LOCUS       sequence-350000-        6831 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Vector to create landing pad 1 (LP1) by homologous recombination in 
            Aspergillus nidulans Delta-St site. The strain A. nidulans LP1 is 
            used for recombinase mediated chromosomal integration..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6831)
  AUTHORS   Roux I, Chooi YH
  TITLE     Cre/lox-Mediated Chromosomal Integration of Biosynthetic Gene 
            Clusters for Heterologous Expression in Aspergillus nidulans.
  JOURNAL   ACS Synth Biol. 2022 Feb 16. doi: 10.1021/acssynbio.1c00458.
  PUBMED    35168324
REFERENCE   2  (bases 1 to 6831)
  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: "ACS Synth 
            Biol. 2022 Feb 16. doi: 10.1021/acssynbio.1c00458."
COMMENT     Sequence Label: pGem-LP1-loxP-bar-lox2272-71
FEATURES             Location/Qualifiers
     source          1..6831
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      complement(126..714)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(206..225)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(885..1676,1677..1745))
                     /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     1508..1527
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(1746..1850)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      complement(1928..2383)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2065..2086)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     2277..2296
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2509..2531
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     2523..2540
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     2524..2540
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     2547..2566
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        2547..2565
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     regulatory      2596..2605
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     regulatory      2600..2609
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     protein_bind    3745..3778
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     CDS             complement(4170..4721)
                     /codon_start=1
                     /gene="Streptomyces hygroscopicus bar"
                     /product="phosphinothricin acetyltransferase"
                     /label=BlpR
                     /note="confers resistance to bialophos or phosphinothricin"
                     /translation="MSPERRPADIRRATEADMPAVCTIVNHYIETSTVNFRTEPQEPQE
                     WTDDLVRLRERYPWLVAEVDGEVAGIAYAGPWKARNAYDWTAESTVYVSPRHQRTGLGS
                     TLYTHLLKSLEAQGFKSVVAVIGLPNDPSVRMHEALGYAPRGMLRAAGFKHGNWHDVGF
                     WQLDFSLPVPPRPVLPVTEI"
     promoter        complement(4744..5103)
                     /label=trpC promoter
                     /note="promoter for Aspergillus nidulans trpC"
     promoter        complement(6519..6537)
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     complement(6520..6537)
