LOCUS       sequence-421879-        4169 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  For expression of VHH with C-terminal sortase and his tags.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4169)
  AUTHORS   Eyssen LE, Ramadurai S, Abdelkarim S, Buckle I, Cornish K, Lin H, 
            Jones AK, Stephens GJ, Owens RJ
  TITLE     From Llama to Nanobody: A Streamlined Workflow for the Generation of
            Functionalised VHHs.
  JOURNAL   Bio Protoc. 2024 Mar 20;14(6):e4962. doi: 10.21769/BioProtoc.4962. 
            eCollection 2024 Mar 20.
  PUBMED    38841291
REFERENCE   2  (bases 1 to 4169)
  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: "Bio 
            Protoc."; date: "2024-03-20"; volume: "14(6):e4962. doi"; pages: " 
            10.21769/BioProtoc.4962. eCollection 2024 Mar 20"
COMMENT     Sequence Label: pOPINVHH_sortase_his
FEATURES             Location/Qualifiers
     source          1..4169
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        21..125
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(126..194,195..986)
                     /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(344..363)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      1157..1745
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     1646..1665
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     protein_bind    1910..1931
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(1946..1951,1952..1969,1970..1976)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    1984..2000
                     /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)."
     CDS             2089..2262
                     /codon_start=1
                     /gene="lacZ fragment"
                     /product="LacZ-alpha fragment of beta-galactosidase"
                     /label=lacZ-alpha
                     /translation="MTMITDSLAVVLQRRDWENPGVTQLNRLAAHPPFASWRNSEEART
                     DRPSQQLRSLNGE"
     primer_bind     complement(2109..2126)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(2109..2125)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2118..2140)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     CDS             2301..2318
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             2341..2370
                     /codon_start=1
                     /product="Myc (human c-Myc proto-oncogene) epitope tag"
                     /label=Myc
                     /translation="EQKLISEEDL"
     rep_origin      complement(3735..4163)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3845..3866)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     4057..4076
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
ORIGIN
        1 gcacttttcg gggaaatgtg cgcggaaccc ctatttgttt atttttctaa atacattcaa
       61 atatgtatcc gctcatgaga caataaccct gataaatgct tcaataatat tgaaaaagga
      121 agagtatgag tattcaacat ttccgtgtcg cccttattcc cttttttgcg gcattttgcc
      181 ttcctgtttt tgctcaccca gaaacgctgg tgaaagtaaa agatgctgaa gatcagttgg
      241 gtgcacgagt gggttacatc gaactggatc tcaacagcgg taagatcctt gagagttttc
      301 gccccgaaga acgttttcca atgatgagca cttttaaagt tctgctatgt ggcgcggtat
      361 tatcccgtat tgacgccggg caagagcaac tcggtcgccg catacactat tctcagaatg
      421 acttggttga gtactcacca gtcacagaaa agcatcttac ggatggcatg acagtaagag
      481 aattatgcag tgctgccata accatgagtg ataacactgc ggccaactta cttctgacaa
      541 cgatcggagg accgaaggag ctaaccgctt ttttgcacaa catgggggat catgtaactc
      601 gccttgatcg ttgggaaccg gagctgaatg aagccatacc aaacgacgag cgtgacacca
      661 cgatgcctgt agcaatggca acaacgttgc gcaaactatt aactggcgaa ctacttactc
      721 tagcttcccg gcaacaatta atagactgga tggaggcgga taaagttgca ggaccacttc
      781 tgcgctcggc gcttccggct ggctggttta ttgctgataa atctggagcc ggtgagcgtg
      841 ggtctcgcgg tatcattgca gcactggggc cagatggtaa gccctcccgt atcgtagtta
      901 tctacacgac ggggagtcag gcaactatgg atgaacgaaa tagacagatc gctgagatag
      961 gtgcctcact gattaagcat tggtaactgt cagaccaagt ttactcatat atactttaga
