LOCUS       sequence-380545-        5215 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Low copy vector backbone with G418 selectivity, contains MCS.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5215)
  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 5215)
  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: pRSII41K
FEATURES             Location/Qualifiers
     source          1..5215
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(91..110)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      904..1492
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     1393..1412
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     1646..1663
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    1780..1801
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(1816..1821,1822..1839,1840..1846)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    1854..1870
                     /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     1859..1881
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     1878..1894
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     1878..1894
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     CDS             1890..2270
                     /codon_start=1
                     /gene="lacZ fragment"
                     /product="LacZ-alpha fragment of beta-galactosidase"
                     /label=lacZ-alpha
                     /translation="MTMITPSAQLTLTKGNKSWSSTAVAAALELVDPPGCRNSISSLSI
                     PSTSRGGPVPNSPYSESYYARSLAVVLQRRDWENPGVTQLNRLAAHPPFASWRNSEEAR
                     TDRPSQQLRSLNGEWTRPVAAH"
     primer_bind     1913..1933
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        1915..1933
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     misc_feature    1946..2053
                     /label=MCS
                     /note="pBluescript multiple cloning site"
     primer_bind     1970..1986
                     /label=SK primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     1970..1986
                     /label=pBluescriptSK
                     /note="For pBluescript vector"
     primer_bind     complement(2020..2036)
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2021..2037)
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     primer_bind     complement(2061..2080)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(2062..2080)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(2090..2107)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(2090..2106)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2099..2121)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      2247..2702
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2334..2353)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2544..2565
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     gene            complement(2790..4146)
                     /label=kanMX
                     /note="yeast selectable marker conferring kanamycin 
                     resistance (Wach et al., 1994)"
     terminator      complement(2790..2987)
                     /label=TEF terminator
                     /note="Ashbya gossypii TEF terminator"
     CDS             complement(2993..3802)
                     /codon_start=1
                     /gene="aph(3')-Ia"
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin"
                     /translation="MGKEKTHVSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYGKP
                     DAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGKTA
                     FQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDASD
                     FDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGIAD
