LOCUS       sequence-496976-        6792 bp    DNA     circular SYN 12-JAN-2026
DEFINITION  Expresses trypansoma brucei 6 phosphogluconate dehydrogenase under 
            T7 with HIS tag in bacterial cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6792)
  TITLE     Josh Beckham Lab plasmids 2025
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 6792)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Jan 12, 2026 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: 
            "Unpublished"
COMMENT     Sequence Label: TB6PDGH-pNIC-Bsa4
FEATURES             Location/Qualifiers
     source          1..6792
                     /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             92..109
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             134..154
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQS"
     CDS             1657..1674
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(1727..1745)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      1741..1788
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     rep_origin      1825..2280
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1912..1931)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2122..2143
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     CDS             complement(2372..3187)
                     /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(2372..3187)
                     /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     3095..3114
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     3179..3198
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     rep_origin      3309..3897
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3798..3817
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     4051..4068
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    4083..4222
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(4148..4167)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     4308..4330
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(4324..4515)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     CDS             complement(5324..6406)
                     /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     6368..6387
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(6407..6484)
                     /gene="lacI"
                     /label=lacI promoter
     primer_bind     6690..6709
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
ORIGIN
        1 taatacgact cactataggg gaattgtgag cggataacaa ttcccctcta gaaataattt
       61 tgtttaactt taagaaggag atatacatat gcaccatcat catcatcatt cttctggtgt
      121 agatctgggt accgagaacc tgtacttcca atccatgtct atggacgttg gtgtagttgg
      181 cctgggtgtt atgggtgcga acctggcgct gaatattgcg gaaaaaggtt tcaaagttgc
      241 ggttttcaac cgtacctact ctaaatctga agaatttatg aaagcgaacg cttctgcacc
      301 gtttgcgggc aacctgaagg cgtttgagac catggaagca tttgcggcgt ctctcaagaa
      361 accgcgtaaa gcgctcattc tggtacaagc gggtgcggcc accgactcta ccatcgaaca
      421 gctcaaaaaa gtatttgaaa agggtgacat cctggtcgac accggtaacg cgcatttcaa
      481 ggaccagggt cgtcgtgcgc agcagctgga ggctgcgggt ctccgtttcc tcggcatggg
      541 tatttctggt ggtgaagaag gtgcgcgtaa gggtccggcg ttcttcccgg gtggtaccct
      601 gtctgtatgg gaagagattc gtccgatcgt tgaagctgcc gctgcgaaag cggacgatgg
      661 tcgcccgtgc gtaacgatga atggttccgg tggtgctggt tcttgcgtta aaatgtacca
      721 caacagcggt gaatacgcga tcctgcaaat ctggggtgaa gtattcgata tcctgcgcgc
      781 gatgggcctg aataatgacg aagtagccgc ggttctggaa gactggaaaa gcaaaaactt
      841 tctcaaatcc tatatgctgg acatctccat cgctgcggca cgcgctaaag acaaggacgg
      901 ctcctacctg accgagcacg tgatggaccg tatcggtagc aaaggtaccg gtctgtggtc
