LOCUS       sequence-181151-        3969 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Linker to construct BB2 from BB1(FS1 - FS4) with FSE and FSF (in 
            with BbsI, out with BsaI).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 3969)
  AUTHORS   Sarkari P, Marx H, Blumhoff ML, Mattanovich D, Sauer M, Steiger MG
  TITLE     An efficient tool for metabolic pathway construction and gene 
            integration for Aspergillus niger.
  JOURNAL   Bioresour Technol. 2017 May 4. pii: S0960-8524(17)30643-0. doi: 
            10.1016/j.biortech.2017.05.004.
  PUBMED    28533066
REFERENCE   2  (bases 1 to 3969)
  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: "Bioresour 
            Technol. 2017 May 4. pii: S0960-8524(17)30643-0. doi: 
            10.1016/j.biortech.2017.05.004."
COMMENT     Sequence Label: BB2_L_EF_syn_BbsI
FEATURES             Location/Qualifiers
     source          1..3969
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     10..27
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    144..165
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(180..185,186..203,204..210)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    218..234
                     /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     223..245
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     242..258
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     242..258
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     277..296
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        277..295
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     promoter        complement(555..573)
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     complement(556..573)
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     primer_bind     complement(587..604)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(587..603)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(596..618)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      complement(744..1199)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(881..902)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     1093..1112
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        1226..1330
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(1331..1399,1400..2191)
                     /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(1549..1568)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     primer_bind     complement(2326..2345)
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     CDS             2337..3152
                     /codon_start=1
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     CDS             2337..3152
                     /codon_start=1
                     /gene="aph(3')-Ia"
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     primer_bind     complement(2410..2429)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     rep_origin      3237..3825
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3726..3745
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 accgagcgca gcgagtcagt gagcgaggaa gcggaagagc gcccaatacg caaaccgcct
       61 ctccccgcgc gttggccgat tcattaatgc agctggcacg acaggtttcc cgactggaaa
      121 gcgggcagtg agcgcaacgc aattaatgtg agttagctca ctcattaggc accccaggct
      181 ttacacttta tgcttccggc tcgtatgttg tgtggaattg tgagcggata acaatttcac
      241 acaggaaaca gctatgacca tgattacgcc aagctctaat acgactcact atagggaaag
      301 ctcggtacca cgcatgctgc agacgcgtta cgtatcggat ccagaattcg tgatgatcac
      361 cggtggtctc caagcagcgg cgtcttcctc gagggatccg aagacgcctc cctccggaga
      421 ccgcgaaaaa accccgccct gtcaggggcg gggttttttg cgactagtga tcatctgaat
      481 tcgtcgacaa gcttctcgag cctaggctag ctctagacca cacgtgtggg ggcccgagct
      541 cgcggccgct gtattctata gtgtcaccta aatggccgca caattcactg gccgtcgttt
      601 tacaacgtcg tgactgggaa aaccctggcg ttacccaact taatcgcctt gcagcacatc
      661 cccctttcgc cagctggcgt aatagcgaag aggcccgcac cgatcgccct tcccaacagt
      721 tgcgcagcct gaatggcgaa tggaaattgt aagcgttaat attttgttaa aattcgcgtt
