LOCUS       sequence-6931-ex        5871 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 5871)
  TITLE     Cross lab plasmids
REFERENCE   2  (bases 1 to 5871)
  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"
COMMENT     Sequence Label: pRS428
FEATURES             Location/Qualifiers
     source          1..5871
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(29..51)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     151..170
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     gene            complement(249..1378)
                     /label=natMX6
                     /note="yeast selectable marker conferring nourseothricin 
                     resistance"
     terminator      complement(249..446)
                     /label=TEF terminator
                     /note="Ashbya gossypii TEF terminator"
     CDS             complement(462..1034)
                     /codon_start=1
                     /gene="Streptomyces noursei nat1"
                     /product="nourseothricin acetyltransferase"
                     /label=NrsR
                     /note="confers resistance to nourseothricin"
                     /translation="MTTLDDTAYRYRTSVPGDAEAIEALDGSFTTDTVFRVTATGDGFT
                     LREVPVDPPLTKVFPDDESDDESDDGEDGDPDSRTFVAYGDDGDLAGFVVISYSAWNRR
                     LTVEDIEVAPEHRGHGVGRALMGLATEFAGERGAGHLWLEVTNVNAPAIHAYRRMGFTL
                     CGLDTALYDGTASDGERQALYMSMPCP"
     promoter        complement(1035..1378)
                     /label=TEF promoter
                     /note="Ashbya gossypii TEF promoter"
     rep_origin      complement(1495..1950)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1632..1653)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     CDS             complement(1733..2311)
                     /codon_start=1
                     /gene="lacZ fragment"
                     /product="LacZ-alpha fragment of beta-galactosidase"
                     /label=lacZ-alpha
                     /translation="MTMITPSAQLTLTKGNKSWSSTAVAAALELVDPPGCRNSISSLSI
                     PSTSRGGPVPNSPYSESYYARSLAVVLQRRDWENPGVTQLNRLAAHPPFASWRNSEEAR
                     TDRPSQQLRSLNGEWRDAPCSGALSAAGVVVTRSVTATLASALAPAPFAFFPSFLATFA
                     GFPRQALNRGLPLGFRFSALRHLDPKKLD"
     primer_bind     1844..1863
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2080..2102
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     2094..2111
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     2095..2111
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     2121..2140
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        2121..2139
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     misc_feature    2148..2255
                     /label=MCS
                     /note="pBluescript multiple cloning site"
     primer_bind     2164..2180
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     primer_bind     2165..2181
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2215..2231)
                     /label=SK primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2215..2231)
                     /label=pBluescriptSK
                     /note="For pBluescript vector"
     primer_bind     complement(2268..2288)
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        complement(2268..2286)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(2307..2323)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2307..2323)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(2320..2342)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    2331..2347
                     /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)."
     promoter        complement(join(2355..2361,2362..2379,2380..2385))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    2400..2421
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(2538..2555)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(2709..3297)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2789..2808)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(3468..4259,4260..4328))
