LOCUS       sequence-381545-        5814 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  High copy vector backbone with nourseothricin selectivity, contains 
            MCS.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5814)
  AUTHORS   Smith JR, Sislak CD, Fernandez Mendoza P, Carmichael L, Lewis AG, 
            Chen A, Jiang GZ, Gibney PA
  TITLE     Auxotrophy-Independent Plasmid Shuttle Vectors for Applications in 
            Diverse Yeasts
  JOURNAL   Appl Microbiol. 2024; 4(1):453-469. doi: 
            10.3390/applmicrobiol4010031
REFERENCE   2  (bases 1 to 5814)
  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: "Appl 
            Microbiol."; date: "2024"; volume: " 4(1):453-469. doi"; pages: " 
            10.3390/applmicrobiol401003"
COMMENT     Sequence Label: pRSII42N
FEATURES             Location/Qualifiers
     source          1..5814
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(105..124)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      918..1506
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     1407..1426
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     1660..1677
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    1794..1815
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(1830..1835,1836..1853,1854..1860)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    1868..1884
                     /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     1873..1895
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     1892..1908
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     1892..1908
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     CDS             1904..2284
                     /codon_start=1
                     /gene="lacZ fragment"
                     /product="LacZ-alpha fragment of beta-galactosidase"
                     /label=lacZ-alpha
                     /translation="MTMITPSAQLTLTKGNKSWSSTAVAAALELVDPPGCRNSISSLSI
                     PSTSRGGPVPNSPYSESYYARSLAVVLQRRDWENPGVTQLNRLAAHPPFASWRNSEEAR
                     TDRPSQQLRSLNGEWTRPVAAH"
     primer_bind     1927..1947
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        1929..1947
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     misc_feature    1960..2067
                     /label=MCS
                     /note="pBluescript multiple cloning site"
     primer_bind     1984..2000
                     /label=SK primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     1984..2000
                     /label=pBluescriptSK
                     /note="For pBluescript vector"
     primer_bind     complement(2034..2050)
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2035..2051)
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     primer_bind     complement(2075..2094)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(2076..2094)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(2104..2121)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(2104..2120)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2113..2135)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      2261..2689
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2348..2367)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2558..2579
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     terminator      complement(2804..3001)
                     /label=TEF terminator
                     /note="Ashbya gossypii TEF terminator"
     promoter        complement(3583..3926)
                     /label=TEF promoter
                     /note="Ashbya gossypii TEF promoter"
     primer_bind     complement(3985..4004)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     4104..4126
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(4164..4182)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     rep_origin      4223..5569