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     primer_bind     complement(6555..6571)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(6555..6571)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(6568..6590)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    6579..6595
                     /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(6603..6609,6610..6627,6628..6633))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    6648..6669
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(6786..6803)
                     /label=L4440
                     /note="L4440 vector, forward primer"
ORIGIN
        1 gtatcagctc actcaaaggc ggtaatacgg ttatccacag aatcagggga taacgcagga
       61 aagaacatgt gagcaaaagg ccagcaaaag gccaggaacc gtaaaaaggc cgcgttgctg
      121 gcgtttttcc ataggctccg cccccctgac gagcatcaca aaaatcgacg ctcaagtcag
      181 aggtggcgaa acccgacagg actataaaga taccaggcgt ttccccctgg aagctccctc
      241 gtgcgctctc ctgttccgac cctgccgctt accggatacc tgtccgcctt tctcccttcg
      301 ggaagcgtgg cgctttctca tagctcacgc tgtaggtatc tcagttcggt gtaggtcgtt
      361 cgctccaagc tgggctgtgt gcacgaaccc cccgttcagc ccgaccgctg cgccttatcc
      421 ggtaactatc gtcttgagtc caacccggta agacacgact tatcgccact ggcagcagcc
      481 actggtaaca ggattagcag agcgaggtat gtaggcggtg ctacagagtt cttgaagtgg
      541 tggcctaact acggctacac tagaagaaca gtatttggta tctgcgctct gctgaagcca
      601 gttaccttcg gaaaaagagt tggtagctct tgatccggca aacaaaccac cgctggtagc
      661 ggtggttttt ttgtttgcaa gcagcagatt acgcgcagaa aaaaaggatc tcaagaagat
      721 cctttgatct tttctacggg gtctgacgct cagtggaacg aaaactcacg ttaagggatt
      781 ttggtcatga gattatcaaa aaggatcttc acctagatcc ttttaaatta aaaatgaagt
      841 tttaaatcaa tctaaagtat atatgagtaa acttggtctg acagttacca atgcttaatc
      901 agtgaggcac ctatctcagc gatctgtcta tttcgttcat ccatagttgc ctgactcccc
      961 gtcgtgtaga taactacgat acgggagggc ttaccatctg gccccagtgc tgcaatgata
     1021 ccgcgagacc cacgctcacc ggctccagat ttatcagcaa taaaccagcc agccggaagg
     1081 gccgagcgca gaagtggtcc tgcaacttta tccgcctcca tccagtctat taattgttgc
     1141 cgggaagcta gagtaagtag ttcgccagtt aatagtttgc gcaacgttgt tgccattgct
     1201 acaggcatcg tggtgtcacg ctcgtcgttt ggtatggctt cattcagctc cggttcccaa
     1261 cgatcaaggc gagttacatg atcccccatg ttgtgcaaaa aagcggttag ctccttcggt
     1321 cctccgatcg ttgtcagaag taagttggcc gcagtgttat cactcatggt tatggcagca
     1381 ctgcataatt ctcttactgt catgccatcc gtaagatgct tttctgtgac tggtgagtac
     1441 tcaaccaagt cattctgaga atagtgtatg cggcgaccga gttgctcttg cccggcgtca
     1501 atacgggata ataccgcgcc acatagcaga actttaaaag tgctcatcat tggaaaacgt
     1561 tcttcggggc gaaaactctc aaggatctta ccgctgttga gatccagttc gatgtaaccc
     1621 actcgtgcac ccaactgatc ttcagcatct tttactttca ccagcgtttc tgggtgagca
     1681 aaaacaggaa ggcaaaatgc cgcaaaaaag ggaataaggg cgacacggaa atgttgaata
     1741 ctcatactct tcctttttca atattattga agcatttatc agggttattg tctcatgagc
     1801 ggatacatat ttgaatgtat ttagaaaaat aaacaaatag gggttccgcg cacatttccc
     1861 cgaaaagtgc cacctgatgc ggtgtgaaat accgcacaga tgcgtaagga gaaaataccg
     1921 catcaggaaa ttgtaagcgt taatattttg ttaaaattcg cgttaaattt ttgttaaatc
