LOCUS       sequence-318922-        7028 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses B. malayi iPGM containing a C-terminal biotinylation site 
            and His tag.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7028)
  AUTHORS   Wiedmann M, Dranchak PK, Aitha M, Queme B, Collmus CD, Kashipathy 
            MM, Kanter L, Lamy L, Rogers JM, Tao D, Battaile KP, Rai G, Lovell 
            S, Suga H, Inglese J
  TITLE     Structure-activity relationship of ipglycermide binding to 
            phosphoglycerate mutases.
  JOURNAL   J Biol Chem. 2021 Apr 1:100628. doi: 10.1016/j.jbc.2021.100628.
  PUBMED    33812994
REFERENCE   2  (bases 1 to 7028)
  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: "J Biol 
            Chem. 2021 Apr 1:100628. doi: 10.1016/j.jbc.2021.100628."
COMMENT     Sequence Label: pET21a Bm-iPGM-bio
FEATURES             Location/Qualifiers
     source          1..7028
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     1..20
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        1..19
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    20..44
                     /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             59..81
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             89..121
                     /codon_start=1
                     /product="leader peptide from bacteriophage T7 gene 10"
                     /label=T7 tag (gene 10 leader)
                     /note="promotes efficient translation in E. coli"
                     /translation="MASMTGGQQMG"
     CDS             1682..1726
                     /codon_start=1
                     /product="peptide tag that allows for enzymatic 
                     biotinylation"
                     /label=AviTag(TM)
                     /translation="GLNDIFEAQKIEWHE"
     CDS             1727..1744
                     /codon_start=1
                     /product="thrombin recognition and cleavage site"
                     /label=thrombin site
                     /translation="LVPRGS"
     CDS             1760..1777
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(1830..1848)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      1844..1891
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     rep_origin      1928..2383
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2015..2034)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2225..2246
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        2409..2513
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(2514..2582,2583..3374)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPAAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     complement(2732..2751)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      3545..4133
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4034..4053
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     4287..4304
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    4319..4458
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(4384..4403)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     4544..4566
