LOCUS Exported 5477 bp ds-DNA circular SYN 03-OCT-2023 DEFINITION attR4-attR3 Destination cassette in Tol1 backbone. ACCESSION . VERSION . KEYWORDS pToneDest SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 5477) AUTHORS Shin M, Male I, Beane TJ, Villefranc JA, Kok FO, Zhu LJ, Lawson ND TITLE Vegfc acts through ERK to induce sprouting and differentiation of trunk lymphatic progenitors. JOURNAL Development. 2016 Oct 15;143(20):3785-3795. Epub 2016 Sep 12. PUBMED 27621059 REFERENCE 2 (bases 1 to 5477) AUTHORS . TITLE Direct Submission JOURNAL Exported Oct 3, 2023 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..5477 /organism="synthetic DNA construct" /mol_type="other DNA" 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" primer_bind 364..386 /label=M13/pUC Forward /note="In lacZ gene" primer_bind 378..395 /label=M13 Forward /note="In lacZ gene. Also called M13-F20 or M13 (-21) Forward" primer_bind 379..395 /label=M13 fwd /note="common sequencing primer, one of multiple similar variants" protein_bind 627..751 /gene="mutant version of attR" /label=attR4 /bound_moiety="LR Clonase(TM)" /note="recombination site for the Gateway(R) LR reaction" CDS complement(1099..1404) /codon_start=1 /gene="ccdB" /product="CcdB, a bacterial toxin that poisons DNA gyrase" /label=ccdB /note="Plasmids containing the ccdB gene cannot be propagated in standard E. coli strains." /translation="MQFKVYTYKRESRYRLFVDVQSDIIDTPGRRMVIPLASARLLSDK VSRELYPVVHIGDESWRMMTTDMASVPVSVIGEEVADLSHRENDIKNAINLMFWGI" primer_bind complement(1201..1220) /label=ccdB-fwd /note="ccdB gene, forward primer" CDS complement(1754..2407) /codon_start=1 /gene="cat" /product="chloramphenicol acetyltransferase" /label=CmR /note="confers resistance to chloramphenicol" /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGG" primer_bind 2322..2341 /label=CAT-R /note="Chloramphenicol resistance gene, reverse primer" promoter complement(2408..2510) /label=cat promoter /note="promoter of the E. coli cat gene encoding chloramphenicol acetyltransferase" protein_bind complement(2656..2779) /gene="mutant version of attR" /label=attR3 /bound_moiety="LR Clonase(TM)" /note="recombination site for the Gateway(R) LR reaction" polyA_signal 2801..2935 /label=SV40 poly(A) signal /note="SV40 polyadenylation signal" primer_bind complement(2825..2844) /label=EBV-rev /note="SV40 polyA terminator, reverse primer" primer_bind 2879..2898 /label=SV40pA-R /note="SV40 polyA, reverse primer" primer_bind complement(3204..3220) /label=M13 rev /note="common sequencing primer, one of multiple similar variants" primer_bind complement(3256..3272) /label=M13 rev /note="common sequencing primer, one of multiple similar variants" primer_bind complement(3269..3291) /label=M13/pUC Reverse /note="In lacZ gene" protein_bind 3280..3296 /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(3304..3334) /label=lac promoter /note="promoter for the E. coli lac operon" protein_bind 3349..3370 /label=CAP binding site /bound_moiety="E. coli catabolite activator protein" /note="CAP binding activates transcription in the presence of cAMP." primer_bind complement(3487..3504) /label=L4440 /note="L4440 vector, forward primer" rep_origin complement(3658..4246) /direction=LEFT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" primer_bind