038/s42003-021-02541-z"
COMMENT     Sequence Label: pHFX-B60-EPSUD
FEATURES             Location/Qualifiers
     source          1..5695
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             complement(join(515..1306,1307..1375))
                     /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     1138..1157
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(1376..1480)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      complement(1506..1961)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1643..1664)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     1855..1874
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2088..2110
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     2102..2119
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     2103..2119
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     promoter        2133..2151
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(5006..5025)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(5007..5025)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(5049..5065)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5049..5065)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(5062..5084)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5073..5089
                     /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)."
     promoter        complement(join(5097..5103,5104..5121,5122..5127))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5142..5163
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5280..5297)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(join(5451..5695,1..344))
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5531..5550)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 tgggctgtgt gcacgaaccc cccgttcagc ccgaccgctg cgccttatcc ggtaactatc
       61 gtcttgagtc caacccggta agacacgact tatcgccact ggcagcagcc actggtaaca
      121 ggattagcag agcgaggtat gtaggcggtg ctacagagtt cttgaagtgg tggcctaact
      181 acggctacac tagaagaaca gtatttggta tctgcgctct gctgaagcca gttaccttcg
      241 gaaaaagagt tggtagctct tgatccggca aacaaaccac cgctggtagc ggtggttttt
      301 ttgtttgcaa gcagcagatt acgcgcagaa aaaaaggatc tcaagaagat cctttgatct
      361 tttctacggg gtctgacgct cagtggaacg aaaactcacg ttaagggatt ttggtcatga
      421 gattatcaaa aaggatcttc acctagatcc ttttaaatta aaaatgaagt tttaaatcaa
      481 tctaaagtat atatgagtaa acttggtctg acagttacca atgcttaatc agtgaggcac
      541 ctatctcagc gatctgtcta tttcgttcat ccatagttgc ctgactcccc gtcgtgtaga
      601 taactacgat acgggagggc ttaccatctg gccccagtgc tgcaatgata ccgcgagacc
      661 cacgctcacc ggctccagat ttatcagcaa taaaccagcc agccggaagg gccgagcgca
      721 gaagtggtcc tgcaacttta tccgcctcca tccagtctat taattgttgc cgggaagcta
      781 gagtaagtag ttcgccagtt aatagtttgc gcaacgttgt tgccattgct acaggcatcg
      841 tggtgtcacg ctcgtcgttt ggtatggctt cattcagctc cggttcccaa cgatcaaggc
      901 gagttacatg atcccccatg ttgtgcaaaa aagcggttag ctccttcggt cctccgatcg
