LOCUS Exported 2415 bp ds-DNA circular SYN 13-MAY-2021 DEFINITION Alpha level R6K_ origin GB Cloning vector, Conditionally replicative ori, requires pir gene expression requires EC100D or EC100D116 bacterial strain (DH10B derived) uses blue-white screening. ACCESSION . VERSION . KEYWORDS pR6Kg-Alpha1 SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 2415) TITLE Multiplexed drug-based selection and counterselection genetic manipulations in Drosophila REFERENCE 2 (bases 1 to 2415) AUTHORS . TITLE Direct Submission JOURNAL Exported May 13, 2021 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..2415 /organism="synthetic DNA construct" /mol_type="other DNA" terminator 15..42 /label=rrnB T2 terminator /note="transcription terminator T2 from the E. coli rrnB gene" primer_bind 183..200 /label=M13 Forward /note="In lacZ gene. Also called M13-F20 or M13 (-21) Forward" primer_bind 184..200 /label=M13 fwd /note="common sequencing primer, one of multiple similar variants" primer_bind 207..226 /label=T7 /note="T7 promoter, forward primer" promoter 207..225 /label=T7 promoter /note="promoter for bacteriophage T7 RNA polymerase" promoter 274..321 /label=EM7 promoter /note="synthetic bacterial promoter " primer_bind complement(702..721) /label=pRS-marker /note="pRS vectors, use to sequence yeast selectable marker" promoter complement(772..790) /label=SP6 promoter /note="promoter for bacteriophage SP6 RNA polymerase" primer_bind complement(773..790) /label=SP6 /note="SP6 promoter, forward primer" primer_bind complement(808..824) /label=M13 rev /note="common sequencing primer, one of multiple similar variants" primer_bind complement(808..824) /label=M13 Reverse /note="In lacZ gene. Also called M13-rev" terminator 895..967 /label=rrnB T1 terminator /note="transcription terminator T1 from the E. coli rrnB gene" rep_origin 1023..1379 /label=R6K gamma ori /note="gamma replication origin from E. coli plasmid R6K; requires the R6K initiator protein pi for replication" CDS complement(1493..2308) /codon_start=1 /gene="aph(3')-Ia" /product="aminoglycoside phosphotransferase" /label=KanR /note="confers resistance to kanamycin in bacteria or G418 (Geneticin(R)) in eukaryotes" /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF" primer_bind 2216..2235 /label=Kan-R /note="Kanamycin resistance gene, reverse primer" promoter complement(2309..2411) /label=cat promoter /note="promoter of the E. coli cat gene encoding chloramphenicol acetyltransferase" ORIGIN 1 attgtagaaa cgcaaaaagg ccatccgtca ggatggcctt ctgcttaatt tgatgcctgg 61 cagtttatgg cgggcgtcct gcccgccacc ctttcagcca aaaaacttaa gaccgccggt 121 cttgtccact accttgcagt aatgcggtgg acaggatcgg cggttttctt ttctcttctc 181 aatgtaaaac gacggccagt gaattgtaat acgactcact atagggttcg aattctatcg 241 gtaagcacaa atcgtctcag gagagagacc cctgttgaca attaatcatc ggcatagtat 301 atcggcatag tataatacga caaggtgagg aactaaaccc aggaggcaga gatcatgacc 361 atgattacgc caagctatct gggcgatact atcgaatact ctagcttgca tgcgtgcagg 421 tcgactctag aagatccccg ggtaccgagc tctaattcgc cgtacagcga gtcttactac 481 aattctctgg cggtggtctt gcaacgtcgt gactgggaaa accctggcgt tacccaactt 541 aatcgccttg cagcacatcc ccctttcgcc agctggcgta atagcgaaga ggcccgcacc 601 gatcgccctt cccaacagtt gcgcagcctg aatggcgaat gacgcctgat gcggtatttt 661 ctccttacgt tactgtgcgg tatttcacac gtgcacctca gtacaatctg ctctgatgcc 721 gcatagggtc tctcgctgtc atgagacgaa ttctgacttg tattccgcgg ttctatagtg 781 tcacctaaat agcttggcgt aatttaggtc atagctgttt cctgtcggac caaaacgaaa 841 aaaggccccc ctttcgggag gcctcttttc tggaatttgg taccgagcag gcatcaaata 901 aaacgaaagg ctcagtcgaa agactgggcc tttcgtttta tctgttgttt gtcggttgaa 961 cgctctctga cgtaccgcgg ccgcgtagag gatctgttga tcagcagttc aacctgttga 1021 tagtacgtac taagctctca tgtttcacgt actaagctct catgtttaac gtactaagct 1081 ctcatgttta acgaactaaa ccctcatggc taacgtacta agctctcatg gctaacgtac 1141 taagctctca tgtttcacgt actaagctct catgtttgaa caataaaatt aatataaatc 1201 agcaacttaa atagcctcta aggttttaag ttttataaga aaaaaaagaa tatataaggc 1261 ttttaaagct tttaaggttt aacggttgtg gacaacaagc cagggatgta acgcactgag 1321 aagcccttag agcctctcaa agcaattttg agtgacacag gaacacttaa cggctgacat 1381 gtgatcatgt ttgacagctt atcatcgaat ttctgccatt catccgctta ttatcactta 1441 ttcaggcgta gcaaccaggc gtttaagggc accaataact gccttaaaaa aattagaaaa 1501 actcatcgag catcaaatga aactgcaatt tattcatatc aggattatca ataccatatt 1561 tttgaaaaag ccgtttctgt aatgaaggag aaaactcacc gaggcagttc cataggatgg 1621 caagatcctg gtatcggtct gcgattccga ctcgtccaac atcaatacaa cctatcaatt 1681 tcccctcgtc aaaaataagg ttatcaagtg agaaatcacc atgagtgact actgaatccg 1741 gtgagaatgg caaaagtttg tgcatttctt tccagacttg ttcaacaggc cagccattac 1801 gctcgtcatc aaaatcactc gcatcaacca aaccgttatt cattcgtgat tgcgcctgag 1861 cgagtcgaaa tacgcggtcg ctgttaaaag gacaattaca aacaggaatc gaatgcaacc 1921 ggcgcaggaa cactgccagc gcatcaacaa tgttttcacc tgaatcagga tattcttcta 1981 atacctggaa tgctgttttc cctgggatcg cagtggtgag taaccacgca tcatcaggag 2041 tacggataaa atgcttgatg gtcggaagag gcataaattc cgtcagccag tttagtctga 2101 ccatctcatc tgtaacatca ttggcaacgc tacctttgcc atgtttcaga aacaactctg 2161 gcgcatcggg cttcccatac aatcggtaga ttgtcgcacc tgattgcccg acattatcgc 2221 gagcccattt atacccatat aaatcagcat ccatgttgga atttaatcgc ggccttgagc 2281 aagacgtttc ccgttgaata tggctcattt tagcttcctt agctcctgaa aatctcgata 2341 actcaaaaaa tacgcccggt agtgatctta tttcattatg gtgaaagttg gaacctctta 2401 cgtgccgatc ataac //