complement(3738..3757) /label=pBR322ori-F /note="pBR322 origin, forward primer" CDS complement(4417..5277) /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 5040..5059 /label=Amp-R /note="Ampicillin resistance gene, reverse primer" promoter complement(5278..5382) /gene="bla" /label=AmpR promoter primer_bind 5450..5468 /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 accatatgcg gtgtgaaata ccgcacagat gcgtaaggag aaaataccgc atcaggcgcc 241 attcgccatt caggctgcgc aactgttggg aagggcgatc ggtgcgggcc tcttcgctat 301 tacgccagct ggcgaaaggg ggatgtgctg caaggcgatt aagttgggta acgccagggt 361 tttcccagtc acgacgttgt aaaacgacgg ccagtgaatt ccctttagcc agtagcggtt 421 ctaggcacgg gccgtccggg cggtggcctg gggcggaaaa ctgaaggggg gcggcaccgg 481 cggctcagcc ctttgtaata tattaatatg caccactatt ggtttactta tgtcacagtt 541 tgtaagtttg taacagcctg aacctggccg cgcgccattg ggatggaacc ctcgaggggc 601 ccgctagcga tatccctgca gggctatcaa ctttgtatag aaaagttgaa cgagaaacgt 661 aaaatgatat aaatatcaat atattaaatt agattttgca taaaaaacag actacataat 721 actgtaaaac acaacatatc cagtcactat ggtcgaccga cagccttcca aatgttcttc 781 tcaaacggaa tcgtcgtatc cagcctactc gctattgtcc tcaatgccgt attaaatcat 841 aaaaagaaat aagaaaaaga ggtgcgagcc tcttttttgt gtgacaaaat aaaaacatct 901 acctattcat atacgctagt gtcatagtcc tgaaaatcat ctgcatcaag aacaatttca 961 caactcttat acttttctct tacaagtcgt tcggcttcat ctggattttc agcctctata 1021 cttactaaac gtgataaagt ttctgtaatt tctactgtat cgacctgcag actggctgtg 1081 tataagggag cctgacattt atattcccca gaacatcagg ttaatggcgt ttttgatgtc 1141 attttcgcgg tggctgagat cagccacttc ttccccgata acggagaccg gcacactggc 1201 catatcggtg gtcatcatgc gccagctttc atccccgata tgcaccaccg ggtaaagttc 1261 acgggagact ttatctgaca gcagacgtgc actggccagg gggatcacca tccgtcgccc 1321 gggcgtgtca ataatatcac tctgtacatc cacaaacaga cgataacggc tctctctttt 1381 ataggtgtaa accttaaact gcatttcacc agcccctgtt ctcgtcagca aaagagccgt 1441 tcatttcaat aaaccgggcg acctcagcca tcccttcctg attttccgct ttccagcgtt 1501 cggcacgcag acgacgggct tcattctgca tggttgtgct taccagaccg gagatattga 1561 catcatatat gccttgagca actgatagct gtcgctgtca actgtcactg taatacgctg 1621 cttcatagca tacctctttt tgacatactt cgggtataca tatcagtata tattcttata 1681 ccgcaaaaat cagcgcgcaa atacgcatac tgttatctgg cttttagtaa gccggatcca 1741 cgcggcgttt acgccccccc tgccactcat cgcagtactg ttgtaattca ttaagcattc 1801 tgccgacatg gaagccatca caaacggcat gatgaacctg aatcgccagc ggcatcagca 1861 ccttgtcgcc ttgcgtataa tatttgccca tggtgaaaac gggggcgaag aagttgtcca 1921 tattggccac gtttaaatca aaactggtga aactcaccca gggattggct gagacgaaaa 1981 acatattctc aataaaccct ttagggaaat aggccaggtt ttcaccgtaa cacgccacat 2041 cttgcgaata tatgtgtaga aactgccgga aatcgtcgtg gtattcactc cagagcgatg 2101 aaaacgtttc agtttgctca tggaaaacgg tgtaacaagg gtgaacacta tcccatatca 2161 ccagctcacc gtctttcatt gccatacgga attccggatg agcattcatc aggcgggcaa 2221 gaatgtgaat aaaggccgga taaaacttgt gcttattttt ctttacggtc tttaaaaagg 2281 ccgtaatatc cagctgaacg gtctggttat aggtacattg agcaactgac tgaaatgcct 2341 caaaatgttc tttacgatgc cattgggata tatcaacggt ggtatatcca gtgatttttt 2401 tctccatttt agcttcctta gctcctgaaa atctcgataa ctcaaaaaat acgcccggta 2461 gtgatcttat ttcattatgg tgaaagttgg aacctcttac gtgccgatca acgtctcatt 2521 ttcgccaaaa gttggcccag ggcttcccgg tatcaacagg gacaccagga tttatttatt 2581 ctgcgaagtg atcttccgtc acaggtattt attcggcgca aagtgcgtcg ggtgatgctg 2641 ccaacttacg gccgccatag tgactggata tgttgtgttt