      781 aaatttttgt taaatcagct cattttttaa ccaataggcc gaaatcggca aaatccctta
      841 taaatcaaaa gaatagaccg agatagggtt gagtgttgtt ccagtttgga acaagagtcc
      901 actattaaag aacgtggact ccaacgtcaa agggcgaaaa accgtctatc agggcgatgg
      961 cccactacgt gaaccatcac cctaatcaag ttttttgggg tcgaggtgcc gtaaagcact
     1021 aaatcggaac cctaaaggga gcccccgatt tagagcttga cggggaaagc cggcgaacgt
     1081 ggcgagaaag gaagggaaga aagcgaaagg agcgggcgct agggcgctgg caagtgtagc
     1141 ggtcacgctg cgcgtaacca ccacacccgc cgcgcttaat gcgccgctac agggcgcgtc
     1201 aggtggcact tttcggggaa atgtgcgcgg aacccctatt tgtttatttt tctaaataca
     1261 ttcaaatatg tatccgctca tgagacaata accctgataa atgcttcaat aatattgaaa
     1321 aaggaagagt atgagtattc aacatttccg tgtcgccctt attccctttt ttgcggcatt
     1381 ttgccttcct gtttttgctc acccagaaac gctggtgaaa gtaaaagatg ctgaagatca
     1441 gttgggtgca cgagtgggtt acatcgaact ggatctcaac agcggtaaga tccttgagag
     1501 ttttcgcccc gaagaacgtt ttccaatgat gagcactttt aaagttctgc tatgtggcgc
     1561 ggtattatcc cgtattgacg ccgggcaaga gcaactcggt cgccgcatac actattctca
     1621 gaatgacttg gttgagtact caccagtcac agaaaagcat cttacggatg gcatgacagt
     1681 aagagaatta tgcagtgctg ccataaccat gagtgataac actgcggcca acttacttct
     1741 gacaacgatc ggaggaccga aggagctaac cgcttttttg cacaacatgg gggatcatgt
     1801 aactcgcctt gatcgttggg aaccggagct gaatgaagcc ataccaaacg acgagcgtga
     1861 caccacgatg cctgtagcaa tggcaacaac gttgcgcaaa ctattaactg gcgaactact
     1921 tactctagct tcccggcaac aattaataga ctggatggag gcggataaag ttgcaggacc
     1981 acttctgcgc tcggcccttc cggctggctg gtttattgct gataaatctg gagccggtga
     2041 gcgtgggtct cgcggtatca ttgcagcact ggggccagat ggtaagccct cccgtatcgt
     2101 agttatctac acgacgggga gtcaggcaac tatggatgaa cgaaatagac agatcgctga
     2161 gataggtgcc tcactgatta agcattggta actgtcagac caagtttact catatatact
     2221 ttagattgat ttaaaacttc atttttaatt taaaaggatc taggtgaaga tcctttttga
     2281 taatctcatg aacaataaaa ctgtctgctt acataaacag taatacaagg ggtgttatga
     2341 gccatattca acgggaaacg tcttgctcta ggccgcgatt aaattccaac atggatgctg
     2401 atttatatgg gtataaatgg gctcgcgata atgtcgggca atcaggtgcg acaatctatc
     2461 gattgtatgg gaagcccgat gcgccagagt tgtttctgaa acatggcaaa ggtagcgttg
     2521 ccaatgatgt tacagatgag atggtcagac taaactggct gacggaattt atgcctcttc
     2581 cgaccatcaa gcattttatc cgtactcctg atgatgcatg gttactcacc actgcgatcc
     2641 ccgggaaaac agcattccag gtattagaag aatatcctga ttcaggtgaa aatattgttg
     2701 atgcgctggc agtgttcctg cgccggttgc attcgattcc tgtttgtaat tgtcctttta
     2761 acagcgatcg cgtatttcgt ctcgctcagg cgcaatcacg aatgaataac ggtttggttg
     2821 atgcgagtga ttttgatgac gagcgtaatg gctggcctgt tgaacaagtc tggaaagaaa
     2881 tgcataaact tttgccattc tcaccggatt cagtcgtcac tcatggtgat ttctcacttg
     2941 ataaccttat ttttgacgag gggaaattaa taggttgtat tgatgttgga cgagtcggaa
     3001 tcgcagaccg ataccaggat cttgccatcc tatggaactg cctcggtgag ttttctcctt
     3061 cattacagaa acggcttttt caaaaatatg gtattgataa tcctgatatg aataaattgc
     3121 agtttcattt gatgctcgat gagtttttct aagaattaat tcatgaccaa aatcccttaa
     3181 cgtgagtttt cgttccactg agcgtcagac cccgtagaaa agatcaaagg atcttcttga
     3241 gatccttttt ttctgcgcgt aatctgctgc ttgcaaacaa aaaaaccacc gctaccagcg
     3301 gtggtttgtt tgccggatca agagctacca actctttttc cgaaggtaac tggcttcagc
     3361 agagcgcaga taccaaatac tgttcttcta gtgtagccgt agttaggcca ccacttcaag
     3421 aactctgtag caccgcctac atacctcgct ctgctaatcc tgttaccagt ggctgctgcc
     3481 agtggcgata agtcgtgtct taccgggttg gactcaagac gatagttacc ggataaggcg
     3541 cagcggtcgg gctgaacggg gggttcgtgc acacagccca gcttggagcg aacgacctac
     3601 accgaactga gatacctaca gcgtgagcta tgagaaagcg ccacgcttcc cgaagggaga
     3661 aaggcggaca ggtatccggt aagcggcagg gtcggaacag gagagcgcac gagggagctt
     3721 ccagggggaa acgcctggta tctttatagt cctgtcgggt ttcgccacct ctgacttgag
     3781 cgtcgatttt tgtgatgctc gtcagggggg cggagcctat ggaaaaacgc cagcaacgcg
     3841 gcctttttac ggttcctggc cttttgctgg ccttttgctc acatgttctt tcctgcgtta
     3901 tcccctgatt ctgtggataa ccgtattacc gcctttgagt gagctgatac cgctcgccgc
     3961 agccgaacg
//
