ID PRS316 preliminary; circular DNA; SYN; 4887 BP. XX AC U03442; ATCC77145; XX DT 01-JUL-1993 (Rel. 7, Created) DT 01-JUL-1995 (Rel. 12, Last updated, Version 1) XX DE Saccharomyces/E.coli phagemid vector pRS316 - complete. XX KW cloning vector. XX OS Cloning vector OC Artificial sequences; Cloning vehicles. XX RN [1] RP 1-4887 RC pRSS56 from pBluescript KS+ & BlueScribe RC pRS303 from pRSS56 & HIS3 gene RC pRS304 from pRSS56 & TRP1 gene RC pRS305 from pRSS56 & LEU2 gene RC pRS306 from pRSS56 & URA3 gene RC pUC19-CEN6:32 from pUC19 & CEN6 RC pRSS83 from pUC19-CEN6:32 & pAB9, ARS RC pRSS84 from pRSS83 RC pRS313 from pRS303 & pRSS84 RC pRS314 from pRS304 & pRSS84 RC pRS315 from pRS305 & pRSS84 RC pRS316 from pRS306 & pRSS84 RC pRSS93 from pRSS84 & pYCF5 RA Sikorski R.S., Hieter P.; RT "A system of shuttle vectors and yeast host strains designed for RT efficient manipulation of DNA in Saccharomyces cerevisiae"; RL Genetics 122:19-27(1989). XX RN [2] RC from YEp24 RC from YCp50 RC from YIp5 RA Strathern J.N., Higgins D.R.; RT "Recovery of plasmids from yeast into Escherichia coli: shuttle RT vectors"; RL Meth. Enzymol. 194:319-329(1991). XX RN [3] RC pRS316 RA Stillman D.J.; RT ; RL Submitted (10-NOV-1993) by: RL Stillman D.J., Dept. of Cellular, Viral and Molecular Biology, RL University of Utah Medical Center, Salt Lake City, UT 84132 USA. XX CC One of a series of pBluescript-based centromere vectors (ATCC 77142 CC ATCC 77145, ATCC 77157 to ATCC 77158) differing in the yeast CC selectable marker gene. [1] CC The order of the major features in this plasmid is: URA3 - f1 ori CC (NaeI) - T7 promoter - lacZ'/MCS - T3 promoter - pMB1 ori - bla - CC CEN6 - ARSH4. CC A fragment (1.112 kb) containing the URA3 gene was inserted into the CC NdeI site and a cassette containing CEN6 and the ARS associated with CC histone 4 (ARSH4) was inserted into the AatII site of pRSS56. All ends CC were blunted. CC pRSS56, constructed by ligating a PvuI fragment (bp 498-2412) of CC pBluescript KS+ to a PvuI fragment (bp 2850-730) of pBS(+), contains CC the KS MCS from pBluescript KS+ and the unique NdeI and AatII sites CC between bla and f1 origin of pBS(+). [1] CC YC-type centromere vector permitting visual detection of recombinants CC and production of ssDNA in E.coli. Contains promoters for in vitro CC RNA synthesis, priming sites useful for sequencing, and encodes the CC lacZ alpha (lacZ') peptide. [1] CC Useful in plasmid shuffle experiments. CC Restriction digests of the clone give the following sizes (kb): CC EcoRI--5.0; BamHI--5.0, PvuII--4.4, 0.5. (ATCC staff) CC Medium is 1227 LB plus ampicillin. CC NCBI gi: 417973 CC NM (pRS316) CC CM (yes) CC NA (ds-DNA) CC TP (circular) CC ST () CC TY (phagemid) CC SP (ATCC) CC HO (E.coli HB101)(Saccharomyces cerevisiae)(E.coli) CC HO (Saccharomyces cerevisiae YPH499)(Saccharomyces cerevisiae YPH500) CC HO (Saccharomyces cerevisiae YPH501) CC CP () CC FN (cloning) CC SE () CC PA () CC BR (pRS314)(pRS315)(pRS313) CC OF () CC OR () XX FH Key Location/Qualifiers FH FT misc_feature 0..0 FT /note="1. pUC19 HincII 2686bp 432..432 FT 2. yeast MboII-MboII 125bp, CEN6 FT -> pUC19-CEN6:32 2811bp FT 1. pUC8 BamHI 2665bp 241..241 FT 2. yeast Sau3AI-Sau3AI 374bp, histone H4 ARS/ARSH4 FT -> pAB9 3039bp FT 1. pUC19-CEN6:32 EcoRI-ScaI 1908bp 397..-..2180, CEN6 FT 2. pAB9 ScaI-EcoRI 1314bp 231..