tacagtatta tgtagtctgt 2701 tttttatgca aaatctaatt taatatattg atatttatat cattttacgt ttctcgttca 2761 actttattat acatagttga taaggcgcgc caggcctgta gatccagaca tgataagata 2821 cattgatgag tttggacaaa ccacaactag aatgcagtga aaaaaatgct ttatttgtga 2881 aatttgtgat gctattgctt tatttgtaac cattataagc tgcaataaac aagttaacaa 2941 caacaattgc attcatttta tgtttcaggt tcagggggag gtgtgggagg ttttttaatt 3001 aaaacatttc agttgacgaa gacaaacaaa gttctgttgt gactatgggg ggggggggcg 3061 cctggggatg gtctcgcccg gggagtaatt cagggtagaa ccgccactgg ctaaagggag 3121 ctcggtacct cgcgaatgca tctagatatc ggatcccggg cccgtcgact gcagaggcct 3181 gcatgcaagc ttgcgtaatc atggtcatag ctgtttcctg tatcccaatg gcgcgccgag 3241 cttggcgtaa tcatggtcat agctgtttcc tgtgtgaaat tgttatccgc tcacaattcc 3301 acacaacata cgagccggaa gcataaagtg taaagcctgg ggtgcctaat gagtgagcta 3361 actcacatta attgcgttgc gctcactgcc cgctttccag tcgggaaacc tgtcgtgcca 3421 gctgcattaa tgaatcggcc aacgcgcggg gagaggcggt ttgcgtattg ggcgctcttc 3481 cgcttcctcg ctcactgact cgctgcgctc ggtcgttcgg ctgcggcgag cggtatcagc 3541 tcactcaaag gcggtaatac ggttatccac agaatcaggg gataacgcag gaaagaacat 3601 gtgagcaaaa ggccagcaaa aggccaggaa ccgtaaaaag gccgcgttgc tggcgttttt 3661 ccataggctc cgcccccctg acgagcatca caaaaatcga cgctcaagtc agaggtggcg 3721 aaacccgaca ggactataaa gataccaggc gtttccccct ggaagctccc tcgtgcgctc 3781 tcctgttccg accctgccgc ttaccggata cctgtccgcc tttctccctt cgggaagcgt 3841 ggcgctttct catagctcac gctgtaggta tctcagttcg gtgtaggtcg ttcgctccaa 3901 gctgggctgt gtgcacgaac cccccgttca gcccgaccgc tgcgccttat ccggtaacta 3961 tcgtcttgag tccaacccgg taagacacga cttatcgcca ctggcagcag ccactggtaa 4021 caggattagc agagcgaggt atgtaggcgg tgctacagag ttcttgaagt ggtggcctaa 4081 ctacggctac actagaagaa cagtatttgg tatctgcgct ctgctgaagc cagttacctt 4141 cggaaaaaga gttggtagct cttgatccgg caaacaaacc accgctggta gcggtggttt 4201 ttttgtttgc aagcagcaga ttacgcgcag aaaaaaagga tctcaagaag atcctttgat 4261 cttttctacg gggtctgacg ctcagtggaa cgaaaactca cgttaaggga ttttggtcat 4321 gagattatca aaaaggatct tcacctagat ccttttaaat taaaaatgaa gttttaaatc 4381 aatctaaagt atatatgagt aaacttggtc tgacagttac caatgcttaa tcagtgaggc 4441 acctatctca gcgatctgtc tatttcgttc atccatagtt gcctgactcc ccgtcgtgta 4501 gataactacg atacgggagg gcttaccatc tggccccagt gctgcaatga taccgcgaga 4561 cccacgctca ccggctccag atttatcagc aataaaccag ccagccggaa gggccgagcg 4621 cagaagtggt cctgcaactt tatccgcctc catccagtct attaattgtt gccgggaagc 4681 tagagtaagt agttcgccag ttaatagttt gcgcaacgtt gttgccattg ctacaggcat 4741 cgtggtgtca cgctcgtcgt ttggtatggc ttcattcagc tccggttccc aacgatcaag 4801 gcgagttaca tgatccccca tgttgtgcaa aaaagcggtt agctccttcg gtcctccgat 4861 cgttgtcaga agtaagttgg ccgcagtgtt atcactcatg gttatggcag cactgcataa 4921 ttctcttact gtcatgccat ccgtaagatg cttttctgtg actggtgagt actcaaccaa 4981 gtcattctga gaatagtgta tgcggcgacc gagttgctct tgcccggcgt caatacggga 5041 taataccgcg ccacatagca gaactttaaa agtgctcatc attggaaaac gttcttcggg 5101 gcgaaaactc tcaaggatct taccgctgtt gagatccagt tcgatgtaac ccactcgtgc 5161 acccaactga tcttcagcat cttttacttt caccagcgtt tctgggtgag caaaaacagg 5221 aaggcaaaat gccgcaaaaa agggaataag ggcgacacgg aaatgttgaa tactcatact 5281 cttccttttt caatattatt gaagcattta tcagggttat tgtctcatga gcggatacat 5341 atttgaatgt atttagaaaa ataaacaaat aggggttccg cgcacatttc cccgaaaagt 5401 gccacctgac gtctaagaaa ccattattat catgacatta acctataaaa ataggcgtat 5461 cacgaggccc tttcgtc //