     1981 agctcatttt ttaaccaata ggccgaaatc ggcaaaatcc cttataaatc aaaagaatag
     2041 accgagatag ggttgagtgt tgttccagtt tggaacaaga gtccactatt aaagaacgtg
     2101 gactccaacg tcaaagggcg aaaaaccgtc tatcagggcg atggcccact acgtgaacca
     2161 tcaccctaat caagtttttt ggggtcgagg tgccgtaaag cactaaatcg gaaccctaaa
     2221 gggagccccc gatttagagc ttgacgggga aagccggcga acgtggcgag aaaggaaggg
     2281 aagaaagcga aaggagcggg cgctagggcg ctggcaagtg tagcggtcac gctgcgcgta
     2341 accaccacac ccgccgcgct taatgcgccg ctacagggcg cgtccattcg ccattcaggc
     2401 tgcgcaactg ttgggaaggg cgatcggtgc gggcctcttc gctattacgc cagctggcga
     2461 aagggggatg tgctgcaagg cgattaagtt gggtaacgcc agggttttcc cagtcacgac
     2521 gttgtaaaac gacggccagt gaattgtaat acgactcact atagggcgaa ttgggcccga
     2581 cgtcgcatgc tcccggccgc catggcggcc gcgggaattc acgtcgtcgt cgcccagcat
     2641 tggacagcgc gttaatgggt cgatagacac tcatttgtat gcactgacgc gttaagggat
     2701 ggcttgaaac ttgcatgaag cgagtgtggt atcagtgtgc agaattcgtc cggttggtgt
     2761 gcgagtcccc cagttcagca aaaacacaca accaacctcc gatcaggggc cggtcgcgct
     2821 cttagaattt cctggtagta tctgataata gatgcttgtc ttggtaaaca taaacagggt
     2881 atcggggctt gaactggaat tggctgtcga ttcaaaattg tggctgcgag ccattacgtc
     2941 acagcgcctg accaaggcat aactgtggca gacagtgcat ccactttcta agtctacacc
     3001 atagggcaga aaggatggta ttgattcttg cttcatttgc ctagattata tcctagacaa
     3061 actttttggt agatagacag gccatcagaa agaacgagca aagggtaaac gattggccta
     3121 ttcttctccc tggaataaag atgattttcc tatgtcatta acttgaactt caccaggatc
     3181 ctcacagcaa ctagcccgtt cggtgactat cttagggctc ccttacatcc ttgtgaaatg
     3241 tggatgtgcg ggtcaagtat tgcgatgact cgtcattgga ctgaaatgga ctaccggtct
     3301 tatatgagct cgtcattgcc caggagcatt cactggccgg ttgttagcgt gatgcttatt
     3361 ttggctgatt tgcgaattga ctttcgatcc aaaggctctt acgtgcgatg aaccgcctcc
     3421 aaagtgagaa caatacgtag tcaccagatg agttgaggac attgtgaact aagccagcca
     3481 aacccagaca aggcgtaaga tagctgctgc tttcgatata tacactaaga aagctggtta
     3541 tacctgggac tgggaccaga gcatccctgc ttctagccca ctgacagccc aacatctgac
     3601 tgcggtaata cggcatttat gaccagggcc ccaaccaagg atattcccct accgacgacc
     3661 caacctttgg gtctgaagag gagtttacgt ccagccaagc tagcttggct gcaggtcgag
     3721 cggccgcgat ccggaaccct taatataact tcgtataatg tatgctatac gaagttatta
     3781 ggtccctcga ccggccgcgg cggttgtttg caagaccgag aaaaggctag caagaatcgg
     3841 gtcattgtag cgtatgcgcc tgtgaacatt ctcttcaaca agtttgattc cattgcggtg
     3901 aaatggtaaa agtcaacccc ctgcgatgta tattttcctg tacaatcaat caaaaagcca
     3961 aatgatttag cattatcttt acatcttgtt attttacaga ttttatgttt agatccacta
     4021 gtttctcggt actatgcatt acatgttaat tattacatgc ttaacgtaat tcaacagaaa
     4081 ttatatgata atcatcgcaa gaccggcaac aggattcaat cttaagaaac tttattgcca
     4141 aatgtttgaa cgatctgcag gtcgacggat cagatctcgg tgacgggcag gaccggacgg
     4201 ggcggtaccg gcaggctgaa gtccagctgc cagaaaccca cgtcatgcca gttcccgtgc
     4261 ttgaagccgg ccgcccgcag catgccgcgg ggggcatatc cgagcgcctc gtgcatgcgc
     4321 acgctcgggt cgttgggcag cccgatgaca gcgaccacgc tcttgaagcc ctgtgcctcc