-..1171, FT \ histone H4 ARS/ARSH4 FT -> pRSS83 3222bp [in MCS] FT 1. pRSS83 remove SmaI-XbaI 9bp, MCS/pUC19 415..424 FT \ 3213bp FT Klenow:Klenow FT -> pRSS84 3213bp FT 1. pRSS84 HincII-PstI, CEN6/ARSH4 FT \ pUC19 440..2686..432 FT T4 DNA polymerase:T4 DNA polymerase FT 2. pRS306 AatII 4373bp 848..848 FT T4 DNA polymerase:T4 DNA polymerase FT -> pRS316 4887bp" FT misc_binding 0..0 FT /note="MCS KpnI-ApaI-XhoI-SalI-ClaI-HindIII-EcoRV- FT EcoRI-PstI-SmaI- FT BamHI-SpeI-XbaI-EagI-NotI-BstXI-SacII-SacI" FT misc_binding 0..0 FT /note="SIT unique KpnI-ApaI-XhoI-SalI-ClaI-EcoRI- FT PstI-SmaI-BamHI-SpeI-EagI-NotI-SacII-SacI-NaeI" FT rep_origin 0..0 FT /note="ORI E. coli pMB1 (ColE1 and pBR322)" FT rep_origin 0..0 FT /note="ORI bacteriophage f1" FT promoter 0..0 FT /note="PRO E. coli lac gene" FT promoter 0..0 FT /note="PRO bacteriophage T3" FT promoter 0..0 FT /note="PRO bacteriophage T7" FT CDS 0..0 FT /note="GEN E. coli beta-galactosidase gene (lacZ'); FT reporter gene" FT CDS 0..0 FT /note="ANT E. coli beta-lactamase gene (bla) FT ampicillin resistance gene (apr/amp)" FT CDS 0..0 FT /note="ANT yeast URA3 gene" FT rep_origin 0..0 FT /note="ORI yeast ARSH4" FT misc_feature 0..0 FT /note="yeast centromere CEN6" XX SQ Sequence 4887 BP; 1361 A; 1069 C; 1167 G; 1290 T; 0 other; tcgcgcgttt cggtgatgac ggtgaaaacc tctgacacat gcagctcccg gagacggtca cagcttgtct gtaagcggat gccgggagca gacaagcccg tcagggcgcg tcagcgggtg ttggcgggtg tcggggctgg cttaactatg cggcatcaga gcagattgta ctgagagtgc accacgcttt tcaattcaat tcatcatttt ttttttattc ttttttttga tttcggtttc tttgaaattt ttttgattcg gtaatctccg aacagaagga agaacgaagg aaggagcaca gacttagatt ggtatatata cgcatatgta gtgttgaaga aacatgaaat tgcccagtat tcttaaccca actgcacaga acaaaaacct gcaggaaacg aagataaatc atgtcgaaag ctacatataa ggaacgtgct gctactcatc ctagtcctgt tgctgccaag ctatttaata tcatgcacga aaagcaaaca aacttgtgtg cttcattgga tgttcgtacc accaaggaat tactggagtt agttgaagca ttaggtccca aaatttgttt actaaaaaca catgtggata tcttgactga tttttccatg gagggcacag ttaagccgct aaaggcatta tccgccaagt acaatttttt actcttcgaa gacagaaaat ttgctgacat tggtaataca gtcaaattgc agtactctgc gggtgtatac agaatagcag aatgggcaga cattacgaat gcacacggtg tggtgggccc aggtattgtt agcggtttga agcaggcggc agaagaagta acaaaggaac ctagaggcct tttgatgtta gcagaattgt catgcaaggg ctccctatct actggagaat atactaaggg tactgttgac attgcgaaga gcgacaaaga ttttgttatc ggctttattg ctcaaagaga catgggtgga agagatgaag gttacgattg gttgattatg acacccggtg tgggtttaga tgacaaggga gacgcattgg gtcaacagta tagaaccgtg gatgatgtgg tctctacagg atctgacatt attattgttg gaagaggact atttgcaaag ggaagggatg ctaaggtaga gggtgaacgt tacagaaaag caggctggga agcatatttg agaagatgcg gccagcaaaa ctaaaaaact gtattataag taaatgcatg tatactaaac tcacaaatta gagcttcaat ttaattatat cagttattac cctgcggtgt gaaataccgc acagatgcgt aaggagaaaa taccgcatca ggaaattgta aacgttaata ttttgttaaa attcgcgtta aatttttgtt aaatcagctc attttttaac caataggccg aaatcggcaa aatcccttat aaatcaaaag aatagaccga gatagggttg agtgttgttc cagtttggaa caagagtcca ctattaaaga acgtggactc caacgtcaaa gggcgaaaaa ccgtctatca gggcgatggc ccactacgtg aaccatcacc ctaatcaagt tttttggggt cgaggtgccg taaagcacta aatcggaacc ctaaagggag cccccgattt agagcttgac ggggaaagcc ggcgaacgtg gcgagaaagg aagggaagaa agcgaaagga gcgggcgcta gggcgctggc aagtgtagcg gtcacgctgc gcgtaaccac cacacccgcc gcgcttaatg cgccgctaca gggcgcgtcg cgccattcgc cattcaggct gcgcaactgt tgggaagggc gatcggtgcg