LOCUS       sequence-307045-        6253 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  synthetic circular DNA
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6253)
  AUTHORS   Schulze-Gahmen U, Pelaschier J, Yokota H, Kim R, Kim SH
  TITLE     Crystal structure of a hypothetical protein, TM841 of Thermotoga 
            maritima, reveals its function as a fatty acid-binding protein.
  JOURNAL   Proteins. 2003 Mar 1. 50(4):526-30.
  PUBMED    12577257
REFERENCE   2  (bases 1 to 6253)
  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: "Proteins. 
            2003 Mar 1. 50(4):526-30."
COMMENT     Sequence Label: pSKB3.TM1468
FEATURES             Location/Qualifiers
     source          1..6253
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     terminator      complement(26..73)
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     primer_bind     69..87
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     CDS             complement(140..157)
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             complement(1116..1136)
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQG"
     CDS             complement(1158..1175)
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     RBS             1194..1216
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     protein_bind    1231..1255
                     /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     complement(1255..1274)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(1256..1274)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(1358..1377)
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     promoter        1583..1660
                     /gene="lacI"
                     /label=lacI promoter
     CDS             1661..2743
                     /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     complement(1680..1699)
                     /label=LacI-R
                     /note="LacI, reverse primer"
     CDS             3552..3743
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     primer_bind     complement(3737..3759)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     misc_feature    3845..3984
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     3900..3919
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     complement(3999..4016)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4170..4758)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4250..4269)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     complement(4869..4888)
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     CDS             4880..5695
                     /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             4880..5695
                     /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(4953..4972)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     rep_origin      complement(5787..6242)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(5924..5945)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     6136..6155
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
ORIGIN
        1 atccggatat agttcctcct ttcagcaaaa aacccctcaa gacccgttta gaggccccaa
       61 ggggttatgc tagttattgc tcagcggtgg cagcagccaa ctcagcttcc tttcgggctt
      121 tgttagcagc cggatctcag tggtggtggt ggtggtgctc gagtgcggcc gcaagcttgt
      181 cgacggagct cgaattcgga tccactagta acggccgcca gtgtgctgga attcgccctt
      241 ggtccttatc tttttcgttc taaaacctcg attccaaacc caaccgtccc gggtcccaca
      301 tgagtcgtga tcacctttcc catcggcgat atgatctcgt cgacaacctc gtaacttttt
      361 ctgagtgtgt tcaaaagctc cacgactcca gcctcgttgt ccgcgtgaac acctatcact
      421 ctcagcttcg agccttcagg agtatcctca cgcagttttt ctatgagggc ttctatcgcc