      961 tgcccaggaa gcgctggaaa tcggtgtacc tgcgccatcc ctgaacatgg cggtagtttc
     1021 ccgccaattc accatgtaca agaccgaacg tcaggcgaat gcgtctaacg ctccgggtat
     1081 cacccagtct cctggttaca ccctgaaaaa caaatctccg tctggtccgg aaatcaaaca
     1141 actgtacgac tctgtttgca ttgcgatcat ctcttgctac gcccaaatgt tccaatgcct
     1201 gcgtgaaatg gacaaagttc acaatttcgg cctcaacctc ccagcgacta tcgcgacctt
     1261 tcgtgcgggc tgtatcctcc agggttacct gctgaaaccg atgacggagg cattcgagaa
     1321 aaatccgaac atctctaacc tcatgtgcgc attccaaact gagatccgtg ccggcctcca
     1381 aaactatcgt gatatggttg ctctgattac ctctaaactg gaagtttcta tcccagttct
     1441 gagcgcgagc ctgaactacg taaccgctat gttcactccg acgctgaaat atggccagct
     1501 cgtttctctg cagcgtgacg ttttcggtcg tcatggttac gaacgtgttg acaaagatgg
     1561 ccgcgaatct tttcagtggc cggagctgca ataacagtaa aggtggatac ggatccgaat
     1621 tcgagctccg tcgacaagct tgcggccgca ctcgagcacc accaccacca ccactgagat
     1681 ccggctgcta acaaagcccg aaaggaagct gagttggctg ctgccaccgc tgagcaataa
     1741 ctagcataac cccttggggc ctctaaacgg gtcttgaggg gttttttgct gaaaggagga
     1801 actatatccg gattggcgaa tgggacgcgc cctgtagcgg cgcattaagc gcggcgggtg
     1861 tggtggttac gcgcagcgtg accgctacac ttgccagcgc cctagcgccc gctcctttcg
     1921 ctttcttccc ttcctttctc gccacgttcg ccggctttcc ccgtcaagct ctaaatcggg
     1981 ggctcccttt agggttccga tttagtgctt tacggcacct cgaccccaaa aaacttgatt
     2041 agggtgatgg ttcacgtagt gggccatcgc cctgatagac ggtttttcgc cctttgacgt
     2101 tggagtccac gttctttaat agtggactct tgttccaaac tggaacaaca ctcaacccta
     2161 tctcggtcta ttcttttgat ttataaggga ttttgccgat ttcggcctat tggttaaaaa
     2221 atgagctgat ttaacaaaaa tttaacgcga attttaacaa aatattaacg cttacaattt
     2281 aggtggcact tttcggggaa atgtgcgcgg aacccctatt tgtttatttt tctaaataca
     2341 ttcaaatatg tatccgctca tgaattaatt cttagaaaaa ctcatcgagc atcaaatgaa
     2401 actgcaattt attcatatca ggattatcaa taccatattt ttgaaaaagc cgtttctgta
     2461 atgaaggaga aaactcaccg aggcagttcc ataggatggc aagatcctgg tatcggtctg
     2521 cgattccgac tcgtccaaca tcaatacaac ctattaattt cccctcgtca aaaataaggt
     2581 tatcaagtga gaaatcacca tgagtgacga ctgaatccgg tgagaatggc aaaagtttat
     2641 gcatttcttt ccagacttgt tcaacaggcc agccattacg ctcgtcatca aaatcactcg
     2701 catcaaccaa accgttattc attcgtgatt gcgcctgagc gagacgaaat acgcgatcgc
     2761 tgttaaaagg acaattacaa acaggaatcg aatgcaaccg gcgcaggaac actgccagcg
     2821 catcaacaat attttcacct gaatcaggat attcttctaa tacctggaat gctgttttcc
     2881 cggggatcgc agtggtgagt aaccatgcat catcaggagt acggataaaa tgcttgatgg
     2941 tcggaagagg cataaattcc gtcagccagt ttagtctgac catctcatct gtaacatcat
     3001 tggcaacgct acctttgcca tgtttcagaa acaactctgg cgcatcgggc ttcccataca
     3061 atcgatagat tgtcgcacct gattgcccga cattatcgcg agcccattta tacccatata
     3121 aatcagcatc catgttggaa tttaatcgcg gcctagagca agacgtttcc cgttgaatat
     3181 ggctcataac accccttgta ttactgttta tgtaagcaga cagttttatt gttcatgacc
     3241 aaaatccctt aacgtgagtt ttcgttccac tgagcgtcag accccgtaga aaagatcaaa
     3301 ggatcttctt gagatccttt ttttctgcgc gtaatctgct gcttgcaaac aaaaaaacca
     3361 ccgctaccag cggtggtttg tttgccggat caagagctac caactctttt tccgaaggta
     3421 actggcttca gcagagcgca gataccaaat actgtccttc tagtgtagcc gtagttaggc
     3481 caccacttca agaactctgt agcaccgcct acatacctcg ctctgctaat cctgttacca
     3541 gtggctgctg ccagtggcga taagtcgtgt cttaccgggt tggactcaag acgatagtta
     3601 ccggataagg cgcagcggtc gggctgaacg gggggttcgt gcacacagcc cagcttggag
     3661 cgaacgacct acaccgaact gagataccta cagcgtgagc tatgagaaag cgccacgctt
     3721 cccgaaggga gaaaggcgga caggtatccg gtaagcggca gggtcggaac aggagagcgc
     3781 acgagggagc ttccaggggg aaacgcctgg tatctttata gtcctgtcgg gtttcgccac
     3841 ctctgacttg agcgtcgatt tttgtgatgc tcgtcagggg ggcggagcct atggaaaaac