ggcctcttcg ctattacgcc agctggcgaa ggggggatgt gctgcaaggc gattaagttg ggtaacgcca gggttttccc agtcacgacg ttgtaaaacg acggccagtg aattgtaata cgactcacta tagggcgaat tggagctcca ccgcggtggc ggccgctcta gaactagtgg atcccccggg ctgcaggaat tcgatatcaa gcttatcgat accgtcgacc tcgagggggg gcccggtacc cagcttttgt tccctttagt gagggttaat tccgagcttg gcgtaatcat ggtcatagct gtttcctgtg tgaaattgtt atccgctcac aattccacac aacataggag ccggaagcat aaagtgtaaa gcctggggtg cctaatgagt gaggtaactc acattaattg cgttgcgctc actgcccgct ttccagtcgg gaaacctgtc gtgccagctg cattaatgaa tcggccaacg cgcggggaga ggcggtttgc gtattgggcg ctcttccgct tcctcgctca ctgactcgct gcgctcggtc gttcggctgc ggcgagcggt atcagctcac tcaaaggcgg taatacggtt atccacagaa tcaggggata acgcaggaaa gaacatgtga gcaaaaggcc agcaaaaggc caggaaccgt aaaaaggccg cgttgctggc gtttttccat aggctcggcc cccctgacga gcatcacaaa aatcgacgct caagtcagag gtggcgaaac ccgacaggac tataaagata ccaggcgttc ccccctggaa gctccctcgt gcgctctcct gttccgaccc tgccgcttac cggatacctg tccgcctttc tcccttcggg aagcgtggcg ctttctcaat gctcacgctg taggtatctc agttcggtgt aggtcgttcg ctccaagctg ggctgtgtgc acgaaccccc cgttcagccc gaccgctgcg ccttatccgg taactatcgt cttgagtcca acccggtaag acacgactta tcgccactgg cagcagccac tggtaacagg attagcagag cgaggtatgt aggcggtgct acagagttct tgaagtggtg gcctaactac ggctacacta gaaggacagt atttggtatc tgcgctctgc tgaagccagt taccttcgga aaaagagttg gtagctcttg atccggcaaa caaaccaccg ctggtagcgg tggttttttt gtttgcaagc agcagattac gcgcagaaaa aaaggatctc aagaagatcc tttgatcttt tctacggggt ctgacgctca gtggaacgaa aactcacgtt aagggatttt ggtcatgaga ttatcaaaaa ggatcttcac ctagatcctt ttaaattaaa aatgaagttt taaatcaatc taaagtatat atgagtaaac ttggtctgac agttaccaat gcttaatcag tgaggcacct atctcagcga tctgtctatt tcgttcatcc atagttgcct gactgcccgt cgtgtagata actacgatac gggagggctt accatctggc cccagtgctg caatgatacc gcgagaccca cgctcaccgg ctccagattt atcagcaata aaccagccag ccggaagggc cgagcgcaga agtggtcctg caactttatc cgcctccatc cagtctatta attgttgccg ggaagctaga gtaagtagtt cgccagttaa tagtttgcgc aacgttgttg ccattgctac aggcatcgtg gtgtcacgct cgtcgtttgg tatggcttca ttcagctccg gttcccaacg atcaaggcga gttacatgat cccccatgtt gtgaaaaaaa gcggttagct ccttcggtcc tccgatcgtt gtcagaagta agttggccgc agtgttatca ctcatggtta tggcagcact gcataattct cttactgtca tgccatccgt aagatgcttt tctgtgactg gtgagtactc aaccaagtca ttctgagaat agtgtatgcg gcgaccgagt tgctcttgcc cggcgtcaat acgggataat accgcgccac atagcagaac tttaaaagtg ctcatcattg gaaaacgttc ttcggggcga aaactctcaa ggatcttacc gctgttgaga tccagttcga tgtaacccac tcgtgcaccc aactgatctt cagcatcttt tactttcacc agcgtttctg ggtgagcaaa aacaggaagg caaaatgccg caaaaaaggg aataagggcg acacggaaat gttgaatact catactcttc ctttttcaat attattgaag catttatcag ggttattgtc tcatgagcgg atacatattt gaatgtattt agaaaaataa acaaataggg gttccgcgca catttccccg aaaagtgcca cctgggtcct tttcatcacg tgctataaaa ataattataa tttaaatttt ttaatataaa tatataaatt aaaaatagaa agtaaaaaaa gaaattaaag aaaaaatagt ttttgttttc cgaagatgta aaagactcta gggggatcgc caacaaatac taccttttat cttgctcttc ctgctctcag gtattaatgc cgaattgttt catcttgtct gtgtagaaga ccacacacga aaatcctgtg attttacatt ttacttatcg ttaatcgaat gtatatctat ttaatctgct tttcttgtct aataaatata tatgtaaagt acgctttttg ttgaaatttt ttaaaccttt gtttattttt ttttcttcat tccgtaactc ttctaccttc tttatttact ttctaaaatc caaatacaaa acataaaaat aaataaacac agagtaaatt cccaaattat tccatcatta aaagatacga ggcgcgtgta agttacaggc aagcgatccg tcctaagaaa ccattattat catgacatta acctataaaa ataggcgtat cacgaggccc tttcgtc //