                     RYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     primer_bind     3716..3735
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     promoter        complement(3803..4146)
                     /label=TEF promoter
                     /note="Ashbya gossypii TEF promoter"
     primer_bind     complement(4205..4224)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     4324..4346
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(4384..4402)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     misc_feature    4443..4946
                     /label=CEN/ARS
                     /note="S. cerevisiae CEN6 centromere fused to an 
                     autonomously replicating sequence"
     promoter        4983..5087
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5088..5156,5157..5215,1..733)
                     /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 ttacatcgaa ctggatctca acagcggtaa gatccttgag agttttcgcc ccgaagaacg
       61 ttttccaatg atgagcactt ttaaagttct gctatgtggc gcggtattat cccgtattga
      121 cgccgggcaa gagcaactcg gtcgccgcat acactattct cagaatgact tggttgagta
      181 ctcaccagtc acagaaaagc atcttacgga tggcatgaca gtaagagaat tatgcagtgc
      241 tgccataacc atgagtgata acactgcggc caacttactt ctgacaacga tcggaggacc
      301 gaaggagcta accgcttttt tgcacaacat gggggatcat gtaactcgcc ttgatcgttg
      361 ggaaccggag ctgaatgaag ccataccaaa cgacgagcgt gacaccacga tgcctgtagc
      421 aatggcaaca acgttgcgca aactattaac tggcgaacta cttactctag cttcccggca
      481 acaattaata gactggatgg aggcggataa agttgcagga ccacttctgc gctcggccct
      541 tccggctggc tggtttattg ctgataaatc tggagccggt gagcgtgggt ctcgcggtat
      601 cattgcagca ctggggccag atggtaagcc ctcccgtatc gtagttatct acacgacggg
      661 gagtcaggca actatggatg aacgaaatag acagatcgct gagataggtg cctcactgat
      721 taagcattgg taactgtcag accaagttta ctcatatata ctttagattg atttaaaact
      781 tcatttttaa tttaaaagga tctaggtgaa gatccttttt gataatctca tgaccaaaat
      841 cccttaacgt gagttttcgt tccactgagc gtcagacccc gtagaaaaga tcaaaggatc
      901 ttcttgagat cctttttttc tgcgcgtaat ctgctgcttg caaacaaaaa aaccaccgct
      961 accagcggtg gtttgtttgc cggatcaaga gctaccaact ctttttccga aggtaactgg
     1021 cttcagcaga gcgcagatac caaatactgt tcttctagtg tagccgtagt taggccacca
     1081 cttcaagaac tctgtagcac cgcctacata cctcgctctg ctaatcctgt taccagtggc
     1141 tgctgccagt ggcgataagt cgtgtcttac caggttggac tcaagacgat agttaccgga
     1201 taaggcgcag cggtcgggct gaacgggggg ttcgtgcaca cagcccagct tggagcgaac
     1261 gacctacacc gaactgagat acctacagcg tgagctatga gaaagcgcca cgcttcccga
     1321 agggagaaag gcggacaggt atccggtaag cggcagggtc ggaacaggag agcgcacgag
     1381 ggagcttcca gggggaaacg cctggtatct ttatagtcct gtcgggtttc gccacctctg
     1441 acttgagcgt cgatttttgt gatgctcgtc aggggggcgg agcctatgga aaaacgccag
     1501 caacgcggcc tttttacggt tcctggcctt ttgctggcct tttgctcaca tgttctttcc
     1561 tgcgttatcc cctgattctg tggataaccg tattaccgcc tttgagtgag ctgataccgc
     1621 tcgccgcagc cgaacgaccg agcgcagcga gtcagtgagc gaggaagcgg aagagcgccc
     1681 aatacgcaaa ccgcctctcc ccgcgcgttg gccgattcat taatgcagct ggcacgacag
     1741 gtttcccgac tggaaagcgg gcagtgagcg caacgcaatt aatgtgagtt agctcactca
     1801 ttaggcaccc caggctttac actttatgct tccggctcgt atgttgtgtg gaattgtgag
     1861 cggataacaa tttcacacag gaaacagcta tgaccatgat tacgccaagc gcgcaattaa
     1921 ccctcactaa agggaacaaa agctggagct ccaccgcggt ggcggccgct ctagaactag
     1981 tggatccccc gggctgcagg aattcgatat caagcttatc gataccgtcg acctcgaggg
     2041 ggggcccggt acccaattcg ccctatagtg agtcgtatta cgcgcgctca ctggccgtcg
     2101 ttttacaacg tcgtgactgg gaaaaccctg gcgttaccca acttaatcgc cttgcagcac
     2161 atcccccttt cgccagctgg cgtaatagcg aagaggcccg caccgatcgc ccttcccaac
     2221 agttgcgcag cctgaatggc gaatggacgc gccctgtagc ggcgcattaa gcgcggcggg