     3901 gccagcaacg cggccttttt acggttcctg gccttttgct ggccttttgc tcacatgttc
     3961 tttcctgcgt tatcccctga ttctgtggat aaccgtatta ccgcctttga gtgagctgat
     4021 accgctcgcc gcagccgaac gaccgagcgc agcgagtcag tgagcgagga agcggaagag
     4081 cgcctgatgc ggtattttct ccttacgcat ctgtgcggta tttcacaccg caatggtgca
     4141 ctctcagtac aatctgctct gatgccgcat agttaagcca gtatacactc cgctatcgct
     4201 acgtgactgg gtcatggctg cgccccgaca cccgccaaca cccgctgacg cgccctgacg
     4261 ggcttgtctg ctcccggcat ccgcttacag acaagctgtg accgtctccg ggagctgcat
     4321 gtgtcagagg ttttcaccgt catcaccgaa acgcgcgagg cagctgcggt aaagctcatc
     4381 agcgtggtcg tgaagcgatt cacagatgtc tgcctgttca tccgcgtcca gctcgttgag
     4441 tttctccaga agcgttaatg tctggcttct gataaagcgg gccatgttaa gggcggtttt
     4501 ttcctgtttg gtcactgatg cctccgtgta agggggattt ctgttcatgg gggtaatgat
     4561 accgatgaaa cgagagagga tgctcacgat acgggttact gatgatgaac atgcccggtt
     4621 actggaacgt tgtgagggta aacaactggc ggtatggatg cggcgggacc agagaaaaat
     4681 cactcagggt caatgccagc gcttcgttaa tacagatgta ggtgttccac agggtagcca
     4741 gcagcatcct gcgatgcaga tccggaacat aatggtgcag ggcgctgact tccgcgtttc
     4801 cagactttac gaaacacgga aaccgaagac cattcatgtt gttgctcagg tcgcagacgt
     4861 tttgcagcag cagtcgcttc acgttcgctc gcgtatcggt gattcattct gctaaccagt
     4921 aaggcaaccc cgccagccta gccgggtcct caacgacagg agcacgatca tgcgcacccg
     4981 tggggccgcc atgccggcga taatggcctg cttctcgccg aaacgtttgg tggcgggacc
     5041 agtgacgaag gcttgagcga gggcgtgcaa gattccgaat accgcaagcg acaggccgat
     5101 catcgtcgcg ctccagcgaa agcggtcctc gccgaaaatg acccagagcg ctgccggcac
     5161 ctgtcctacg agttgcatga taaagaagac agtcataagt gcggcgacga tagtcatgcc
     5221 ccgcgcccac cggaaggagc tgactgggtt gaaggctctc aagggcatcg gtcgagatcc
     5281 cggtgcctaa tgagtgagct aacttacatt aattgcgttg cgctcactgc ccgctttcca
     5341 gtcgggaaac ctgtcgtgcc agctgcatta atgaatcggc caacgcgcgg ggagaggcgg
     5401 tttgcgtatt gggcgccagg gtggtttttc ttttcaccag tgagacgggc aacagctgat
     5461 tgcccttcac cgcctggccc tgagagagtt gcagcaagcg gtccacgctg gtttgcccca
     5521 gcaggcgaaa atcctgtttg atggtggtta acggcgggat ataacatgag ctgtcttcgg
     5581 tatcgtcgta tcccactacc gagatatccg caccaacgcg cagcccggac tcggtaatgg
     5641 cgcgcattgc gcccagcgcc atctgatcgt tggcaaccag catcgcagtg ggaacgatgc
     5701 cctcattcag catttgcatg gtttgttgaa aaccggacat ggcactccag tcgccttccc
     5761 gttccgctat cggctgaatt tgattgcgag tgagatattt atgccagcca gccagacgca
     5821 gacgcgccga gacagaactt aatgggcccg ctaacagcgc gatttgctgg tgacccaatg
     5881 cgaccagatg ctccacgccc agtcgcgtac cgtcttcatg ggagaaaata atactgttga
     5941 tgggtgtctg gtcagagaca tcaagaaata acgccggaac attagtgcag gcagcttcca
     6001 cagcaatggc atcctggtca tccagcggat agttaatgat cagcccactg acgcgttgcg
     6061 cgagaagatt gtgcaccgcc gctttacagg cttcgacgcc gcttcgttct accatcgaca
     6121 ccaccacgct ggcacccagt tgatcggcgc gagatttaat cgccgcgaca atttgcgacg
     6181 gcgcgtgcag ggccagactg gaggtggcaa cgccaatcag caacgactgt ttgcccgcca
     6241 gttgttgtgc cacgcggttg ggaatgtaat tcagctccgc catcgccgct tccacttttt
     6301 cccgcgtttt cgcagaaacg tggctggcct ggttcaccac gcgggaaacg gtctgataag
     6361 agacaccggc atactctgcg acatcgtata acgttactgg tttcacattc accaccctga
     6421 attgactctc ttccgggcgc tatcatgcca taccgcgaaa ggttttgcgc cattcgatgg
     6481 tgtccgggat ctcgacgctc tcccttatgc gactcctgca ttaggaagca gcccagtagt
     6541 aggttgaggc cgttgagcac cgccgccgca aggaatggtg catgcaagga gatggcgccc
     6601 aacagtcccc cggccacggg gcctgccacc atacccacgc cgaaacaagc gctcatgagc
     6661 ccgaagtggc gagcccgatc ttccccatcg gtgatgtcgg cgatataggc gccagcaacc
     6721 gcacctgtgg cgccggtgat gccggccacg atgcgtccgg cgtagaggat cgagatctcg
     6781 atcccgcgaa at
//