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     4091..4110
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(4329..4433)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      4460..5802
                     /label=2u ori
                     /note="yeast 2u plasmid origin of replication"
     protein_bind    complement(5423..5470)
                     /label=FRT
                     /bound_moiety="FLP recombinase from the Saccharomyces 
                     cerevisiae 2u plasmid"
                     /note="FLP-mediated recombination occurs in the 8-bp core 
                     sequence TCTAGAAA (Turan and Bode, 2011)."
     primer_bind     5844..5862
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
ORIGIN
        1 tcgcgcgttt cggtgatgac ggtgaaaacc tctgacacat gcagctcccg gagacggtca
       61 cagcttgtct gtaagcggat gccgggagca gacaagcccg tcagggcgcg tcagcgggtg
      121 ttggcgggtg tcggggctgg cttaactatg cggcatcaga gcagattgta ctgagagtgc
      181 accataatca tcgatgaatt cgagctcgtt ttcgacactg gatggcggcg ttagtatcga
      241 atcgacagca gtatagcgac cagcattcac atacgattga cgcatgatat tactttctgc
      301 gcacttaact tcgcatctgg gcagatgatg tcgaggcgaa aaaaaatata aatcacgcta
      361 acatttgatt aaaatagaac aactacaata taaaaaaact atacaaatga caagttcttg
      421 aaaacaagaa tctttttatt gtcagtactg attagaaaaa cctaggggca gggcatgctc
      481 atgtagagcg cctgccgctc gccgtccgag gcggtgccgt cgtacagggc ggtgtccagg
      541 ccgcagaggg tgaaccccat ccgccggtac gcgtggatcg ccggtgcgtt gacgttggtg
      601 acctccagcc agaggtgccc ggcgccccgc tcgccggcga actccgtcgc gagccccatc
      661 aacgcgcgcc cgaccccgtg cccccggtgc tccggggcga cctcgatgtc ctcgacggtc
      721 agccggcggt tccacgccga gtacgagatg accacgaagc ccgccaggtc gccgtcgtcc
      781 ccgtacgcga cgaacgtccg ggagtccggg tcgccgtcct ccccgtcgtc cgattcgtcg
      841 tccgattcgt cgtcggggaa caccttggtc aggggcgggt ccaccggcac ctcccgcagg
      901 gtgaagccgt ccccggtggc ggtgacgcgg aagacggtgt cggtggtgaa ggacccatcc
      961 agtgcctcga tggcctcggc gtcccccggg acactggtgc ggtaccggta agccgtgtcg
     1021 tcaagagtgg tcatggttgt ttatgttcgg atgtgatgtg agaactgtat cctagcaaga
     1081 ttttaaaagg aagtatatga aagaagaacc tcagtggcaa atcctaacct tttatatttc
     1141 tctacagggg cgcggcgtgg ggacaattca acgcgtctgt gaggggagcg tttccctgct
     1201 cgcaggtctg cagcgaggag ccgtaatttt tgcttcgcgc cgtgcggcca tcaaaatgta
     1261 tggatgcaaa tgattataca tggggatgta tgggctaaat gtacgggcga cagtcacatc
     1321 atgcccctga gctgcgcacg tcaagactgt caaggagggt attctgggcc tccatgtcgc
     1381 tggccgggtg acccggcggg gacgaggcaa gctaaacaga tctggcgcgc cttaattaac
     1441 cctatgcggt gtgaaatacc gcacagatgc gtaaggagaa aataccgcat caggaaattg
     1501 taaacgttaa tattttgtta aaattcgcgt taaatttttg ttaaatcagc tcatttttta
     1561 accaataggc cgaaatcggc aaaatccctt ataaatcaaa agaatagacc gagatagggt
     1621 tgagtgttgt tccagtttgg aacaagagtc cactattaaa gaacgtggac tccaacgtca
     1681 aagggcgaaa aaccgtctat cagggcgatg gcccactacg tgaaccatca ccctaatcaa
     1741 gttttttggg gtcgaggtgc cgtaaagcac taaatcggaa ccctaaaggg agcccccgat
     1801 ttagagcttg acggggaaag ccggcgaacg tggcgagaaa ggaagggaag aaagcgaaag
     1861 gagcgggcgc tagggcgctg gcaagtgtag cggtcacgct gcgcgtaacc accacacccg
     1921 ccgcgcttaa tgcgccgcta cagggcgcgt cgcgccattc gccattcagg ctgcgcaact
     1981 gttgggaagg gcgatcggtg cgggcctctt cgctattacg ccagctggcg aaagggggat
     2041 gtgctgcaag gcgattaagt tgggtaacgc cagggttttc ccagtcacga cgttgtaaaa
     2101 cgacggccag tgagcgcgcg taatacgact cactataggg cgaattgggt accgggcccc
     2161 ccctcgaggt cgacggtatc gataagcttg atatcgaatt cctgcagccc gggggatcca
     2221 ctagttctag agcggccgcc accgcggtgg agctccagct tttgttccct ttagtgaggg
     2281 ttaattgcgc gcttggcgta atcatggtca tagctgtttc ctgtgtgaaa ttgttatccg
     2341 ctcacaattc cacacaacat acgagccgga agcataaagt gtaaagcctg gggtgcctaa
     2401 tgagtgagct aactcacatt aattgcgttg cgctcactgc ccgctttcca gtcgggaaac
     2461 ctgtcgtgcc agctgcatta atgaatcggc caacgcgcgg ggagaggcgg tttgcgtatt
     2521 gggcgctctt ccgcttcctc gctcactgac tcgctgcgct cggtcgttcg gctgcggcga
     2581 gcggtatcag ctcactcaaa ggcggtaata cggttatcca cagaatcagg ggataacgca
