LOCUS       sequence-263742-        7186 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Integration into fission yeast genome at ura4 locus with natMX 
            selection cassette.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7186)
  AUTHORS   Vjestica A, Marek M, Nkosi PJ, Merlini L, Liu G, Berard M, 
            Billault-Chaumartin I, Martin SG
  TITLE     A toolbox of stable integration vectors in the fission yeast 
            Schizosaccharomyces pombe.
  JOURNAL   J Cell Sci. 2020 Jan 8;133(1). pii: jcs.240754. doi: 
            10.1242/jcs.240754.
  PUBMED    31801797
REFERENCE   2  (bases 1 to 7186)
  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: "J Cell 
            Sci."; date: "2020-01-8"; volume: "133(1). pii: jcs.240754. doi"; 
            pages: " 10.1242/jcs.240754"
COMMENT     Sequence Label: pAV0584
FEATURES             Location/Qualifiers
     source          1..7186
                     /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"
     CDS             1640..2434
                     /codon_start=1
                     /gene="Schizosaccharomyces pombe ura4"
                     /product="orotidine 5'-phosphate decarboxylase, required 
                     for uracil biosynthesis"
                     /label=S. pombe ura4
                     /translation="MDARVFQSYSARAEGMKNPIAKELLALMEEKQSNLSVAVDLTKKS
                     EILELVDKIGPYVCVIKTHIDVVEDFDQDMVEKLVALGKKHRFLIFEDRKFADIGNTVK
                     LQYASGVYKIASWAHITNCHTVPGEGIIQGLKEVGLPLGRGLLLLAEMSSKGSLATGSY
                     TEKTLEWFEKHTDFCFGFIAGRRFPNLQSDYITMSPGIGLDVKGDGLGQQYRTPEEVIV
                     NCGSDIIIVGRGVYGAGRNPVVEAKRYREAGWKAYQQRLSQH"
     rep_origin      complement(2926..3381)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3063..3084)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     3275..3294
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3507..3529
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     3521..3538
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     3522..3538
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     3548..3567
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        3548..3566
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     3591..3607
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     primer_bind     3592..3608
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3642..3658)
                     /label=SK primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3642..3658)
                     /label=pBluescriptSK
                     /note="For pBluescript vector"
     gene            3745..4864
                     /label=natMX6
                     /note="yeast selectable marker conferring nourseothricin 
                     resistance"
     promoter        3745..4088
                     /label=TEF promoter
                     /note="Ashbya gossypii TEF promoter"
     CDS             4089..4661
                     /codon_start=1
                     /gene="Streptomyces noursei nat1"
                     /product="nourseothricin acetyltransferase"
                     /label=NrsR
                     /note="confers resistance to nourseothricin"
                     /translation="MGTTLDDTAYRYRTSVPGDAEAIEALDGSFTTDTVFRVTATGDGF
                     TLREVPVDPPLTKVFPDDESDDESDDGEDGDPDSRTFVAYGDDGDLAGFVVVSYSGWNR