      961 ttgtcagaag taagttggcc gcagtgttat cactcatggt tatggcagca ctgcataatt
     1021 ctcttactgt catgccatcc gtaagatgct tttctgtgac tggtgagtac tcaaccaagt
     1081 cattctgaga atagtgtatg cggcgaccga gttgctcttg cccggcgtca atacgggata
     1141 ataccgcgcc acatagcaga actttaaaag tgctcatcat tggaaaacgt tcttcggggc
     1201 gaaaactctc aaggatctta ccgctgttga gatccagttc gatgtaaccc actcgtgcac
     1261 ccaactgatc ttcagcatct tttactttca ccagcgtttc tgggtgagca aaaacaggaa
     1321 ggcaaaatgc cgcaaaaaag ggaataaggg cgacacggaa atgttgaata ctcatactct
     1381 tcctttttca atattattga agcatttatc agggttattg tctcatgagc ggatacatat
     1441 ttgaatgtat ttagaaaaat aaacaaatag gggttccgcg cacatttccc cgaaaagtgc
     1501 cacctaaatt gtaagcgtta atattttgtt aaaattcgcg ttaaattttt gttaaatcag
     1561 ctcatttttt aaccaatagg ccgaaatcgg caaaatccct tataaatcaa aagaatagac
     1621 cgagataggg ttgagtgttg ttccagtttg gaacaagagt ccactattaa agaacgtgga
     1681 ctccaacgtc aaagggcgaa aaaccgtcta tcagggcgat ggcccactac gtgaaccatc
     1741 accctaatca agttttttgg ggtcgaggtg ccgtaaagca ctaaatcgga accctaaagg
     1801 gagcccccga tttagagctt gacggggaaa gccggcgaac gtggcgagaa aggaagggaa
     1861 gaaagcgaaa ggagcgggcg ctagggcgct ggcaagtgta gcggtcacgc tgcgcgtaac
     1921 caccacaccc gccgcgctta atgcgccgct acagggcgcg tcccattcgc cattcaggct
     1981 gcgcaactgt tgggaagggc gatcggtgcg ggcctcttcg ctattacgcc agctggcgaa
     2041 agggggatgt gctgcaaggc gattaagttg ggtaacgcca gggttttccc agtcacgacg
     2101 ttgtaaaacg acggccagtg aattaggcct gtaattaacc ctcactaaag ggagagaatt
     2161 cgttttcttg tgtgcggttc ccttgcgtac agttcccagc gaccttcccg gctgcgacga
     2221 agggaaaacc ccttacatac ggtcgtcgca ctcgcgggca atgggcttct ttgacgacct
     2281 tcaggaccgc atcgagaagg tggacagtgt cgtctcggtc ggactcgacg ccgacctcga
     2341 ccggattccc gaacacctcc gcgacgagga cctgccgcgg tgggcgttca accgccgcat
     2401 catcgacgcg acgcacgaac acgcggcggc ctacaagccg aacgcggcct tctacgagga
     2461 cgaagacggc tggcgctcgc tccgcgagac catcgcctac gcacacggaa aggatgttcc
     2521 cgtcctcctc gacgccaagc gcgccgacat cggcaacacg acgcgacagt acgcgaacct
     2581 gcttgacgac gccgatgcca tcaccgtcaa cccctacatg ggccgcgatt ccatccagcc
     2641 gttcctcgcc cgcgaggaca agggcgtgtt cgtcctctgt cgcacctcta acccgggcag
     2701 cgcggacctc cagtcgctcg aactcgcctc cggagaaccc ctgtacgagc gcgtcgcggc
     2761 gctctgcgac ctctggaacg aaaacgacaa cgtcggactc gtcgtcggcg cgaccggccc
     2821 cgaagagttg gaagacctcc gcgagcaggt ccccgacctg ccgtttctcg tccccggcgt
     2881 cggcgcacag ggtggcgatg ccgaggcggc agtcgagtac ggactggcag acggcgtcgg
     2941 tctcgtgaac tcctcacgcg gcatcatctt cgccggcgaa gaccagggag aacacttcga
     3001 caaggctgcg ggacaggcgg cgaagcgact caaagagcgg ctgaaccagt accggtaagg
     3061 taccatctaa tggtgtcgaa gggaacatat atgttactgc aggtacaaca ccgagttagg
     3121 agatgggatc catgcactcc gataacatcc aaaccagcaa atcgatacag caggagaccg
     3181 gccggacgtt tcacctcgcg acgcgtctgc ttccggagcg tatccgccac ccgacctacg
     3241 tcatgtacgc ctttttccgg gtcgcggacg aggtcgtcga ccagacggat gggccgcccc
     3301 cgaccgtcca gcacgagcaa ctggaggtaa tccgcgaggc cgcgctcggg aacgtcgacc