     4381 agggacttca gcaggtgggt gtagagcgtg gagcccagtc ccgtccgctg gtggcggggg
     4441 gagacgtaca cggtcgactc ggccgtccag tcgtaggcgt tgcgtgcctt ccaggggccc
     4501 gcgtaggcga tgccggcgac ctcgccgtcc acctcggcga cgagccaggg atagcgctcc
     4561 cgcagacgga cgaggtcgtc cgtccactcc tgcggttcct gcggctcggt acggaagttg
     4621 accgtgcttg tctcgatgta gtggttgacg atggtgcaga ccgccggcat gtccgcctcg
     4681 gtggcacggc ggatgtcggc cgggcgtcgt tctgggctca tggttacttc ctaatcgaag
     4741 ctttgcttgg gtagaatagg taagtcagat tgaatctgaa ataaagggag gaagggcgaa
     4801 cttaagaagg tatgaccggg tcgttcactt accttgcttg acaaacgcac caagttatcg
     4861 tgcaccaagc agcagatgat aataatgtcc tcgttcctgt ctgctaataa gagtcacact
     4921 tcgagcgccg ccgctactgc tacaagtggg gctgatctga ccagttgcct aaatgaacca
     4981 tcttgtcaaa cgacacaaat tttgtgctca ccgcctggac gactaaacca aaataggcat
     5041 tcattgttga cctccactag ctccagccaa gcccaaaaag tgctccttca atatcatctt
     5101 ctgtcgagtc tagagcacat tctgcggcct ctgtgctctg caagccgcaa actttcacca
     5161 atggaccaga actacctgtg aaattaataa cagacatact ccaagctgcc tttgtgtgct
     5221 taatcacgta tactcacatt atagatcata acttcgtata aagtatccta tacgaacggt
     5281 agctagctgt acgtcaggtg gcacttttcg gggaaatgtg cgcggaaccc ctatttgttt
     5341 atttttctaa atacattcaa atatgtatcc gctcatgaga caataaccct gtcccacctt
     5401 gatctacctg ctgtcattta gtaagctacg atgacacgta cccaatacag gacctcgaag
     5461 cattgcgtcg tttgccgttg ctgatcgtga actgcacttt ttgtagaact cagacaacgt
     5521 tcatcgctat ttactggacc atgaaaatca aacaaaaaat aactaatcca tatacgtctg
     5581 aaataccgag actatcgttt gtaagtgggg aattaatcgt gctgtgccag gacactgtct
     5641 gaccaaacta ggagaacttg ggcttctcaa tgagttcctg cgtctcttgg ccaagcctgt
     5701 tccggcgcgt tactgctcct gtccgttata aaatggttca ggctcggtat gcaatgaagg
     5761 cattggtttg ctgtggtagt aagtagtacg cggaacctgc atccgcgact gttgactgta
     5821 ttatgacggt atataccttt ctagatacgc tctgtcatgg catttttcgt tgttatgtta
     5881 tatttcaaca ggtatattta ggactatatg gtttgactag agcatctata accgcaagcc
     5941 agttttatcg gtgcgatcca ggcaggtcga gaatgggatc tgatcaaagt gctgagagat
     6001 gcttgcccga aaacgtctct tacaggatac ccaagatttg gcgttgcatt agtctaacta
     6061 tgcttgtcct attaatacct tctcatatta tcggtatcct tacctggtta acaggctgat
     6121 gagatgccag gtacaatacc attttctaaa cattaacagt gacatgacca atcacttagc
     6181 tagtttcatc tatttgccga cggtttcgca ttctgggcat aggccagaaa aaggcaacag
     6241 caggaagaca ttaaaagaga tccttaaagt atcaaaagtc atgctaggcg cgctgtatta
     6301 tatttaatac ggtcatggac aacgtaaatg cctcgctggc agaggcagac accaaaccat
     6361 tgcaacataa gattgcagag atatataatt atatctatgt aactttccag caatcctaat
     6421 cattacctca cttgggcgac acgacgagga attcgcggcc gcctgcaggt cgaccatatg
     6481 ggagagctcc caacgcgttg gatgcatagc ttgagtattc tatagtgtca cctaaatagc
     6541 ttggcgtaat catggtcata gctgtttcct gtgtgaaatt gttatccgct cacaattcca
     6601 cacaacatac gagccggaag cataaagtgt aaagcctggg gtgcctaatg agtgagctaa
     6661 ctcacattaa ttgcgttgcg ctcactgccc gctttccagt cgggaaacct gtcgtgccag
     6721 ctgcattaat gaatcggcca acgcgcgggg agaggcggtt tgcgtattgg gcgctcttcc
     6781 gcttcctcgc tcactgactc gctgcgctcg gtcgttcggc tgcggcgagc g
//