      481 ttcttgtctc ctctgacctt tctgtaaggg atgagctcac cgttctctat gtgaagacac
      541 actcttatct tgagaagatt tcccacaaat ccctggaact tcgacaccct tcccccctta
      601 acaagataat cgaaattcga aacataaaaa atcgctttga agtccttgtt cttcattctc
      661 tcatcgagtt ttttcagtac ttcctcgatg gtggcaccat tttcgagcat ttcacgagcc
      721 acacgggccg gaaggggtat cgcacccgaa gcgagaagag tatccacaac gtaaacgggt
      781 atgtccacct ctttggatgc gagaacagcg gaattgtaag ttccggagag tttcgatgat
      841 aaagtgagaa ccaggacaac atcgtaatcc tcttctttgt acttcagata tctcttcttg
      901 aaatcctcca cactcggctg ggaagtcttt ggtacactgc ctgcttctct tattctcttg
      961 taaaagttca ttatctcttc cggttctctt tcgtccggct cggatcttcc gtcttcccac
     1021 accacgtaaa gtgggatgct atctatatcg tatttttcca tccaggaaaa aggaacatcg
     1081 gccgtgctgt caacgagaat ctttaccttc atatggccct ggaagtaaag gttttcggtc
     1141 gttgggatat cgtaatcgtg atgatgatga tgatggctgc tgcccatggt atatctcctt
     1201 cttaaagtta aacaaaatta tttctagagg ggaattgtta tccgctcaca attcccctat
     1261 agtgagtcgt attaatttcg cgggatcgag atctcgatcc tctacgccgg acgcatcgtg
     1321 gccggcatca ccggcgccac aggtgcggtt gctggcgcct atatcgccga catcaccgat
     1381 ggggaagatc gggctcgcca cttcgggctc atgagcgctt gtttcggcgt gggtatggtg
     1441 gcaggccccg tggccggggg actgttgggc gccatctcct tgcatgcacc attccttgcg
     1501 gcggcggtgc tcaacggcct caacctacta ctgggctgct tcctaatgca ggagtcgcat
     1561 aagggagagc gtcgagatcc cggacaccat cgaatggcgc aaaacctttc gcggtatggc
     1621 atgatagcgc ccggaagaga gtcaattcag ggtggtgaat gtgaaaccag taacgttata
     1681 cgatgtcgca gagtatgccg gtgtctctta tcagaccgtt tcccgcgtgg tgaaccaggc
     1741 cagccacgtt tctgcgaaaa cgcgggaaaa agtggaagcg gcgatggcgg agctgaatta
     1801 cattcccaac cgcgtggcac aacaactggc gggcaaacag tcgttgctga ttggcgttgc
     1861 cacctccagt ctggccctgc acgcgccgtc gcaaattgtc gcggcgatta aatctcgcgc
     1921 cgatcaactg ggtgccagcg tggtggtgtc gatggtagaa cgaagcggcg tcgaagcctg
     1981 taaagcggcg gtgcacaatc ttctcgcgca acgcgtcagt gggctgatca ttaactatcc
     2041 gctggatgac caggatgcca ttgctgtgga agctgcctgc actaatgttc cggcgttatt
     2101 tcttgatgtc tctgaccaga cacccatcaa cagtattatt ttctcccatg aagacggtac
     2161 gcgactgggc gtggagcatc tggtcgcatt gggtcaccag caaatcgcgc tgttagcggg
     2221 cccattaagt tctgtctcgg cgcgtctgcg tctggctggc tggcataaat atctcactcg
     2281 caatcaaatt cagccgatag cggaacggga aggcgactgg agtgccatgt ccggttttca
     2341 acaaaccatg caaatgctga atgagggcat cgttcccact gcgatgctgg ttgccaacga
     2401 tcagatggcg ctgggcgcaa tgcgcgccat taccgagtcc gggctgcgcg ttggtgcgga
     2461 tatctcggta gtgggatacg acgataccga agacagctca tgttatatcc cgccgttaac
     2521 caccatcaaa caggattttc gcctgctggg gcaaaccagc gtggaccgct tgctgcaact
     2581 ctctcagggc caggcggtga agggcaatca gctgttgccc gtctcactgg tgaaaagaaa
     2641 aaccaccctg gcgcccaata cgcaaaccgc ctctccccgc gcgttggccg attcattaat
     2701 gcagctggca cgacaggttt cccgactgga aagcgggcag tgagcgcaac gcaattaatg
     2761 taagttagct cactcattag gcaccgggat ctcgaccgat gcccttgaga gccttcaacc
     2821 cagtcagctc cttccggtgg gcgcggggca tgactatcgt cgccgcactt atgactgtct
     2881 tctttatcat gcaactcgta ggacaggtgc cggcagcgct ctgggtcatt ttcggcgagg
     2941 accgctttcg ctggagcgcg acgatgatcg gcctgtcgct tgcggtattc ggaatcttgc
     3001 acgccctcgc tcaagccttc gtcactggtc ccgccaccaa acgtttcggc gagaagcagg
     3061 ccattatcgc cggcatggcg gccccacggg tgcgcatgat cgtgctcctg tcgttgagga
     3121 cccggctagg ctggcggggt tgccttactg gttagcagaa tgaatcaccg atacgcgagc
     3181 gaacgtgaag cgactgctgc tgcaaaacgt ctgcgacctg agcaacaaca tgaatggtct
     3241 tcggtttccg tgtttcgtaa agtctggaaa cgcggaagtc agcgccctgc accattatgt
     3301 tccggatctg catcgcagga tgctgctggc taccctgtgg aacacctaca tctgtattaa