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(4560..4751)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     CDS             complement(5560..6642)
                     /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     6604..6623
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(6643..6720)
                     /gene="lacI"
                     /label=lacI promoter
     primer_bind     6926..6945
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
ORIGIN
        1 taatacgact cactataggg gaattgtgag cggataacaa ttcccctcta gaaataattt
       61 tgtttaactt taagaaggag atatacatat ggctagcatg actggtggac agcaaatggg
      121 tcgcggatcc atggccgaag caaagaatcg agtatgtctg gtagtgattg atggttgggg
      181 aatcagtaac gaaactaaag gcaatgcaat actaaatgct aaaacacctg taatggatga
      241 gctttgtgta atgaattcgc atccaattca agcacatggc ttgcatgttg gtttaccgga
      301 aggacttatg ggcaattcag aagtgggtca tttaaacatt ggcgctggcc gtgttgttta
      361 tcaggatatt gtacgcataa atttggcggt caagaataag actttggtgg aaaataagca
      421 tttgaaggaa gctgctgaac gtgcaattaa agggaatggc cgcatgcact tatgtggttt
      481 ggtcagcgat ggtggtgtgc attcacatat tgatcatttg tttgctttga taacagcttt
      541 gaaacaactt aaagtaccga agctttacat tcaattcttt ggagatggtc gtgatacgag
      601 tccaacaagc ggagttggtt tccttcaaca gctaattgat ttcgtcaaca aggaacaata
      661 tggtgaaata tcaacaatag tagggcgcta ctatgcgatg gacagagata aacggtggga
      721 acgaattcgg gtatgttatg atgcactaat tggtggagtt ggtgagaaga ctacaattga
      781 taaggcgatt gatgttatca aaggacgata tgcaaaggat gagactgatg aattcctaaa
      841 accaataatt ctttcggatg aaggacgtac aaaagatggt gatactttga tattctttga
      901 ttatcgtgct gatcgtatgc gagaaatcac tgaatgcatg ggtatggaac gatacaaaga
      961 tcttaattct aatattaaac atccaaagaa tatgcaagta attggaatga ctcagtacaa
     1021 ggcagaattt acctttcctg cactttttcc tccggaatct cataaaaatg tattggcgga
     1081 atggttatct gtaaatggat taacacaatt ccattgtgct gaaacagaaa aatatgcgca
     1141 cgttacattc ttcttcaatg gtggtgtgga aaaacaattt gcaaatgaag agcgttgttt
     1201 agtagtatct ccgaaagttg ccacttatga tcttgaacca ccaatgagtt cagctgctgt
     1261 agctgataag gtgattgagc aattgcatat gaaaaaacat ccatttgtta tgtgcaattt
     1321 tgcacctccc gatatggttg gccatactgg agtttatgaa gcagccgtga aagcagttga
     1381 agcaactgat attgctatcg gacgaatata tgaagcatgt aagaagaatg actacatact
     1441 gatggtaact gctgatcatg gaaatgctga gaaaatgatg gcaccagatg gtagcaagca
     1501 tactgctcac acttgcaatt tagtgccatt cacttgttcc tcaatgaaat acaaattcat
     1561 ggacaagtta ccggatcggg agatggctct ttgtgatgtt gctccaacag ttctaaaagt
     1621 tatgggtgtg ccattgccat ccgagatgac cggacagcca ttggttaatg aagccctcga
     1681 gggtctgaac gacatcttcg aggctcagaa aatcgaatgg cacgaattag tgcctcgcgg
     1741 aagcgctgca gggctcgagc accaccacca ccaccactga gatccggctg ctaacaaagc
     1801 ccgaaaggaa gctgagttgg ctgctgccac cgctgagcaa taactagcat aaccccttgg
     1861 ggcctctaaa cgggtcttga ggggtttttt gctgaaagga ggaactatat ccggattggc
     1921 gaatgggacg cgccctgtag cggcgcatta agcgcggcgg gtgtggtggt tacgcgcagc
     1981 gtgaccgcta cacttgccag cgccctagcg cccgctcctt tcgctttctt cccttccttt
     2041 ctcgccacgt tcgccggctt tccccgtcaa gctctaaatc gggggctccc tttagggttc
