LOCUS       sequence-129516-        5773 bp    DNA     circular SYN 14-APR-2025
DEFINITION  synthetic circular DNA
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5773)
  TITLE     various new plasmids
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 5773)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Apr 14, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: 
            "Unpublished"
COMMENT     Sequence Label: Pin1
FEATURES             Location/Qualifiers
     source          1..5773
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(18..37)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     178..200
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(194..385)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     CDS             complement(1194..2276)
                     /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     2238..2257
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(2277..2354)
                     /gene="lacI"
                     /label=lacI promoter
     primer_bind     2560..2579
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     primer_bind     2667..2686
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        2667..2685
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    2686..2710
                     /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             2725..2747
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             2758..2775
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             2800..2820
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQS"
     CDS             3376..3393
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(3446..3464)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      3460..3507
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     primer_bind     complement(3631..3650)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        3817..3920
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(3921..3989,3990..4781)
                     /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(4139..4158)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      4952..5540
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5441..5460
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     5694..5711
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    join(5726..5773,1..92)
                     /label=bom
                     /note="basis of mobility region from pBR322"
ORIGIN
        1 caatggtgca ctctcagtac aatctgctct gatgccgcat agttaagcca gtatacactc
       61 cgctatcgct acgtgactgg gtcatggctg cgccccgaca cccgccaaca cccgctgacg
      121 cgccctgacg ggcttgtctg ctcccggcat ccgcttacag acaagctgtg accgtctccg
      181 ggagctgcat gtgtcagagg ttttcaccgt catcaccgaa acgcgcgagg cagctgcggt
      241 aaagctcatc agcgtggtcg tgaagcgatt cacagatgtc tgcctgttca tccgcgtcca
      301 gctcgttgag tttctccaga agcgttaatg tctggcttct gataaagcgg gccatgttaa
      361 gggcggtttt ttcctgtttg gtcactgatg cctccgtgta agggggattt ctgttcatgg
      421 gggtaatgat accgatgaaa cgagagagga tgctcacgat acgggttact gatgatgaac
      481 atgcccggtt actggaacgt tgtgagggta aacaactggc ggtatggatg cggcgggacc
      541 agagaaaaat cactcagggt caatgccagc gcttcgttaa tacagatgta ggtgttccac