                     /label=2u ori
                     /note="yeast 2u plasmid origin of replication"
     promoter        5596..5700
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5701..5769,5770..5814,1..747)
                     /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"
ORIGIN
        1 ggtgcacgag tgggttacat cgaactggat ctcaacagcg gtaagatcct tgagagtttt
       61 cgccccgaag aacgttttcc aatgatgagc acttttaaag ttctgctatg tggcgcggta
      121 ttatcccgta ttgacgccgg gcaagagcaa ctcggtcgcc gcatacacta ttctcagaat
      181 gacttggttg agtactcacc agtcacagaa aagcatctta cggatggcat gacagtaaga
      241 gaattatgca gtgctgccat aaccatgagt gataacactg cggccaactt acttctgaca
      301 acgatcggag gaccgaagga gctaaccgct tttttgcaca acatggggga tcatgtaact
      361 cgccttgatc gttgggaacc ggagctgaat gaagccatac caaacgacga gcgtgacacc
      421 acgatgcctg tagcaatggc aacaacgttg cgcaaactat taactggcga actacttact
      481 ctagcttccc ggcaacaatt aatagactgg atggaggcgg ataaagttgc aggaccactt
      541 ctgcgctcgg cccttccggc tggctggttt attgctgata aatctggagc cggtgagcgt
      601 gggtctcgcg gtatcattgc agcactgggg ccagatggta agccctcccg tatcgtagtt
      661 atctacacga cggggagtca ggcaactatg gatgaacgaa atagacagat cgctgagata
      721 ggtgcctcac tgattaagca ttggtaactg tcagaccaag tttactcata tatactttag
      781 attgatttaa aacttcattt ttaatttaaa aggatctagg tgaagatcct ttttgataat
      841 ctcatgacca aaatccctta acgtgagttt tcgttccact gagcgtcaga ccccgtagaa
      901 aagatcaaag gatcttcttg agatcctttt tttctgcgcg taatctgctg cttgcaaaca
      961 aaaaaaccac cgctaccagc ggtggtttgt ttgccggatc aagagctacc aactcttttt
     1021 ccgaaggtaa ctggcttcag cagagcgcag ataccaaata ctgttcttct agtgtagccg
     1081 tagttaggcc accacttcaa gaactctgta gcaccgccta catacctcgc tctgctaatc
     1141 ctgttaccag tggccgctgc cagtggcgat aagtcgtatc ttaccgggtt ggactcaaga
     1201 cgatagttac cggataaggc gcagcggtcg ggctgaacgg ggggttcgtg cacacagccc
     1261 agcttggagc gaacgaccta caccgaactg agatacctac agcgtgagct atgagaaagc
     1321 gccacgcttc ccgaagggag aaaggcggac agatatccgg taagcggcag ggtcggaaca
     1381 ggagagcgca cgagggagct tccagtggga aacgcctggt atctttatag tcctgtcggg
     1441 tttcgccacc tctgacttga gcgtcgattt ttgtgatgct cgtcaggggg gcggagccta
     1501 tggaaaaacg ccagcaacgc ggccttttta cggttcctgg ccttttgctg gccttttgct
     1561 cacatgttct ttcctgcgtt atcccctgat tctgtggata accgtattac cgcctttgag
     1621 tgagctgata ccgctcgccg cagccgaacg accgagcgca gcgagtcagt gagcgaggaa
     1681 gcggaagagc gcccaatacg caaaccgcct ctccccgcgc gttggccgat tcattaatgc
     1741 agctggcacg acaggtttcc cgactggaaa gcgggcagtg agcgcaacgc aattaatgtg
     1801 agttagctca ctcattaggc accccaggct ttacacttta tgcttccggc tcgtatgttg
     1861 tgtggaattg tgagcggata acaatttcac acaggaaaca gctatgacca tgattacgcc
     1921 aagcgcgcaa ttaaccctca ctaaagggaa caaaagctgg agctccaccg cggtggcggc
     1981 cgctctagaa ctagtggatc ccccgggctg caggaattcg atatcaagct tatcgatacc
     2041 gtcgacctcg agggggggcc cggtacccaa ttcgccctat agtgagtcgt attacgcgcg
     2101 ctcactggcc gtcgttttac aacgtcgtga ctgggaaaac cctggcgtta cccaacttaa
     2161 tcgccttgca gcacatcccc ctttcgccag ctggcgtaat agcgaagagg cccgcaccga
     2221 tcgcccttcc caacagttgc gcagcctgaa tggcgaatgg acgcgccctg tagcggcgca
     2281 ttaagcgcgg cgggtgtggt ggttacgcgc agcgtgaccg ctacacttgc cagcgcccta
     2341 gcgcccgctc ctttcgcttt cttcccttcc tttctcgcca cgttcgccgg ctttccccgt
     2401 caagctctaa atcgggggct ccctttaggg ttccgattta gtgctttacg gcacctcgac
     2461 cccaaaaaac ttgattaggg tgatggttca cgtagtgggc catcgccctg atagacggtt
     2521 tttcgccctt tgacgttgga gtccacgttc tttaatagtg gactcttgtt ccaaactgga
     2581 acaacactca accctatctc ggtctattct tttgatttat aagggatttt gccgatttcg
     2641 gcctattggt taaaaaatga gctgatttaa caaaaattta acgcgaattt taacaaaata
     2701 ttaacgctta cagtttcctg atgcggtatt ttctccttac gcatctgtgc ggtatttcac