     2641 ggaaagaaca tgtgagcaaa aggccagcaa aaggccagga accgtaaaaa ggccgcgttg
     2701 ctggcgtttt tccataggct ccgcccccct gacgagcatc acaaaaatcg acgctcaagt
     2761 cagaggtggc gaaacccgac aggactataa agataccagg cgtttccccc tggaagctcc
     2821 ctcgtgcgct ctcctgttcc gaccctgccg cttaccggat acctgtccgc ctttctccct
     2881 tcgggaagcg tggcgctttc tcatagctca cgctgtaggt atctcagttc ggtgtaggtc
     2941 gttcgctcca agctgggctg tgtgcacgaa ccccccgttc agcccgaccg ctgcgcctta
     3001 tccggtaact atcgtcttga gtccaacccg gtaagacacg acttatcgcc actggcagca
     3061 gccactggta acaggattag cagagcgagg tatgtaggcg gtgctacaga gttcttgaag
     3121 tggtggccta actacggcta cactagaagg acagtatttg gtatctgcgc tctgctgaag
     3181 ccagttacct tcggaaaaag agttggtagc tcttgatccg gcaaacaaac caccgctggt
     3241 agcggtggtt tttttgtttg caagcagcag attacgcgca gaaaaaaagg atctcaagaa
     3301 gatcctttga tcttttctac ggggtctgac gctcagtgga acgaaaactc acgttaaggg
     3361 attttggtca tgagattatc aaaaaggatc ttcacctaga tccttttaaa ttaaaaatga
     3421 agttttaaat caatctaaag tatatatgag taaacttggt ctgacagtta ccaatgctta
     3481 atcagtgagg cacctatctc agcgatctgt ctatttcgtt catccatagt tgcctgactc
     3541 cccgtcgtgt agataactac gatacgggag ggcttaccat ctggccccag tgctgcaatg
     3601 ataccgcgag acccacgctc accggctcca gatttatcag caataaacca gccagccgga
     3661 agggccgagc gcagaagtgg tcctgcaact ttatccgcct ccatccagtc tattaattgt
     3721 tgccgggaag ctagagtaag tagttcgcca gttaatagtt tgcgcaacgt tgttgccatt
     3781 gctacaggca tcgtggtgtc acgctcgtcg tttggtatgg cttcattcag ctccggttcc
     3841 caacgatcaa ggcgagttac atgatccccc atgttgtgca aaaaagcggt tagctccttc
     3901 ggtcctccga tcgttgtcag aagtaagttg gccgcagtgt tatcactcat ggttatggca
     3961 gcactgcata attctcttac tgtcatgcca tccgtaagat gcttttctgt gactggtgag
     4021 tactcaacca agtcattctg agaatagtgt atgcggcgac cgagttgctc ttgcccggcg
     4081 tcaatacggg ataataccgc gccacatagc agaactttaa aagtgctcat cattggaaaa
     4141 cgttcttcgg ggcgaaaact ctcaaggatc ttaccgctgt tgagatccag ttcgatgtaa
     4201 cccactcgtg cacccaactg atcttcagca tcttttactt tcaccagcgt ttctgggtga
     4261 gcaaaaacag gaaggcaaaa tgccgcaaaa aagggaataa gggcgacacg gaaatgttga
     4321 atactcatac tcttcctttt tcaatattat tgaagcattt atcagggtta ttgtctcatg
     4381 agcggataca tatttgaatg tatttagaaa aataaacaaa taggggttcc gcgcacattt
     4441 ccccgaaaag tgccacctga acgaagcatc tgtgcttcat tttgtagaac aaaaatgcaa
     4501 cgcgagagcg ctaatttttc aaacaaagaa tctgagctgc atttttacag aacagaaatg
     4561 caacgcgaaa gcgctatttt accaacgaag aatctgtgct tcatttttgt aaaacaaaaa
     4621 tgcaacgcga gagcgctaat ttttcaaaca aagaatctga gctgcatttt tacagaacag
     4681 aaatgcaacg cgagagcgct attttaccaa caaagaatct atacttcttt tttgttctac
     4741 aaaaatgcat cccgagagcg ctatttttct aacaaagcat cttagattac tttttttctc
     4801 ctttgtgcgc tctataatgc agtctcttga taactttttg cactgtaggt ccgttaaggt
     4861 tagaagaagg ctactttggt gtctattttc tcttccataa aaaaagcctg actccacttc
     4921 ccgcgtttac tgattactag cgaagctgcg ggtgcatttt ttcaagataa aggcatcccc
     4981 gattatattc tataccgatg tggattgcgc atactttgtg aacagaaagt gatagcgttg
     5041 atgattcttc attggtcaga aaattatgaa cggtttcttc tattttgtct ctatatacta
     5101 cgtataggaa atgtttacat tttcgtattg ttttcgattc actctatgaa tagttcttac
     5161 tacaattttt ttgtctaaag agtaatacta gagataaaca taaaaaatgt agaggtcgag
     5221 tttagatgca agttcaagga gcgaaaggtg gatgggtagg ttatataggg atatagcaca
     5281 gagatatata gcaaagagat acttttgagc aatgtttgtg gaagcggtat tcgcaatatt
     5341 ttagtagctc gttacagtcc ggtgcgtttt tggttttttg aaagtgcgtc ttcagagcgc
     5401 ttttggtttt caaaagcgct ctgaagttcc tatactttct agagaatagg aacttcggaa
     5461 taggaacttc aaagcgtttc cgaaaacgag cgcttccgaa aatgcaacgc gagctgcgca
     5521 catacagctc actgttcacg tcgcacctat atctgcgtgt tgcctgtata tatatataca
     5581 tgagaagaac ggcatagtgc gtgtttatgc ttaaatgcgt acttatatgc gtctatttat
     5641 gtaggatgaa aggtagtcta gtacctcctg tgatattatc ccattccatg cggggtatcg
     5701 tatgcttcct tcagcactac cctttagctg ttctatatgc tgccactcct caattggatt
     5761 agtctcatcc ttcaatgcta tcatttcctt tgatattgga tcatatctaa gaaaccatta
     5821 ttatcatgac attaacctat aaaaataggc gtatcacgag gccctttcgt c
//