     2101 cgatttagtg ctttacggca cctcgacccc aaaaaacttg attagggtga tggttcacgt
     2161 agtgggccat cgccctgata gacggttttt cgccctttga cgttggagtc cacgttcttt
     2221 aatagtggac tcttgttcca aactggaaca acactcaacc ctatctcggt ctattctttt
     2281 gatttataag ggattttgcc gatttcggcc tattggttaa aaaatgagct gatttaacaa
     2341 aaatttaacg cgaattttaa caaaatatta acgcttacaa tttaggtggc acttttcggg
     2401 gaaatgtgcg cggaacccct atttgtttat ttttctaaat acattcaaat atgtatccgc
     2461 tcatgagaca ataaccctga taaatgcttc aataatattg aaaaaggaag agtatgagta
     2521 ttcaacattt ccgtgtcgcc cttattccct tttttgcggc attttgcctt cctgtttttg
     2581 ctcacccaga aacgctggtg aaagtaaaag atgctgaaga tcagttgggt gcacgagtgg
     2641 gttacatcga actggatctc aacagcggta agatccttga gagttttcgc cccgaagaac
     2701 gttttccaat gatgagcact tttaaagttc tgctatgtgg cgcggtatta tcccgtattg
     2761 acgccgggca agagcaactc ggtcgccgca tacactattc tcagaatgac ttggttgagt
     2821 actcaccagt cacagaaaag catcttacgg atggcatgac agtaagagaa ttatgcagtg
     2881 ctgccataac catgagtgat aacactgcgg ccaacttact tctgacaacg atcggaggac
     2941 cgaaggagct aaccgctttt ttgcacaaca tgggggatca tgtaactcgc cttgatcgtt
     3001 gggaaccgga gctgaatgaa gccataccaa acgacgagcg tgacaccacg atgcctgcag
     3061 caatggcaac aacgttgcgc aaactattaa ctggcgaact acttactcta gcttcccggc
     3121 aacaattaat agactggatg gaggcggata aagttgcagg accacttctg cgctcggccc
     3181 ttccggctgg ctggtttatt gctgataaat ctggagccgg tgagcgtggg tctcgcggta
     3241 tcattgcagc actggggcca gatggtaagc cctcccgtat cgtagttatc tacacgacgg
     3301 ggagtcaggc aactatggat gaacgaaata gacagatcgc tgagataggt gcctcactga
     3361 ttaagcattg gtaactgtca gaccaagttt actcatatat actttagatt gatttaaaac
     3421 ttcattttta atttaaaagg atctaggtga agatcctttt tgataatctc atgaccaaaa
     3481 tcccttaacg tgagttttcg ttccactgag cgtcagaccc cgtagaaaag atcaaaggat
     3541 cttcttgaga tccttttttt ctgcgcgtaa tctgctgctt gcaaacaaaa aaaccaccgc
     3601 taccagcggt ggtttgtttg ccggatcaag agctaccaac tctttttccg aaggtaactg
     3661 gcttcagcag agcgcagata ccaaatactg tccttctagt gtagccgtag ttaggccacc
     3721 acttcaagaa ctctgtagca ccgcctacat acctcgctct gctaatcctg ttaccagtgg
     3781 ctgctgccag tggcgataag tcgtgtctta ccgggttgga ctcaagacga tagttaccgg
     3841 ataaggcgca gcggtcgggc tgaacggggg gttcgtgcac acagcccagc ttggagcgaa
     3901 cgacctacac cgaactgaga tacctacagc gtgagctatg agaaagcgcc acgcttcccg
     3961 aagggagaaa ggcggacagg tatccggtaa gcggcagggt cggaacagga gagcgcacga
     4021 gggagcttcc agggggaaac gcctggtatc tttatagtcc tgtcgggttt cgccacctct
     4081 gacttgagcg tcgatttttg tgatgctcgt caggggggcg gagcctatgg aaaaacgcca
     4141 gcaacgcggc ctttttacgg ttcctggcct tttgctggcc ttttgctcac atgttctttc
     4201 ctgcgttatc ccctgattct gtggataacc gtattaccgc ctttgagtga gctgataccg
     4261 ctcgccgcag ccgaacgacc gagcgcagcg agtcagtgag cgaggaagcg gaagagcgcc
     4321 tgatgcggta ttttctcctt acgcatctgt gcggtatttc acaccgcaat ggtgcactct
     4381 cagtacaatc tgctctgatg ccgcatagtt aagccagtat acactccgct atcgctacgt
     4441 gactgggtca tggctgcgcc ccgacacccg ccaacacccg ctgacgcgcc ctgacgggct
     4501 tgtctgctcc cggcatccgc ttacagacaa gctgtgaccg tctccgggag ctgcatgtgt
