LOCUS       sequence-468017-        5678 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Cloning vector with an N-terminal fusion of 10x His-tagged SUMO-1, 
            followed by a Golden Gate assembly site for seamless insertion..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5678)
  TITLE     Molecular biology strategies for optimized expression and 
            purification using SUMO protease-mediated elution.
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 5678)
  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: 
            "Unpublished"
COMMENT     Sequence Label: pAIP4
FEATURES             Location/Qualifiers
     source          1..5678
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             19..36
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(89..107)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      103..150
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     rep_origin      187..642
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(274..293)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     484..505
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     CDS             complement(734..1549)
                     /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             complement(734..1549)
                     /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     1457..1476
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     1541..1560
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     rep_origin      1671..2259
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     2160..2179
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     2413..2430
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    2445..2584
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(2510..2529)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     2670..2692
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(2686..2877)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     CDS             complement(3686..4768)
                     /codon_start=1
                     /gene="lacI"
                     /product="lac repressor"
                     /label=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)."
                     /translation="MKPVTLYDVAEYAGVSYQTVSRVVNQASHVSAKTREKVEAAMAEL
                     NYIPNRVAQQLAGKQSLLIGVATSSLALHAPSQIVAAIKSRADQLGASVVVSMVERSGV
                     EACKAAVHNLLAQRVSGLIINYPLDDQDAIAVEAACTNVPALFLDVSDQTPINSIIFSH
                     EDGTRLGVEHLVALGHQQIALLAGPLSSVSARLRLAGWHKYLTRNQIQPIAEREGDWSA
                     MSGFQQTMQMLNEGIVPTAMLVANDQMALGAMRAITESGLRVGADISVVGYDDTEDSSC
                     YIPPLTTIKQDFRLLGQTSVDRLLQLSQGQAVKGNQLLPVSLVKRKTTLAPNTQTASPR
                     ALADSLMQLARQVSRLESGQ"
     primer_bind     4730..4749
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(4769..4846)
                     /gene="lacI"
                     /label=lacI promoter
     primer_bind     5052..5071
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     primer_bind     5159..5178
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        5159..5177
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    5178..5202
                     /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)."
     RBS             5217..5239
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             5262..5291
                     /codon_start=1
                     /product="10xHis affinity tag"
                     /label=10xHis
                     /translation="HHHHHHHHHH"
ORIGIN