     2761 accgcataga cactggatgg cggcgttagt atcgaatcga cagcagtata gcgaccagca
     2821 ttcacatacg attgacgcat gatattactt tctgcgcact taacttcgca tctgggcaga
     2881 tgatgtcgag gcgaaaaaaa atataaatca cgctaacatt tgattaaaat agaacaacta
     2941 caatataaaa aaactataca aatgacaagt tcttgaaaac aagaatcttt ttattgtcag
     3001 tactgattat caagggcaag gcattgacat atataatgcc tgttctccat ctgaagcagt
     3061 tccgtcatac aaggctgtat caaggccaca taaagtaaaa cccattcttc tgtatgcgtg
     3121 gattgccggt gcatttacgt tggtgacttc caaccacaaa tgacctgcac ctctttcacg
     3181 tgcaaattct gttgcaagtc ccatcagagc cctacccaca ccgtgacccc tatgttcagg
     3241 agcaacttca atgtcttcga ctgtaggtct tctgttccac cccgaatatg agactactac
     3301 gaaaccagct aaatctccgt cgtcgccgta ggccacaaat gttctactat ccgggtctcc
     3361 gtcttcacca tcgtcacttt catcatcgct ttcatcatca ggaaacactt tggttagagg
     3421 tggatcgact ggaacttctc ttaatgtaaa gccgtctcct gtagctgtaa cacggaaaac
     3481 ggtatccgtc gtgaaagaac catcaagagc ttcaatagct tctgcatcac ctgggacaga
     3541 tgtacgatac ctgtaggcgg tgtcgtctaa tgtagtaccc atggttgttt atgttcggat
     3601 gtgatgtgag aactgtatcc tagcaagatt ttaaaaggaa gtatatgaaa gaagaacctc
     3661 agtggcaaat cctaaccttt tatatttctc tacaggggcg cggcgtgggg acaattcaac
     3721 gcgtctgtga ggggagcgtt tccctgctcg caggtctgca gcgaggagcc gtaatttttg
     3781 cttcgcgccg tgcggccatc aaaatgtatg gatgcaaacg attatacatg gggatgtatg
     3841 ggctaaatgt acgggcgaca gtcacatcat gcccctgagc tgcgcacgtc aagactgtca
     3901 aggagggtat tctgggcctc catgtcgctg gccgggtgac ccggcgggga cgaggcaagc
     3961 taaggtggta tggtgcactc tcagtacaat ctgctctgat gccgcatagt taagccagcc
     4021 ccgacacccg ccaacacccg ctgacgcgcc ctgacgggct tgtctgctcc cggcatccgc
     4081 ttacagacaa gctgtgaccg tctccgggag ctgcatgtgt cagaggtttt caccgtcatc
     4141 accgaaacgc gcgagacgaa agggcctcgt gatacgccta tttttatagg ttaatgtcat
     4201 gataataatg gtttcttagt atgatccaat atcaaaggaa atgatagcat tgaaggatga
     4261 gactaatcca attgaggagt ggcagcatat agaacagcta aagggtagtg ctgaaggaag
     4321 catacgatac cccgcatgga atgggataat atcacaggag gtactagact acctttcatc
     4381 ctacataaat agacgcatat aagtacgcat ttaagcataa acacgcacta tgccgttctt
     4441 ctcatgtata tatatataca ggcaacacgc agatataggt gcgacgtgaa cagtgagctg
     4501 tatgtgcgca gctcgcgttg cattttcgga agcgctcgtt ttcggaaacg ctttgaagtt
     4561 cctattccga agttcctatt ctctagctag aaagtatagg aacttcagag cgcttttgaa
     4621 aaccaaaagc gctctgaaga cgcactttca aaaaaccaaa aacgcaccgg actgtaacga
     4681 gctactaaaa tattgcgaat accgcttcca caaacattgc tcaaaagtat ctctttgcta
     4741 tatatctctg tgctatatcc ctatataacc tacccatcca cctttcgctc cttgaacttg
     4801 catctaaact cgacctctac attttttatg tttatctcta gtattactct ttagacaaaa
     4861 aaattgtagt aagaactatt catagagtga atcgaaaaca atacgaaaat gtaaacattt
     4921 cctatacgta gtatatagag acaaaataga agaaaccgtt cataattttc tgaccaatga
     4981 agaatcatca acgctatcac tttctgttca caaagtatgc gcaatccaca tcggtataga
     5041 atataatcgg ggatgccttt atcttgaaaa aatgcacccg cagcttcgct agtaatcagt
     5101 aaacgcggga agtggagtca ggcttttttt atggaagaga aaatagacac caaagtagcc
     5161 ttcttctaac cttaacggac ctacagtgca aaaagttatc aagagactgc attatagagc
     5221 gcacaaagga gaaaaaaagt aatctaagat gctttgttag aaaaatagcg ctctcgggat
     5281 gcatttttgt agaacaaaaa agaagtatag attctttgtt ggtaaaatag cgctctcgcg
     5341 ttgcatttct gttctgtaaa aatgcagctc agattctttg tttgaaaaat tagcgctctc
     5401 gcgttgcatt tttgttttac aaaaatgaag cacagattct tcgttggtaa aatagcgctt
     5461 tcgcgttgca tttctgttct gtaaaaatgc agctcagatt ctttgtttga aaaattagcg
     5521 ctctcgcgtt gcatttttgt tctacaaaat gaagcacaga tgcttcgttc aggtggcact
     5581 tttcggggaa atgtgcgcgg aacccctact tgtttatttt tctaaataca ttcaaatatg
     5641 tatccgctca tgagacaata accctgataa atgcttcaat aatattgaaa aaggaagagt
     5701 atgagtattc aacatttccg tgtcgccctt attccctttt ttgcggcatt ttgccttcct
     5761 gtttttgctc acccagaaac gctggtgaaa gtaaaagatg ctgaagatca gttg
//
