LOCUS       sequence-129110-        6100 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 6100)
  AUTHORS   Lai L, Yokota H, Hung LW, Kim R, Kim SH
  TITLE     Crystal structure of archaeal RNase HII: a homologue of human major 
            RNase H.
  JOURNAL   Structure. 2000 Aug 15. 8(8):897-904.
  PUBMED    10997908
REFERENCE   2  (bases 1 to 6100)
  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: "Structure. 
            2000 Aug 15. 8(8):897-904."
COMMENT     Sequence Label: pET21a.MJ0135
FEATURES             Location/Qualifiers
     source          1..6100
                     /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"
     RBS             904..926
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     protein_bind    941..965
                     /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(965..984)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(966..984)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(1068..1087)
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     promoter        1293..1370
                     /gene="lacI"
                     /label=lacI promoter
     CDS             1371..2453
                     /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(1390..1409)
                     /label=LacI-R
                     /note="LacI, reverse primer"
     CDS             3262..3453
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     primer_bind     complement(3447..3469)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     misc_feature    3555..3697
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     3610..3629
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     complement(3712..3729)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3883..4471)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3963..3982)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4642..5433,5434..5502))
                     /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     5265..5284
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5503..5607)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      complement(5634..6089)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(5771..5792)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     5983..6002
                     /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 tcctcactct ataattagct ttgtctgttt tgatttatcc
      241 aatatccttt tacatgtttt ccagtgagtt cttgcaatat caggaagttt tttatgcttt
      301 ttaaagtaat cttcaagaaa ctttatagtt tttgggtctg atggataacc gctcccaata
      361 tctccataaa tttttttgta gtagtctatt atctcatccc gttctgcctt tgctattatt
      421 gaggcagctg aaacaacagg atacttagca tctgccttat gttcagctat aatttttatg
      481 tttaaatttc tttctttaat tatatcctct atcttatctt taaaactatc ctcaaatttt
      541 ttagtgtttg tactgcatgc atcaatataa atttcaattt catcatccct tatattcaac
      601 ttttctatca aatttttagc aaccttggaa aaggcgttaa tttcaatatc atttaagttt
      661 attgagttca ttaattggtt tatttcctct gcttctaaga tacgtttttc tacataaccc
      721 aaattttcaa gtaacttttt tagataagcc cttttatttt tagttagctc cttactatct
      781 ttaactccca atttctttaa ttcctcttct ctctcttttt caattgcaaa agcacaaaca
      841 accattggac ccaaaacagg ccctcttcca gcttcatcaa taccaataat tatcatatgt