                     RLTVEDIEVAPEHRGHGVGRALMGLATEFARERGAGHLWLEVTNVNAPAIHAYRRMGFT
                     LCGLDTALYDGTASDGEQALYMSMPCP"
     terminator      4667..4864
                     /label=TEF terminator
                     /note="Ashbya gossypii TEF terminator"
     primer_bind     complement(4926..4946)
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        complement(4926..4944)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(4965..4981)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4965..4981)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4978..5000)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4989..5005
                     /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(5013..5019,5020..5037,5038..5043))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5058..5079
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5196..5213)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5367..5955)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5447..5466)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6126..6917,6918..6986))
                     /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     6749..6768
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6987..7091)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     7159..7177
                     /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 accataaccg ggtattagtc aaaactagta tactttttct cggagtataa tacacaatat
      241 cggtgcaaat aggttttaaa attgctccaa tcacatgttc ttaagaaaaa aacgtcaaaa
      301 gaaatctaag tattcccaaa ataattcctc ctgacatgag aagaataaaa aacactaaga
      361 tgcattgtca gtatgaaaca caagccaaag ttgctaacaa taggatcaat tcttacttac
      421 tggcaatgtt acgaggaatc ttggtaggag gatgtttggc aacataacgt tcttttgaat
      481 ttgtttttac acgacgagta atgttcttct gaaactgtgt gtttgcacga cgctgagagg
      541 gagtttgaac cgtctaaaag acatagttag gatccatgat gcacagattt ctaaagataa
      601 aactacttac aggcatgttg gtttattaaa aaagaaaaac aggaatgata agagggaaag
      661 aatacgtgta cagagcgctt acataatgag tgagtgtcat tattgaggaa tgccatgtca
      721 gatttgacac aactcagaac ttattaactt tttatgattg gaacttttgt tttgcccaag
      781 gaatttttaa tggaagaagt tgttttataa atgatattga atttatatcc actcttttta
      841 gcgcatttgc atctctactt taggttcttc tttgactaga ttctaggatg cgtattggat
      901 atactggtct ttgaatcatt gtcgaaaaaa aaattcatgg ttgtacccat gagcaaactg
      961 cttttgtggt tatccatgtg cttcttttgt tcagcatcta tcttcttaat cgcatggaag
     1021 atttgattta aaccaaaaga aatattgtta aactttgaca gaagctttaa attttgattc
     1081 catacattaa atggataaaa ctcctgaagc ttagctacaa atcccactgg ctatatgtat
     1141 gcatttgtgt taaaaaagtt tgtatagatt atttaatcta ctcagcattc tttctctaaa
     1201 taggaatttg ttacttaatg gagaaaaaaa tgtttcgatt tacctagtgt atttgtttgt
     1261 atactcacgt ttaatttcaa acatccattc tatcttgtgt aatttttggc atggtgaaaa
     1321 agataatcag ccttataatc tttacaaaag taagaaattc tgtaaataag ccttaatgcc
     1381 cttgctttaa attaaaatgg ttctttttca tgataatgtt tgcactttgt gaatatattt
     1441 tagatagttc tgtgaggtat aattaagatg ttttagagac ttatacaatt ttgtctttat
     1501 aaattcttaa ttgattttac catcccagtt taactatgct tcgtcggcat ctctgcacat
     1561 gtcgtgtttt cttaccgtat tgtcctacca agaacctctt ttttgcttgg atcgaaatta
     1621 aaggtttaaa agcaaagtta tggatgctag agtatttcaa agctattcag ctagagctga
     1681 ggggatgaaa aatcccattg ccaaggaatt gttggctttg atggaagaaa agcaaagcaa
     1741 cttgtcagtc gcggtcgatt tgacgaagaa atccgaaatc ttagaattgg tagataaaat