     3361 cggccgagac cgaccacgag gcggtcatgg cggcgtttca ggacctggcc gagcgtcacg
     3421 acatctccga ggagacgatc aacgtcttca tcgacgcgat ggagatggac atcgcacagg
     3481 cccgctacga gacgttcgag gacctccgtg agtacatggg cggctccgcc gtcgctgtcg
     3541 gccacatgat gacggaggtg atggacccgc cacagaaggc ggaggccctg ccccacgcga
     3601 cggcgctggc cgaagcgttc cagctatcga acttcctgcg ggacgtccgc gaggacatcc
     3661 acgactacgg ccgggtgtac ctcccacagg agacgctcga ccgccacggc gtcaccgagg
     3721 agcaactggc cgacgccgag gtcgacgacg ccttccgtgc cgtgatgcag gaggaactgg
     3781 ctcggaccga cgaactgtac cgcgagggcg tcgccggcat ccgatacctc ccggaagact
     3841 gccagttcgg cgtgctgctg gctgcggtcc tgtacgctga ccaccaccga ctcatccgcg
     3901 accgtggcta cgacgtgctg acggagacgc cggaactcac tcgtcgtcgc cggctgtggt
     3961 tgctcgcccg tacgtggtgg cactggcggc gcaacggcga ccccgaagcg acgttctaca
     4021 ccgtgagcgc tgtctccgag cgcggccccg gcgagacgcc gacggacgcc cacggccacg
     4081 ggcaacccgc gtggcgtgga tgagatatcg agctcagtgg ttctggattc gctcgcgttt
     4141 gtggccgacc acgatgatga tctcgtcaac actggcagct ttgattgctt cgacgatgtg
     4201 ctccagcaat ggtttgtctc caattgggag catcgctttg ggtcgtgctt gggtaagtgg
     4261 gcgaagccgg gttccctcac cagcagcgag aatgattgct ttcattgtaa tatttttttg
     4321 cgtgtgtcca tttcattttt acgtcacaga ttttttctag gtgagctaaa tttaattatg
     4381 ccactctcat cctcacttac tctcctgttt attcattaat aaatagccgt caattttatg
     4441 ttttggttcg agtatggaca tagtgagaaa gaaaatatca agttcacaga gtgtcgcaat
     4501 cacggctaca ataacacttg taataattcc agtgtgtcgt gcctacctgt cactagtaca
     4561 atactaaact atcccattga aagggtctgt accgaaaacc attttgtcta gtcagacggc
     4621 cattaaatat ggattgggag aaatttgcta ttacactatc gactgtatgt ctttccctta
     4681 cagttgtata tgcgagtttc aatccgtcat caaagtacag cctaagtatt tacgccgaga
     4741 acccgctttt ttggttgggt gttactattg caggtggact tagtatcgtt acgatcgttc
     4801 taggctcgcg tagaagatta tcaattgggg tcctatacct ggttggagta attgttttat
     4861 ttgcaccgac cctcctaggg ttctatttcc atgctggagg tgatggactc actcagttag
     4921 gttggttgaa agcaataaaa gatggtgagt tctccatctt tggacttcga tatatgggtc
     4981 tacaattgat tgcatgcaag cttctcccta tagtgagtcg tattaatttg atatctggcg
     5041 taatcatggt catagctgtt tcctgtgtga aattgttatc cgctcacaat tccacacaac
     5101 atacgagccg gaagcataaa gtgtaaagcc tggggtgcct aatgagtgag ctaactcaca
     5161 ttaattgcgt tgcgctcact gcccgctttc cagtcgggaa acctgtcgtg ccagctgcat
     5221 taatgaatcg gccaacgcgc ggggagaggc ggtttgcgta ttgggcgctc ttccgcttcc
     5281 tcgctcactg actcgctgcg ctcggtcgtt cggctgcggc gagcggtatc agctcactca
     5341 aaggcggtaa tacggttatc cacagaatca ggggataacg caggaaagaa catgtgagca
     5401 aaaggccagc aaaaggccag gaaccgtaaa aaggccgcgt tgctggcgtt tttccatagg
     5461 ctccgccccc ctgacgagca tcacaaaaat cgacgctcaa gtcagaggtg gcgaaacccg
     5521 acaggactat aaagatacca ggcgtttccc cctggaagct ccctcgtgcg ctctcctgtt
     5581 ccgaccctgc cgcttaccgg atacctgtcc gcctttctcc cttcgggaag cgtggcgctt
     5641 tctcatagct cacgctgtag gtatctcagt tcggtgtagg tcgttcgctc caagc
//