     1021 ttgatttaaa acttcatttt taatttaaaa ggatctaggt gaagatcctt tttgataatc
     1081 tcatgaccaa aatcccttaa cgtgagtttt cgttccactg agcgtcagac cccgtagaaa
     1141 agatcaaagg atcttcttga gatccttttt ttctgcgcgt aatctgctgc ttgcaaacaa
     1201 aaaaaccacc gctaccagcg gtggtttgtt tgccggatca agagctacca actctttttc
     1261 cgaaggtaac tggcttcagc agagcgcaga taccaaatac tgttcttcta gtgtagccgt
     1321 agttaggcca ccacttcaag aactctgtag caccgcctac atacctcgct ctgctaatcc
     1381 tgttaccagt ggctgctgcc agtggcgata agtcgtgtct taccgggttg gactcaagac
     1441 gatagttacc ggataaggcg cagcggtcgg gctgaacggg gggttcgtgc acacagccca
     1501 gcttggagcg aacgacctac accgaactga gatacctaca gcgtgagcta tgagaaagcg
     1561 ccacgcttcc cgaagggaga aaggcggaca ggtatccggt aagcggcagg gtcggaacag
     1621 gagagcgcac gagggagctt ccagggggaa acgcctggta tctttatagt cctgtcgggt
     1681 ttcgccacct ctgacttgag cgtcgatttt tgtgatgctc gtcagggggg cggagcctat
     1741 ggaaaaacgc cagcaacgcg gcctttttac ggttcctggc cttttgctgg ccttttgctc
     1801 acatgcgccc aatacgcaaa ccgcctctcc ccgcgcgttg gccgattcat taatgcagct
     1861 ggcacgacag gtttcccgac tggaaagcgg gcagtgagcg caacgcaatt aatgtgagtt
     1921 agctcactca ttaggcaccc caggctttac actttatgct tccggctcgt atgttgtgtg
     1981 gaattgtgag cggataacaa tttgaattca aggagacagt cataatgaaa tacctattgc
     2041 ctacggcggc cgctggattg ttattactcg cggcccagcc ggccatggat gaccatgatt
     2101 acggattcac tggccgtcgt tttacaacgt cgtgactggg aaaaccctgg cgttacccaa
     2161 cttaatcgcc ttgcagcaca tccccctttc gccagctggc gtaatagcga agaggcccgc
     2221 accgatcgcc cttcccaaca gttgcgcagc ctgaatggcg aatggcgcca tcctcaggcg
     2281 gcctgccgga aaccggcggc catcatcatc atcatcatta ataaccacca tcatggcgca
     2341 gaacaaaaac tcatctcaga agaggatctg tcttaggccg aaactgttga aagttgttta
     2401 gcaaaacctc atacagaaaa ttcatttact aacgtctgga aagacgacaa aactttagat
     2461 cgttacgcta actatgaggg ctgtctgtgg aatgctacag gcgttgtggt ttgtactggt
     2521 gacgaaactc agtgttacgg tacatgggtt cctattgggc ttgctatccc tgaaaatgag
     2581 ggtggtggct ctgagggtgg cggttctgag ggtggcggtt ctgagggtgg cggtactaaa
     2641 cctcctgagt acggtgatac acctattccg ggctatactt atatcaaccc tctcgacggc
     2701 acttatccgc ctggtactga gcaaaacccc gctaatccta atccttctct tgaggagtct
     2761 cagcctctta atactttcat gtttcagaat aataggttcc gaaataggca gggtgcatta
     2821 actgtttata cgggcactgt tactcaaggc actgaccccg ttaaaactta ttaccagtac
     2881 actcctgtat catcaaaagc catgtatgac gcttactgga acggtaaatt cagagactgc
     2941 gctttccatt ctggctttaa tgaggatcca ttcgtttgtg aatatcaagg ccaatcgtct
     3001 gacctgcctc aacctcctgt caatgctggc ggcggctctg gtggtggttc tggtggcggc
     3061 tctgagggtg gcggctctga gggtggcggt tctgagggtg gcggctctga gggtggcggt
     3121 tccggtggcg gctccggttc cggtgatttt gattatgaaa aaatggcaaa cgctaataag
     3181 ggggctatga ccgaaaatgc cgatgaaaac gcgctacagt ctgacgctaa aggcaaactt
     3241 gattctgtcg ctactgatta cggtgctgct atcgatggtt tcattggtga cgtttccggc
     3301 cttgctaatg gtaatggtgc tactggtgat tttgctggct ctaattccca aatggctcaa
     3361 gtcggtgacg gtgataattc acctttaatg aataatttcc gtcaatattt accttctttg
     3421 cctcagtcgg ttgaatgtcg cccttatgtc tttggcgctg gtaaaccata tgaattttct
     3481 attgattgtg acaaaataaa cttattccgt ggtgtctttg cgtttctttt atatgttgcc
     3541 acctttatgt atgtattttc gacgtttgct aacatactgc gtaataagga gtcttaagct
     3601 agctaacagt cctatgaatc aactacttag atggtattag tgacctgtaa cagagcatta
     3661 gcgcaaggtg atttttgtct tcttgcgcta attttttgtc atcaaacctg tcgcactcct
     3721 taatattttg ttaaaattcg cgttaaattt ttgttaaatc agctcatttt ttaaccaata
     3781 ggccgaaatc ggcaaaatcc cttataaatc aaaagaatag accgagatag ggttgagtgt
     3841 tgttccagtt tggaacaaga gtccactatt aaagaacgtg gactccaacg tcaaagggcg
     3901 aaaaaccgtc tatcagggcg atggcccact acgtgaacca tcaccctaat caagtttttt
     3961 ggggtcgagg tgccgtaaag cactaaatcg gaaccctaaa gggagccccc gatttagagc
     4021 ttgacgggga aagccggcga acgtggcgag aaaggaaggg aagaaagcga aaggagcggg
     4081 cgctagggcg ctggcaagtg tagcggtcac gctgcgcgta accaccacac ccgccgcgct
     4141 taatgcgccg ctacagggcg cgtcaggtg
//