      901 atatctcctt cttaaagtta aacaaaatta tttctagagg ggaattgtta tccgctcaca
      961 attcccctat agtgagtcgt attaatttcg cgggatcgag atctcgatcc tctacgccgg
     1021 acgcatcgtg gccggcatca ccggcgccac aggtgcggtt gctggcgcct atatcgccga
     1081 catcaccgat ggggaagatc gggctcgcca cttcgggctc atgagcgctt gtttcggcgt
     1141 gggtatggtg gcaggccccg tggccggggg actgttgggc gccatctcct tgcatgcacc
     1201 attccttgcg gcggcggtgc tcaacggcct caacctacta ctgggctgct tcctaatgca
     1261 ggagtcgcat aagggagagc gtcgagatcc cggacaccat cgaatggcgc aaaacctttc
     1321 gcggtatggc atgatagcgc ccggaagaga gtcaattcag ggtggtgaat gtgaaaccag
     1381 taacgttata cgatgtcgca gagtatgccg gtgtctctta tcagaccgtt tcccgcgtgg
     1441 tgaaccaggc cagccacgtt tctgcgaaaa cgcgggaaaa agtggaagcg gcgatggcgg
     1501 agctgaatta cattcccaac cgcgtggcac aacaactggc gggcaaacag tcgttgctga
     1561 ttggcgttgc cacctccagt ctggccctgc acgcgccgtc gcaaattgtc gcggcgatta
     1621 aatctcgcgc cgatcaactg ggtgccagcg tggtggtgtc gatggtagaa cgaagcggcg
     1681 tcgaagcctg taaagcggcg gtgcacaatc ttctcgcgca acgcgtcagt gggctgatca
     1741 ttaactatcc gctggatgac caggatgcca ttgctgtgga agctgcctgc actaatgttc
     1801 cggcgttatt tcttgatgtc tctgaccaga cacccatcaa cagtattatt ttctcccatg
     1861 aagacggtac gcgactgggc gtggagcatc tggtcgcatt gggtcaccag caaatcgcgc
     1921 tgttagcggg cccattaagt tctgtctcgg cgcgtctgcg tctggctggc tggcataaat
     1981 atctcactcg caatcaaatt cagccgatag cggaacggga aggcgactgg agtgccatgt
     2041 ccggttttca acaaaccatg caaatgctga atgagggcat cgttcccact gcgatgctgg
     2101 ttgccaacga tcagatggcg ctgggcgcaa tgcgcgccat taccgagtcc gggctgcgcg
     2161 ttggtgcgga tatctcggta gtgggatacg acgataccga agacagctca tgttatatcc
     2221 cgccgttaac caccatcaaa caggattttc gcctgctggg gcaaaccagc gtggaccgct
     2281 tgctgcaact ctctcagggc caggcggtga agggcaatca gctgttgccc gtctcactgg
     2341 tgaaaagaaa aaccaccctg gcgcccaata cgcaaaccgc ctctccccgc gcgttggccg
     2401 attcattaat gcagctggca cgacaggttt cccgactgga aagcgggcag tgagcgcaac
     2461 gcaattaatg taagttagct cactcattag gcaccgggat ctcgaccgat gcccttgaga
     2521 gccttcaacc cagtcagctc cttccggtgg gcgcggggca tgactatcgt cgccgcactt
     2581 atgactgtct tctttatcat gcaactcgta ggacaggtgc cggcagcgct ctgggtcatt
     2641 ttcggcgagg accgctttcg ctggagcgcg acgatgatcg gcctgtcgct tgcggtattc
     2701 ggaatcttgc acgccctcgc tcaagccttc gtcactggtc ccgccaccaa acgtttcggc
     2761 gagaagcagg ccattatcgc cggcatggcg gccccacggg tgcgcatgat cgtgctcctg
     2821 tcgttgagga cccggctagg ctggcggggt tgccttactg gttagcagaa tgaatcaccg
     2881 atacgcgagc gaacgtgaag cgactgctgc tgcaaaacgt ctgcgacctg agcaacaaca
     2941 tgaatggtct tcggtttccg tgtttcgtaa agtctggaaa cgcggaagtc agcgccctgc
     3001 accattatgt tccggatctg catcgcagga tgctgctggc taccctgtgg aacacctaca
     3061 tctgtattaa cgaagcgctg gcattgaccc tgagtgattt ttctctggtc ccgccgcatc
     3121 cataccgcca gttgtttacc ctcacaacgt tccagtaacc gggcatgttc atcatcagta
     3181 acccgtatcg tgagcatcct ctctcgtttc atcggtatca ttacccccat gaacagaaat
     3241 cccccttaca cggaggcatc agtgaccaaa caggaaaaaa ccgcccttaa catggcccgc
     3301 tttatcagaa gccagacatt aacgcttctg gagaaactca acgagctgga cgcggatgaa
     3361 caggcagaca tctgtgaatc gcttcacgac cacgctgatg agctttaccg cagctgcctc
     3421 gcgcgtttcg gtgatgacgg tgaaaacctc tgacacatgc agctcccgga gacggtcaca
     3481 gcttgtctgt aagcggatgc cgggagcaga caagcccgtc agggcgcgtc agcgggtgtt