     2281 tgtggtggtt acgcgcagcg tgaccgctac acttgccagc gccctagcgc ccgctccttt
     2341 cgctttcttc ccttcctttc tcgccacgtt cgccggcttt ccccgtcaag ctctaaatcg
     2401 ggggctccct ttagggttcc gatttagtgc tttacggcac ctcgacccca aaaaacttga
     2461 ttagggtgat ggttcacgta gtgggccatc gccctgatag acggtttttc gccctttgac
     2521 gttggagtcc acgttcttta atagtggact cttgttccaa actggaacaa cactcaaccc
     2581 tatctcggtc tattcttttg atttataagg gattttgccg atttcggcct attggttaaa
     2641 aaatgagctg atttaacaaa aatttaacgc gaattttaac aaaatattaa cgcttacaat
     2701 ttcctgatgc ggtattttct ccttacgcat ctgtgcggta tttcacaccg catagacact
     2761 ggatggcggc gttagtatcg aatcgacagc agtatagcga ccagcattca catacgattg
     2821 acgcatgata ttactttctg cgcacttaac ttcgcatctg ggcagatgat gtcgaggcga
     2881 aaaaaaatat aaatcacgct aacatttgat taaaatagaa caactacaat ataaaaaaac
     2941 tatacaaatg acaagttctt gaaaacaaga atctttttat tgtcagtact gattagaaaa
     3001 actcatcgag catcaaatga aactgcaatt tattcatatc aggattatca ataccatatt
     3061 tttgaaaaag ccgtttctgt aatgaaggag aaaactcacc gaggcagttc cataggatgg
     3121 caagatcctg gtatcggtct gcgattccga ctcgtccaac atcaatacaa cctattaatt
     3181 tcccctcgtc aaaaataagg ttatcaagtg agaaatcacc atgagtgacg actgaatccg
     3241 gtgagaatgg caaaagctta tgcatttctt tccagacttg ttcaacaggc cagccattac
     3301 gctcgtcatc aaaatcactc gcatcaacca aaccgttatt cattcgtgat tgcgcctgag
     3361 cgagacgaaa tacgcgatcg ctgttaaaag gacaattaca aacaggaatc gaatgcaacc
     3421 ggcgcaggaa cactgccagc gcatcaacaa tattttcacc tgaatcagga tattcttcta
     3481 atacctggaa tgctgttttg ccggggatcg cagtggtgag taaccatgca tcatcaggag
     3541 tacggataaa atgcttgatg gtcggaagag gcataaattc cgtcagccag tttagtctga
     3601 ccatctcatc tgtaacatca ttggcaacgc tacctttgcc atgtttcaga aacaactctg
     3661 gcgcatcggg cttcccatac aatcgataga ttgtcgcacc tgattgcccg acattatcgc
     3721 gagcccattt atacccatat aaatcagcat ccatgttgga atttaatcgc ggcctcgaaa
     3781 cgtgagtctt ttccttaccc atggttgttt atgttcggat gtgatgtgag aactgtatcc
     3841 tagcaagatt ttaaaaggaa gtatatgaaa gaagaacctc agtggcaaat cctaaccttt
     3901 tatatttctc tacaggggcg cggcgtgggg acaattcaac gcgtctgtga ggggagcgtt
     3961 tccctgctcg caggtctgca gcgaggagcc gtaatttttg cttcgcgccg tgcggccatc
     4021 aaaatgtatg gatgcaaatg attatacatg gggatgtatg ggctaaatgt acgggcgaca
     4081 gtcacatcat gcccctgagc tgcgcacgtc aagactgtca aggagggtat tctgggcctc
     4141 catgtcgctg gccgggtgac ccggcgggga cgaggcaagc taaggtggta tggtgcactc
     4201 tcagtacaat ctgctctgat gccgcatagt taagccagcc ccgacacccg ccaacacccg
     4261 ctgacgcgcc ctgacgggct tgtctgctcc cggcatccgc ttacagacaa gctgtgaccg
     4321 tctccgggag ctgcatgtgt cagaggtttt caccgtcatc accgaaacgc gcgagacgaa
     4381 agggcctcgt gatacgccta tttttatagg ttaatgtcat gataataatg gtttcttaga
     4441 cggatcgctt gcctgtaact tacacgcgcc tcgtatcttt taatgatgga ataatttggg
     4501 aatttactct gtgtttattt atttttatgt tttgtatttg gattttagaa agtaaataaa
     4561 gaaggtagaa gagttacgga atgaagaaaa aaaaataaac aaaggtttaa aaaatttcaa
     4621 caaaaagcgt actttacata tatatttatt agacaagaaa agcagattaa atagatatac
     4681 attcgattaa cgataagtaa aatgtaaaat cacaggattt tcgtgtgtgg tcttctacac
     4741 agacaagatg aaacaattcg gcattaatac ctgagagcag gaagagcaag ataaaaggta
     4801 gtatttgttg gcgatccccc tagagtcttt tacatcttcg gaaaacaaaa actatttttt
     4861 ctttaatttc tttttttact ttctattttt aatttatata tttatattaa aaaatttaaa
     4921 ttataattat ttttatagca cgtgatgaaa aggacccagg tggcactttt cggggaaatg
     4981 tgcgcggaac ccctatttgt ttatttttct aaatacattc aaatatgtat ccgctcatga
     5041 gacaataacc ctgataaatg cttcaataat attgaaaaag gaagagtatg agtattcaac
     5101 atttccgtgt cgcccttatt cccttttttg cggcattttg ccttcctgtt tttgctcacc
     5161 cagaaacgct ggtgaaagta aaagatgctg aagatcagtt gggtgcacga gtggg
//