        1 ggttcgtaat gactcgagca ccaccaccac caccactgag atccggctgc taacaaagcc
       61 cgaaaggaag ctgagttggc tgctgccacc gctgagcaat aactagcata accccttggg
      121 gcctctaaac gggtcttgag gggttttttg ctgaaaggag gaactatatc cggattggcg
      181 aatgggacgc gccctgtagc ggcgcattaa gcgcggcggg tgtggtggtt acgcgcagcg
      241 tgaccgctac acttgccagc gccctagcgc ccgctccttt cgctttcttc ccttcctttc
      301 tcgccacgtt cgccggcttt ccccgtcaag ctctaaatcg ggggctccct ttagggttcc
      361 gatttagtgc tttacggcac ctcgacccca aaaaacttga ttagggtgat ggttcacgta
      421 gtgggccatc gccctgatag acggtttttc gccctttgac gttggagtcc acgttcttta
      481 atagtggact cttgttccaa actggaacaa cactcaaccc tatctcggtc tattcttttg
      541 atttataagg gattttgccg atttcggcct attggttaaa aaatgagctg atttaacaaa
      601 aatttaacgc gaattttaac aaaatattaa cgcttacaat ttaggtggca cttttcgggg
      661 aaatgtgcgc ggaaccccta tttgtttatt tttctaaata cattcaaata tgtatccgct
      721 catgaattaa ttcttagaaa aactcatcga gcatcaaatg aaactgcaat ttattcatat
      781 caggattatc aataccatat ttttgaaaaa gccgtttctg taatgaagga gaaaactcac
      841 cgaggcagtt ccataggatg gcaagatcct ggtatcggtc tgcgattccg actcgtccaa
      901 catcaataca acctattaat ttcccctcgt caaaaataag gttatcaagt gagaaatcac
      961 catgagtgac gactgaatcc ggtgagaatg gcaaaagttt atgcatttct ttccagactt
     1021 gttcaacagg ccagccatta cgctcgtcat caaaatcact cgcatcaacc aaaccgttat
     1081 tcattcgtga ttgcgcctga gcgagacgaa atacgcgatc gctgttaaaa ggacaattac
     1141 aaacaggaat cgaatgcaac cggcgcagga acactgccag cgcatcaaca atattttcac
     1201 ctgaatcagg atattcttct aatacctgga atgctgtttt cccggggatc gcagtggtga
     1261 gtaaccatgc atcatcagga gtacggataa aatgcttgat ggtcggaaga ggcataaatt
     1321 ccgtcagcca gtttagtctg accatctcat ctgtaacatc attggcaacg ctacctttgc
     1381 catgtttcag aaacaactct ggcgcatcgg gcttcccata caatcgatag attgtcgcac
     1441 ctgattgccc gacattatcg cgagcccatt tatacccata taaatcagca tccatgttgg
     1501 aatttaatcg cggcctagag caagacgttt cccgttgaat atggctcata acaccccttg
     1561 tattactgtt tatgtaagca gacagtttta ttgttcatga ccaaaatccc ttaacgtgag
     1621 ttttcgttcc actgagcgtc agaccccgta gaaaagatca aaggatcttc ttgagatcct
     1681 ttttttctgc gcgtaatctg ctgcttgcaa acaaaaaaac caccgctacc agcggtggtt
     1741 tgtttgccgg atcaagagct accaactctt tttccgaagg taactggctt cagcagagcg
     1801 cagataccaa atactgtcct tctagtgtag ccgtagttag gccaccactt caagaactct
     1861 gtagcaccgc ctacatacct cgctctgcta atcctgttac cagtggctgc tgccagtggc
     1921 gataagtcgt gtcttaccgg gttggactca agacgatagt taccggataa ggcgcagcgg
     1981 tcgggctgaa cggggggttc gtgcacacag cccagcttgg agcgaacgac ctacaccgaa
     2041 ctgagatacc tacagcgtga gctatgagaa agcgccacgc ttcccgaagg gagaaaggcg
     2101 gacaggtatc cggtaagcgg cagggtcgga acaggagagc gcacgaggga gcttccaggg
     2161 ggaaacgcct ggtatcttta tagtcctgtc gggtttcgcc acctctgact tgagcgtcga
     2221 tttttgtgat gctcgtcagg ggggcggagc ctatggaaaa acgccagcaa cgcggccttt
     2281 ttacggttcc tggccttttg ctggcctttt gctcacatgt tctttcctgc gttatcccct
     2341 gattctgtgg ataaccgtat taccgccttt gagtgagctg ataccgctcg ccgcagccga
     2401 acgaccgagc gcagcgagtc agtgagcgag gaagcggaag agcgcctgat gcggtatttt
     2461 ctccttacgc atctgtgcgg tatttcacac cgcaatggtg cactctcagt acaatctgct
     2521 ctgatgccgc atagttaagc cagtatacac tccgctatcg ctacgtgact gggtcatggc
     2581 tgcgccccga cacccgccaa cacccgctga cgcgccctga cgggcttgtc tgctcccggc
     2641 atccgcttac agacaagctg tgaccgtctc cgggagctgc atgtgtcaga ggttttcacc
     2701 gtcatcaccg aaacgcgcga ggcagctgcg gtaaagctca tcagcgtggt cgtgaagcga
     2761 ttcacagatg tctgcctgtt catccgcgtc cagctcgttg agtttctcca gaagcgttaa