      601 agggtagcca gcagcatcct gcgatgcaga tccggaacat aatggtgcag ggcgctgact
      661 tccgcgtttc cagactttac gaaacacgga aaccgaagac cattcatgtt gttgctcagg
      721 tcgcagacgt tttgcagcag cagtcgcttc acgttcgctc gcgtatcggt gattcattct
      781 gctaaccagt aaggcaaccc cgccagccta gccgggtcct caacgacagg agcacgatca
      841 tgcgcacccg tggggccgcc atgccggcga taatggcctg cttctcgccg aaacgtttgg
      901 tggcgggacc agtgacgaag gcttgagcga gggcgtgcaa gattccgaat accgcaagcg
      961 acaggccgat catcgtcgcg ctccagcgaa agcggtcctc gccgaaaatg acccagagcg
     1021 ctgccggcac ctgtcctacg agttgcatga taaagaagac agtcataagt gcggcgacga
     1081 tagtcatgcc ccgcgcccac cggaaggagc tgactgggtt gaaggctctc aagggcatcg
     1141 gtcgagatcc cggtgcctaa tgagtgagct aacttacatt aattgcgttg cgctcactgc
     1201 ccgctttcca gtcgggaaac ctgtcgtgcc agctgcatta atgaatcggc caacgcgcgg
     1261 ggagaggcgg tttgcgtatt gggcgccagg gtggtttttc ttttcaccag tgagacgggc
     1321 aacagctgat tgcccttcac cgcctggccc tgagagagtt gcagcaagcg gtccacgctg
     1381 gtttgcccca gcaggcgaaa atcctgtttg atggtggtta acggcgggat ataacatgag
     1441 ctgtcttcgg tatcgtcgta tcccactacc gagatatccg caccaacgcg cagcccggac
     1501 tcggtaatgg cgcgcattgc gcccagcgcc atctgatcgt tggcaaccag catcgcagtg
     1561 ggaacgatgc cctcattcag catttgcatg gtttgttgaa aaccggacat ggcactccag
     1621 tcgccttccc gttccgctat cggctgaatt tgattgcgag tgagatattt atgccagcca
     1681 gccagacgca gacgcgccga gacagaactt aatgggcccg ctaacagcgc gatttgctgg
     1741 tgacccaatg cgaccagatg ctccacgccc agtcgcgtac cgtcttcatg ggagaaaata
     1801 atactgttga tgggtgtctg gtcagagaca tcaagaaata acgccggaac attagtgcag
     1861 gcagcttcca cagcaatggc atcctggtca tccagcggat agttaatgat cagcccactg
     1921 acgcgttgcg cgagaagatt gtgcaccgcc gctttacagg cttcgacgcc gcttcgttct
     1981 accatcgaca ccaccacgct ggcacccagt tgatcggcgc gagatttaat cgccgcgaca
     2041 atttgcgacg gcgcgtgcag ggccagactg gaggtggcaa cgccaatcag caacgactgt
     2101 ttgcccgcca gttgttgtgc cacgcggttg ggaatgtaat tcagctccgc catcgccgct
     2161 tccacttttt cccgcgtttt cgcagaaacg tggctggcct ggttcaccac gcgggaaacg
     2221 gtctgataag agacaccggc atactctgcg acatcgtata acgttactgg tttcacattc
     2281 accaccctga attgactctc ttccgggcgc tatcatgcca taccgcgaaa ggttttgcgc
     2341 cattcgatgg tgtccgggat ctcgacgctc tcccttatgc gactcctgca ttaggaagca
     2401 gcccagtagt aggttgaggc cgttgagcac cgccgccgca aggaatggtg catgcaagga
     2461 gatggcgccc aacagtcccc cggccacggg gcctgccacc atacccacgc cgaaacaagc
     2521 gctcatgagc ccgaagtggc gagcccgatc ttccccatcg gtgatgtcgg cgatataggc
     2581 gccagcaacc gcacctgtgg cgccggtgat gccggccacg atgcgtccgg cgtagaggat
     2641 cgagatcgat ctcgatcccg cgaaattaat acgactcact ataggggaat tgtgagcgga
     2701 taacaattcc cctctagaaa taattttgtt taactttaag aaggagatat acatatgcac
     2761 catcatcatc atcattcttc tggtgtagat ctgggtaccg agaacctgta cttccaatcc
     2821 aatgcggacg aggagaagct gccgcccggc tgggagaagc gcatgagccg cagctcaggc
     2881 cgagtgtact acttcaacca catcactaac gccagccagt gggagcggcc cagcggcaac
     2941 agcagcagtg gtggcaaaaa cgggcagggg gagcctgcca gggtccgctg ctcgcacctg
     3001 ctggtgaagc acagccagtc acggcggccc tcgtcctggc ggcaggagaa gatcacccgg
     3061 accaaggagg aggccctgga gctgatcaac ggctacatcc agaagatcaa gtcgggagag
     3121 gaggactttg agtctctggc ctcacagttc agcgactgca gctcagccaa ggccagggga