     4561 cagaggtttt caccgtcatc accgaaacgc gcgaggcagc tgcggtaaag ctcatcagcg
     4621 tggtcgtgaa gcgattcaca gatgtctgcc tgttcatccg cgtccagctc gttgagtttc
     4681 tccagaagcg ttaatgtctg gcttctgata aagcgggcca tgttaagggc ggttttttcc
     4741 tgtttggtca ctgatgcctc cgtgtaaggg ggatttctgt tcatgggggt aatgataccg
     4801 atgaaacgag agaggatgct cacgatacgg gttactgatg atgaacatgc ccggttactg
     4861 gaacgttgtg agggtaaaca actggcggta tggatgcggc gggaccagag aaaaatcact
     4921 cagggtcaat gccagcgctt cgttaataca gatgtaggtg ttccacaggg tagccagcag
     4981 catcctgcga tgcagatccg gaacataatg gtgcagggcg ctgacttccg cgtttccaga
     5041 ctttacgaaa cacggaaacc gaagaccatt catgttgttg ctcaggtcgc agacgttttg
     5101 cagcagcagt cgcttcacgt tcgctcgcgt atcggtgatt cattctgcta accagtaagg
     5161 caaccccgcc agcctagccg ggtcctcaac gacaggagca cgatcatgcg cacccgtggg
     5221 gccgccatgc cggcgataat ggcctgcttc tcgccgaaac gtttggtggc gggaccagtg
     5281 acgaaggctt gagcgagggc gtgcaagatt ccgaataccg caagcgacag gccgatcatc
     5341 gtcgcgctcc agcgaaagcg gtcctcgccg aaaatgaccc agagcgctgc cggcacctgt
     5401 cctacgagtt gcatgataaa gaagacagtc ataagtgcgg cgacgatagt catgccccgc
     5461 gcccaccgga aggagctgac tgggttgaag gctctcaagg gcatcggtcg agatcccggt
     5521 gcctaatgag tgagctaact tacattaatt gcgttgcgct cactgcccgc tttccagtcg
     5581 ggaaacctgt cgtgccagct gcattaatga atcggccaac gcgcggggag aggcggtttg
     5641 cgtattgggc gccagggtgg tttttctttt caccagtgag acgggcaaca gctgattgcc
     5701 cttcaccgcc tggccctgag agagttgcag caagcggtcc acgctggttt gccccagcag
     5761 gcgaaaatcc tgtttgatgg tggttaacgg cgggatataa catgagctgt cttcggtatc
     5821 gtcgtatccc actaccgaga tatccgcacc aacgcgcagc ccggactcgg taatggcgcg
     5881 cattgcgccc agcgccatct gatcgttggc aaccagcatc gcagtgggaa cgatgccctc
     5941 attcagcatt tgcatggttt gttgaaaacc ggacatggca ctccagtcgc cttcccgttc
     6001 cgctatcggc tgaatttgat tgcgagtgag atatttatgc cagccagcca gacgcagacg
     6061 cgccgagaca gaacttaatg ggcccgctaa cagcgcgatt tgctggtgac ccaatgcgac
     6121 cagatgctcc acgcccagtc gcgtaccgtc ttcatgggag aaaataatac tgttgatggg
     6181 tgtctggtca gagacatcaa gaaataacgc cggaacatta gtgcaggcag cttccacagc
     6241 aatggcatcc tggtcatcca gcggatagtt aatgatcagc ccactgacgc gttgcgcgag
     6301 aagattgtgc accgccgctt tacaggcttc gacgccgctt cgttctacca tcgacaccac
     6361 cacgctggca cccagttgat cggcgcgaga tttaatcgcc gcgacaattt gcgacggcgc
     6421 gtgcagggcc agactggagg tggcaacgcc aatcagcaac gactgtttgc ccgccagttg
     6481 ttgtgccacg cggttgggaa tgtaattcag ctccgccatc gccgcttcca ctttttcccg
     6541 cgttttcgca gaaacgtggc tggcctggtt caccacgcgg gaaacggtct gataagagac
     6601 accggcatac tctgcgacat cgtataacgt tactggtttc acattcacca ccctgaattg
     6661 actctcttcc gggcgctatc atgccatacc gcgaaaggtt ttgcgccatt cgatggtgtc
     6721 cgggatctcg acgctctccc ttatgcgact cctgcattag gaagcagccc agtagtaggt
     6781 tgaggccgtt gagcaccgcc gccgcaagga atggtgcatg caaggagatg gcgcccaaca
     6841 gtcccccggc cacggggcct gccaccatac ccacgccgaa acaagcgctc atgagcccga
     6901 agtggcgagc ccgatcttcc ccatcggtga tgtcggcgat ataggcgcca gcaaccgcac
     6961 ctgtggcgcc ggtgatgccg gccacgatgc gtccggcgta gaggatcgag atctcgatcc
     7021 cgcgaaat
//