     2821 tgtctggctt ctgataaagc gggccatgtt aagggcggtt ttttcctgtt tggtcactga
     2881 tgcctccgtg taagggggat ttctgttcat gggggtaatg ataccgatga aacgagagag
     2941 gatgctcacg atacgggtta ctgatgatga acatgcccgg ttactggaac gttgtgaggg
     3001 taaacaactg gcggtatgga tgcggcggga ccagagaaaa atcactcagg gtcaatgcca
     3061 gcgcttcgtt aatacagatg taggtgttcc acagggtagc cagcagcatc ctgcgatgca
     3121 gatccggaac ataatggtgc agggcgctga cttccgcgtt tccagacttt acgaaacacg
     3181 gaaaccgaag accattcatg ttgttgctca ggtcgcagac gttttgcagc agcagtcgct
     3241 tcacgttcgc tcgcgtatcg gtgattcatt ctgctaacca gtaaggcaac cccgccagcc
     3301 tagccgggtc ctcaacgaca ggagcacgat catgcgcacc cgtggggccg ccatgccggc
     3361 gataatggcc tgcttctcgc cgaaacgttt ggtggcggga ccagtgacga aggcttgagc
     3421 gagggcgtgc aagattccga ataccgcaag cgacaggccg atcatcgtcg cgctccagcg
     3481 aaagcggtcc tcgccgaaaa tgacccagag cgctgccggc acctgtccta cgagttgcat
     3541 gataaagaag acagtcataa gtgcggcgac gatagtcatg ccccgcgccc accggaagga
     3601 gctgactggg ttgaaggctc tcaagggcat cggtcgagat cccggtgcct aatgagtgag
     3661 ctaacttaca ttaattgcgt tgcgctcact gcccgctttc cagtcgggaa acctgtcgtg
     3721 ccagctgcat taatgaatcg gccaacgcgc ggggagaggc ggtttgcgta ttgggcgcca
     3781 gggtggtttt tcttttcacc agtgagacgg gcaacagctg attgcccttc accgcctggc
     3841 cctgagagag ttgcagcaag cggtccacgc tggtttgccc cagcaggcga aaatcctgtt
     3901 tgatggtggt taacggcggg atataacatg agctgtcttc ggtatcgtcg tatcccacta
     3961 ccgagatgtc cgcaccaacg cgcagcccgg actcggtaat ggcgcgcatt gcgcccagcg
     4021 ccatctgatc gttggcaacc agcatcgcag tgggaacgat gccctcattc agcatttgca
     4081 tggtttgttg aaaaccggac atggcactcc agtcgccttc ccgttccgct atcggctgaa
     4141 tttgattgcg agtgagatat ttatgccagc cagccagacg cagacgcgcc gagacagaac
     4201 ttaatgggcc cgctaacagc gcgatttgct ggtgacccaa tgcgaccaga tgctccacgc
     4261 ccagtcgcgt accgtcttca tgggagaaaa taatactgtt gatgggtgtc tggtcagaga
     4321 catcaagaaa taacgccgga acattagtgc aggcagcttc cacagcaatg gcatcctggt
     4381 catccagcgg atagttaatg atcagcccac tgacgcgttg cgcgagaaga ttgtgcaccg
     4441 ccgctttaca ggcttcgacg ccgcttcgtt ctaccatcga caccaccacg ctggcaccca
     4501 gttgatcggc gcgagattta atcgccgcga caatttgcga cggcgcgtgc agggccagac
     4561 tggaggtggc aacgccaatc agcaacgact gtttgcccgc cagttgttgt gccacgcggt
     4621 tgggaatgta attcagctcc gccatcgccg cttccacttt ttcccgcgtt ttcgcagaaa
     4681 cgtggctggc ctggttcacc acgcgggaaa cggtctgata agagacaccg gcatactctg
     4741 cgacatcgta taacgttact ggtttcacat tcaccaccct gaattgactc tcttccgggc
     4801 gctatcatgc cataccgcga aaggttttgc gccattcgat ggtgtccggg atctcgacgc
     4861 tctcccttat gcgactcctg cattaggaag cagcccagta gtaggttgag gccgttgagc
     4921 accgccgccg caaggaatgg tgcatgcaag gagatggcgc ccaacagtcc cccggccacg
     4981 gggcctgcca ccatacccac gccgaaacaa gcgctcatga gcccgaagtg gcgagcccga
     5041 tcttccccat cggtgatgtc ggcgatatag gcgccagcaa ccgcacctgt ggcgccggtg
     5101 atgccggcca cgatgcgtcc ggcgtagagg atcgagatcg atctcgatcc cgcgaaatta
     5161 atacgactca ctatagggga attgtgagcg gataacaatt cccctctaga aataattttg
     5221 tttaacttta agaaggagat atacatatga tggcgaagat tcatcaccat catcaccatc
     5281 accatcacca cggtagcctg caagattcgg aagtcaacca agaagcaaag ccggaagtca
     5341 agccggaagt gaaaccggaa acccatatta acctgaaagt tagtgacggc agctctgaaa
     5401 ttttctttaa gatcaaaaag accacgccgc tgcgtcgcct gatggaagcg tttgccaaac
     5461 gtcagggcaa ggaaatggat agcctgcgtt tcctgtatga cggtattcgc atccaggcag
     5521 atcaagcgcc ggaagacctg gacatggaag acaacgacat cattgaagcc caccgtgaac
     5581 agattggcgg ctctggagag accggagcga gacgggagtc gtcttcgctt tccagatctg
     5641 ataacttgtg aagacgacca tcgtctcaca gtggtctc
//