     1801 tggaccctat gtctgtgtta tcaagacaca tattgacgtt gtcgaggatt tcgaccagga
     1861 tatggtagaa aaactggtgg ccttaggtaa aaagcatcgt tttcttatct ttgaggatcg
     1921 caaattcgca gacattggaa ataccgtcaa gctacaatat gcatctggtg tgtacaaaat
     1981 tgcttcttgg gctcatatca caaattgcca tacagtgcca ggcgagggta ttatacaagg
     2041 cctcaaagaa gttggtttac ctttgggacg tggtctcttg cttttggctg aaatgtcttc
     2101 caaaggctct ttggctactg gttcctacac agagaaaacc ttagaatggt ttgagaagca
     2161 taccgatttt tgctttggct ttatagctgg tcgtcgattt cctaaccttc aaagcgacta
     2221 cataactatg tcccctggta tcggcttgga tgttaaagga gacgggctgg gacagcaata
     2281 tcgtactcct gaagaagtga ttgtaaactg cggtagcgat atcatcattg ttggtcgtgg
     2341 agtttatgga gctggtcgta atcctgttgt cgaagccaag agatatagag aagctggttg
     2401 gaaggcatat cagcaaagac tttctcagca ttaaaaaaag actaatgtaa aatttttttg
     2461 gttggttatt gaaaaagtcg atgccttgtt tgcgtttgtt ttcctaggcg ttttatgtca
     2521 gaaggcattt agaattagta tacaagtact ctttggtaaa attttatgta gcgactaaaa
     2581 tattaactat tatagataaa caccttggga ataaaaagta atttgctata gtaatttatt
     2641 aaacatgctc ctacaacatt accacaatct tttctcttgg attgacattg aataagaaaa
     2701 gagtgaattt ttttagactt gtaatgataa ctatgtacaa agccaatgaa agatgtatgt
     2761 agatgaatgt aaaataccat gtagacaaac aagataaaac ttggttataa acattggtgt
     2821 tggaacagaa taaattagat gtcaaaaagt ttcgtcaata tcacaagctt gctagggcgg
     2881 tgtgaaatac cgcacagatg cgtaaggaga aaataccgca tcaggaaatt gtaagcgtta
     2941 atattttgtt aaaattcgcg ttaaattttt gttaaatcag ctcatttttt aaccaatagg
     3001 ccgaaatcgg caaaatccct tataaatcaa aagaatagac cgagataggg ttgagtgttg
     3061 ttccagtttg gaacaagagt ccactattaa agaacgtgga ctccaacgtc aaagggcgaa
     3121 aaaccgtcta tcagggcgat ggcccactac gtgaaccatc accctaatca agttttttgg
     3181 ggtcgaggtg ccgtaaagca ctaaatcgga accctaaagg gagcccccga tttagagctt
     3241 gacggggaaa gccggcgaac gtggcgagaa aggaagggaa gaaagcgaaa ggagcgggcg
     3301 ctagggcgct ggcaagtgta gcggtcacgc tgcgcgtaac caccacaccc gccgcgctta
     3361 atgcgccgct acagggcgcg tccattcgcc attcaggctg cgcaactgtt gggaagggcg
     3421 atcggtgcgg gcctcttcgc tattacgcca gctggcgaaa gggggatgtg ctgcaaggcg
     3481 attaagttgg gtaacgccag ggttttccca gtcacgacgt tgtaaaacga cggccagtga
     3541 gcgcgcgtaa tacgactcac tatagggcga attgggtacc gggccccccc tcgaggtcga
     3601 cggtatcgat aagcttgata tcgaattcct gcagcccggg ggatccacta gttctagagc
     3661 ggccgccacc gcggtggtta attaaggcgc gccagatctg agctcgttta gcttgcctcg
     3721 tccccgccgg gtcacccggc cagcgacatg gaggcccaga ataccctcct tgacagtctt
     3781 gacgtgcgca gctcaggggc atgatgtgac tgtcgcccgt acatttagcc catacatccc
     3841 catgtataat catttgcatc catacatttt gatggccgca cggcgcgaag caaaaattac
     3901 ggctcctcgc tgcagacctg cgagcaggga aacgctcccc tcacagacgc gttgaattgt
     3961 ccccacgccg cgcccctgta gagaaatata aaaggttagg atttgccact gaggttcttc
     4021 tttcatatac ttccttttta aatcttgcta ggatacagtt ctcacatcac atccgaacat
     4081 aaacaaccat gggtaccact cttgacgaca cggcttaccg gtaccgcacc agtgtcccgg
     4141 gggacgccga ggccatcgag gcactggatg ggtccttcac caccgacacc gtcttccgcg
     4201 tcaccgccac cggggacggc ttcaccctgc gggaggtgcc ggtggacccg cccctgacca
     4261 aggtgttccc cgacgacgaa tcggacgacg aatcggacga cggggaggac ggcgacccgg
     4321 actctcggac gttcgtcgcg tacggggacg acggcgacct ggcgggcttc gtggtcgtct
     4381 cgtactccgg ctggaaccgc cggctgaccg tcgaggacat cgaggtcgcc ccggagcacc
     4441 gggggcacgg ggtcgggcgc gcgttgatgg ggctcgcgac ggagttcgcc cgcgagcggg