     3181 gacctgggtg ccttcagcag aggtcagatg cagaagccat ttgaagacgc ctcgtttgcg
     3241 ctgcggacgg gggagatgag cgggcccgtg ttcacggatt ccggcatcca catcatcctc
     3301 cgcactgagt gacattggaa gtggataacg gatccgaatt cgagctccgt cgacaagctt
     3361 gcggccgcac tcgagcacca ccaccaccac cactgagatc cggctgctaa caaagcccga
     3421 aaggaagctg agttggctgc tgccaccgct gagcaataac tagcataacc ccttggggcc
     3481 tctaaacggg tcttgagggg ttttttgctg aaaggaggaa ctatatccgg attggcgaat
     3541 gggacgcgcc ctgtagcggc gcattaagcg cggcgggtgt ggtggttacg cgcagcgtga
     3601 ccgctacact tgccagcgcc ctagcgcccg ctcctttcgc tttcttccct tcctttctcg
     3661 ccacgttcgc cggctttccc cgtcaagctc taaatcgggg gctcccttta gggttccgat
     3721 ttagtgcttt acggcacctc gaccccaaaa aacttgatta gggtgatggt tcacattaac
     3781 gcttacaatt taggtggcac ttttcgggga aatgtgcgcg gaacccctat ttgtttattt
     3841 ttctaaatac attcaaatat gtatccgctc atgagacaat aaccctgata aatgcttcaa
     3901 taatttgaaa aaggaagagt atgagtattc aacatttccg tgtcgccctt attccctttt
     3961 ttgcggcatt ttgccttcct gtttttgctc acccagaaac gctggtgaaa gtaaaagatg
     4021 ctgaagatca gttgggtgca cgagtgggtt acatcgaact ggatctcaac agcggtaaga
     4081 tccttgagag ttttcgcccc gaagaacgtt ttccaatgat gagcactttt aaagttctgc
     4141 tatgtggcgc ggtattatcc cgtattgacg ccgggcaaga gcaactcggt cgccgcatac
     4201 actattctca gaatgacttg gttgagtact caccagtcac agaaaagcat cttacggatg
     4261 gcatgacagt aagagaatta tgcagtgctg ccataaccat gagtgataac actgcggcca
     4321 acttacttct gacaacgatc ggaggaccga aggagctaac cgcttttttg cacaacatgg
     4381 gggatcatgt aactcgcctt gatcgttggg aaccggagct gaatgaagcc ataccaaacg
     4441 acgagcgtga caccacgatg cctgcagcaa tggcaacaac gttgcgcaaa ctattaactg
     4501 gcgaactact tactctagct tcccggcaac aattaataga ctggatggag gcggataaag
     4561 ttgcaggacc acttctgcgc tcggcccttc cggctggctg gtttattgct gataaatctg
     4621 gagccggtga gcgtgggtct cgcggtatca ttgcagcact ggggccagat ggtaagccct
     4681 cccgtatcgt agttatctac acgacgggga gtcaggcaac tatggatgaa cgaaatagac
     4741 agatcgctga gataggtgcc tcactgatta agcattggta actgtcagac caagtttact
     4801 catatatact ttagattgat ttaaaacttc atttttaatt taaaaggatc taggtgaaga
     4861 tcctttttga taatctcatg accaaaatcc cttaacgtga gttttcgttc cactgagcgt
     4921 cagaccccgt agaaaagatc aaaggatctt cttgagatcc tttttttctg cgcgtaatct
     4981 gctgcttgca aacaaaaaaa ccaccgctac cagcggtggt ttgtttgccg gatcaagagc
     5041 taccaactct ttttccgaag gtaactggct tcagcagagc gcagatacca aatactgtcc
     5101 ttctagtgta gccgtagtta ggccaccact tcaagaactc tgtagcaccg cctacatacc
     5161 tcgctctgct aatcctgtta ccagtggctg ctgccagtgg cgataagtcg tgtcttaccg
     5221 ggttggactc aagacgatag ttaccggata aggcgcagcg gtcgggctga acggggggtt
     5281 cgtgcacaca gcccagcttg gagcgaacga cctacaccga actgagatac ctacagcgtg
     5341 agctatgaga aagcgccacg cttcccgaag ggagaaaggc ggacaggtat ccggtaagcg
     5401 gcagggtcgg aacaggagag cgcacgaggg agcttccagg gggaaacgcc tggtatcttt
     5461 atagtcctgt cgggtttcgc cacctctgac ttgagcgtcg atttttgtga tgctcgtcag
     5521 gggggcggag cctatggaaa aacgccagca acgcggcctt tttacggttc ctggcctttt
     5581 gctggccttt tgctcacatg ttctttcctg cgttatcccc tgattctgtg gataaccgta
     5641 ttaccgcctt tgagtgagct gataccgctc gccgcagccg aacgaccgag cgcagcgagt
     5701 cagtgagcga ggaagcggaa gagcgcctga tgcggtattt tctccttacg catctgtgcg
     5761 gtatttcaca ccg
//