     3361 cgaagcgctg gcattgaccc tgagtgattt ttctctggtc ccgccgcatc cataccgcca
     3421 gttgtttacc ctcacaacgt tccagtaacc gggcatgttc atcatcagta acccgtatcg
     3481 tgagcatcct ctctcgtttc atcggtatca ttacccccat gaacagaaat cccccttaca
     3541 cggaggcatc agtgaccaaa caggaaaaaa ccgcccttaa catggcccgc tttatcagaa
     3601 gccagacatt aacgcttctg gagaaactca acgagctgga cgcggatgaa caggcagaca
     3661 tctgtgaatc gcttcacgac cacgctgatg agctttaccg cagctgcctc gcgcgtttcg
     3721 gtgatgacgg tgaaaacctc tgacacatgc agctcccgga gacggtcaca gcttgtctgt
     3781 aagcggatgc cgggagcaga caagcccgtc agggcgcgtc agcgggtgtt ggcgggtgtc
     3841 ggggcgcagc catgacccag tcacgtagcg atagcggagt gtatactggc ttaactatgc
     3901 ggcatcagag cagattgtac tgagagtgca ccattgcggt gtgaaatacc gcacagatgc
     3961 gtaaggagaa aataccgcat caggcgctct tccgcttcct cgctcactga ctcgctgcgc
     4021 tcggtcgttc ggctgcggcg agcggtatca gctcactcaa aggcggtaat acggttatcc
     4081 acagaatcag gggataacgc aggaaagaac atgtgagcaa aaggccagca aaaggccagg
     4141 aaccgtaaaa aggccgcgtt gctggcgttt ttccataggc tccgcccccc tgacgagcat
     4201 cacaaaaatc gacgctcaag tcagaggtgg cgaaacccga caggactata aagataccag
     4261 gcgtttcccc ctggaagctc cctcgtgcgc tctcctgttc cgaccctgcc gcttaccgga
     4321 tacctgtccg cctttctccc ttcgggaagc gtggcgcttt ctcatagctc acgctgtagg
     4381 tatctcagtt cggtgtaggt cgttcgctcc aagctgggct gtgtgcacga accccccgtt
     4441 cagcccgacc gctgcgcctt atccggtaac tatcgtcttg agtccaaccc ggtaagacac
     4501 gacttatcgc cactggcagc agccactggt aacaggatta gcagagcgag gtatgtaggc
     4561 ggtgctacag agttcttgaa gtggtggcct aactacggct acactagaag gacagtattt
     4621 ggtatctgcg ctctgctgaa gccagttacc ttcggaaaaa gagttggtag ctcttgatcc
     4681 ggcaaacaaa ccaccgctgg tagcggtggt ttttttgttt gcaagcagca gattacgcgc
     4741 agaaaaaaag gatctcaaga agatcctttg atcttttcta cggggtctga cgctcagtgg
     4801 aacgaaaact cacgttaagg gattttggtc atgaacaata aaactgtctg cttacataaa
     4861 cagtaataca aggggtgtta tgagccatat tcaacgggaa acgtcttgct ctaggccgcg
     4921 attaaattcc aacatggatg ctgatttata tgggtataaa tgggctcgcg ataatgtcgg
     4981 gcaatcaggt gcgacaatct atcgattgta tgggaagccc gatgcgccag agttgtttct
     5041 gaaacatggc aaaggtagcg ttgccaatga tgttacagat gagatggtca gactaaactg
     5101 gctgacggaa tttatgcctc ttccgaccat caagcatttt atccgtactc ctgatgatgc
     5161 atggttactc accactgcga tccccgggaa aacagcattc caggtattag aagaatatcc
     5221 tgattcaggt gaaaatattg ttgatgcgct ggcagtgttc ctgcgccggt tgcattcgat
     5281 tcctgtttgt aattgtcctt ttaacagcga tcgcgtattt cgtctcgctc aggcgcaatc
     5341 acgaatgaat aacggtttgg ttgatgcgag tgattttgat gacgagcgta atggctggcc
     5401 tgttgaacaa gtctggaaag aaatgcataa acttttgcca ttctcaccgg attcagtcgt
     5461 cactcatggt gatttctcac ttgataacct tatttttgac gaggggaaat taataggttg
     5521 tattgatgtt ggacgagtcg gaatcgcaga ccgataccag gatcttgcca tcctatggaa
     5581 ctgcctcggt gagttttctc cttcattaca gaaacggctt tttcaaaaat atggtattga
     5641 taatcctgat atgaataaat tgcagtttca tttgatgctc gatgagtttt tctaagaatt
     5701 aattcatgag cggatacata tttgaatgta tttagaaaaa taaacaaata ggggttccgc
     5761 gcacatttcc ccgaaaagtg ccacctaaat tgtaagcgtt aatattttgt taaaattcgc
     5821 gttaaatttt tgttaaatca gctcattttt taaccaatag gccgaaatcg gcaaaatccc
     5881 ttataaatca aaagaataga ccgagatagg gttgagtgtt gttccagttt ggaacaagag
     5941 tccactatta aagaacgtgg actccaacgt caaagggcga aaaaccgtct atcagggcga
     6001 tggcccacta cgtgaaccat caccctaatc aagttttttg gggtcgaggt gccgtaaagc
     6061 actaaatcgg aaccctaaag ggagcccccg atttagagct tgacggggaa agccggcgaa
     6121 cgtggcgaga aaggaaggga agaaagcgaa aggagcgggc gctagggcgc tggcaagtgt
     6181 agcggtcacg ctgcgcgtaa ccaccacacc cgccgcgctt aatgcgccgc tacagggcgc
     6241 gtcccattcg cca
//