     3541 ggcgggtgtc ggggcgcagc catgacccag tcacgtagcg atagcggagt gtatactggc
     3601 ttaactatgc ggcatcagag cagattgtac tgagagtgca ccatatatgc ggtgtgaaat
     3661 accgcacaga tgcgtaagga gaaaataccg catcaggcgc tcttccgctt cctcgctcac
     3721 tgactcgctg cgctcggtcg ttcggctgcg gcgagcggta tcagctcact caaaggcggt
     3781 aatacggtta tccacagaat caggggataa cgcaggaaag aacatgtgag caaaaggcca
     3841 gcaaaaggcc aggaaccgta aaaaggccgc gttgctggcg tttttccata ggctccgccc
     3901 ccctgacgag catcacaaaa atcgacgctc aagtcagagg tggcgaaacc cgacaggact
     3961 ataaagatac caggcgtttc cccctggaag ctccctcgtg cgctctcctg ttccgaccct
     4021 gccgcttacc ggatacctgt ccgcctttct cccttcggga agcgtggcgc tttctcatag
     4081 ctcacgctgt aggtatctca gttcggtgta ggtcgttcgc tccaagctgg gctgtgtgca
     4141 cgaacccccc gttcagcccg accgctgcgc cttatccggt aactatcgtc ttgagtccaa
     4201 cccggtaaga cacgacttat cgccactggc agcagccact ggtaacagga ttagcagagc
     4261 gaggtatgta ggcggtgcta cagagttctt gaagtggtgg cctaactacg gctacactag
     4321 aaggacagta tttggtatct gcgctctgct gaagccagtt accttcggaa aaagagttgg
     4381 tagctcttga tccggcaaac aaaccaccgc tggtagcggt ggtttttttg tttgcaagca
     4441 gcagattacg cgcagaaaaa aaggatctca agaagatcct ttgatctttt ctacggggtc
     4501 tgacgctcag tggaacgaaa actcacgtta agggattttg gtcatgagat tatcaaaaag
     4561 gatcttcacc tagatccttt taaattaaaa atgaagtttt aaatcaatct aaagtatata
     4621 tgagtaaact tggtctgaca gttaccaatg cttaatcagt gaggcaccta tctcagcgat
     4681 ctgtctattt cgttcatcca tagttgcctg actccccgtc gtgtagataa ctacgatacg
     4741 ggagggctta ccatctggcc ccagtgctgc aatgataccg cgagacccac gctcaccggc
     4801 tccagattta tcagcaataa accagccagc cggaagggcc gagcgcagaa gtggtcctgc
     4861 aactttatcc gcctccatcc agtctattaa ttgttgccgg gaagctagag taagtagttc
     4921 gccagttaat agtttgcgca acgttgttgc cattgctgca ggcatcgtgg tgtcacgctc
     4981 gtcgtttggt atggcttcat tcagctccgg ttcccaacga tcaaggcgag ttacatgatc
     5041 ccccatgttg tgcaaaaaag cggttagctc cttcggtcct ccgatcgttg tcagaagtaa
     5101 gttggccgca gtgttatcac tcatggttat ggcagcactg cataattctc ttactgtcat
     5161 gccatccgta agatgctttt ctgtgactgg tgagtactca accaagtcat tctgagaata
     5221 gtgtatgcgg cgaccgagtt gctcttgccc ggcgtcaata cgggataata ccgcgccaca
     5281 tagcagaact ttaaaagtgc tcatcattgg aaaacgttct tcggggcgaa aactctcaag
     5341 gatcttaccg ctgttgagat ccagttcgat gtaacccact cgtgcaccca actgatcttc
     5401 agcatctttt actttcacca gcgtttctgg gtgagcaaaa acaggaaggc aaaatgccgc
     5461 aaaaaaggga ataagggcga cacggaaatg ttgaatactc atactcttcc tttttcaata
     5521 ttattgaagc atttatcagg gttattgtct catgagcgga tacatatttg aatgtattta
     5581 gaaaaataaa caaatagggg ttccgcgcac atttccccga aaagtgccac ctgaaattgt
     5641 aaacgttaat attttgttaa aattcgcgtt aaatttttgt taaatcagct cattttttaa
     5701 ccaataggcc gaaatcggca aaatccctta taaatcaaaa gaatagaccg agatagggtt
     5761 gagtgttgtt ccagtttgga acaagagtcc actattaaag aacgtggact ccaacgtcaa
     5821 agggcgaaaa accgtctatc agggcgatgg cccactacgt gaaccatcac cctaatcaag
     5881 ttttttgggg tcgaggtgcc gtaaagcact aaatcggaac cctaaaggga gcccccgatt
     5941 tagagcttga cggggaaagc cggcgaacgt ggcgagaaag gaagggaaga aagcgaaagg
     6001 agcgggcgct agggcgctgg caagtgtagc ggtcacgctg cgcgtaacca ccacacccgc
     6061 cgcgcttaat gcgccgctac agggcgcgtc ccattcgcca
//