     4501 gtgccgggca cctctggctg gaggtcacca acgtcaacgc accggcgatc cacgcgtacc
     4561 ggcggatggg gttcaccctc tgcggcctgg acaccgccct gtacgacggc accgcctcgg
     4621 acggcgagca ggcgctctac atgagcatgc cctgccccta atcagtactg acaataaaaa
     4681 gattcttgtt ttcaagaact tgtcatttgt atagtttttt tatattgtag ttgttctatt
     4741 ttaatcaaat gttagcgtga tttatatttt ttttcgcctc gacatcatct gcccagatgc
     4801 gaagttaagt gcgcagaaag taatatcatg cgtcaatcgt atgtgaatgc tggtcgctat
     4861 actgctgtcg attcgatact aacgccgcca tccacctgca ggtaatcacg tgccagcttt
     4921 tgttcccttt agtgagggtt aattgcgcgc ttggcgtaat catggtcata gctgtttcct
     4981 gtgtgaaatt gttatccgct cacaattcca cacaacatac gagccggaag cataaagtgt
     5041 aaagcctggg gtgcctaatg agtgagctaa ctcacattaa ttgcgttgcg ctcactgccc
     5101 gctttccagt cgggaaacct gtcgtgccag ctgcattaat gaatcggcca acgcgcgggg
     5161 agaggcggtt tgcgtattgg gcgctcttcc gcttcctcgc tcactgactc gctgcgctcg
     5221 gtcgttcggc tgcggcgagc ggtatcagct cactcaaagg cggtaatacg gttatccaca
     5281 gaatcagggg ataacgcagg aaagaacatg tgagcaaaag gccagcaaaa ggccaggaac
     5341 cgtaaaaagg ccgcgttgct ggcgtttttc cataggctcc gcccccctga cgagcatcac
     5401 aaaaatcgac gctcaagtca gaggtggcga aacccgacag gactataaag ataccaggcg
     5461 tttccccctg gaagctccct cgtgcgctct cctgttccga ccctgccgct taccggatac
     5521 ctgtccgcct ttctcccttc gggaagcgtg gcgctttctc atagctcacg ctgtaggtat
     5581 ctcagttcgg tgtaggtcgt tcgctccaag ctgggctgtg tgcacgaacc ccccgttcag
     5641 cccgaccgct gcgccttatc cggtaactat cgtcttgagt ccaacccggt aagacacgac
     5701 ttatcgccac tggcagcagc cactggtaac aggattagca gagcgaggta tgtaggcggt
     5761 gctacagagt tcttgaagtg gtggcctaac tacggctaca ctagaagaac agtatttggt
     5821 atctgcgctc tgctgaagcc agttaccttc ggaaaaagag ttggtagctc ttgatccggc
     5881 aaacaaacca ccgctggtag cggtggtttt tttgtttgca agcagcagat tacgcgcaga
     5941 aaaaaaggat ctcaagaaga tcctttgatc ttttctacgg ggtctgacgc tcagtggaac
     6001 gaaaactcac gttaagggat tttggtcatg agattatcaa aaaggatctt cacctagatc
     6061 cttttaaatt aaaaatgaag ttttaaatca atctaaagta tatatgagta aacttggtct
     6121 gacagttacc aatgcttaat cagtgaggca cctatctcag cgatctgtct atttcgttca
     6181 tccatagttg cctgactccc cgtcgtgtag ataactacga tacgggaggg cttaccatct
     6241 ggccccagtg ctgcaatgat accgcgagac ccacgctcac cggctccaga tttatcagca
     6301 ataaaccagc cagccggaag ggccgagcgc agaagtggtc ctgcaacttt atccgcctcc
     6361 atccagtcta ttaattgttg ccgggaagct agagtaagta gttcgccagt taatagtttg
     6421 cgcaacgttg ttgccattgc tacaggcatc gtggtgtcac gctcgtcgtt tggtatggct
     6481 tcattcagct ccggttccca acgatcaagg cgagttacat gatcccccat gttgtgcaaa
     6541 aaagcggtta gctccttcgg tcctccgatc gttgtcagaa gtaagttggc cgcagtgtta
     6601 tcactcatgg ttatggcagc actgcataat tctcttactg tcatgccatc cgtaagatgc
     6661 ttttctgtga ctggtgagta ctcaaccaag tcattctgag aatagtgtat gcggcgaccg
     6721 agttgctctt gcccggcgtc aatacgggat aataccgcgc cacatagcag aactttaaaa
     6781 gtgctcatca ttggaaaacg ttcttcgggg cgaaaactct caaggatctt accgctgttg
     6841 agatccagtt cgatgtaacc cactcgtgca cccaactgat cttcagcatc ttttactttc
     6901 accagcgttt ctgggtgagc aaaaacagga aggcaaaatg ccgcaaaaaa gggaataagg
     6961 gcgacacgga aatgttgaat actcatactc ttcctttttc aatattattg aagcatttat
     7021 cagggttatt gtctcatgag cggatacata tttgaatgta tttagaaaaa taaacaaata
     7081 ggggttccgc gcacatttcc ccgaaaagtg ccacctgacg tctaagaaac cattattatc
     7141 atgacattaa cctataaaaa taggcgtatc acgaggccct ttcgtc
//
