LOCUS Exported 6721 bp ds-DNA circular SYN 12-MAY-2021 DEFINITION This vector is used for cloning a gene driven by 4xnrUAS, to be co-expressed with EGFP through P2A self-cleaving peptides.. ACCESSION . VERSION . KEYWORDS pBT2-4xUAS:P2A-EGFP SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 6721) AUTHORS Gu Q, Yang X, Lv J, Zhang J, Xia B, Kim JD, Wang R, Xiong F, Meng S, Clements TP, Tandon B, Wagner DS, Diaz MF, Wenzel PL, Miller YI, Traver D, Cooke JP, Li W, Zon LI, Chen K, Bai Y, Fang L TITLE AIBP-mediated cholesterol efflux instructs hematopoietic stem and progenitor cell fate. JOURNAL Science. 2019 Mar 8;363(6431):1085-1088. doi: 10.1126/science.aav1749. Epub 2019 Jan 31. PUBMED 30705153 REFERENCE 2 (bases 1 to 6721) AUTHORS . TITLE Direct Submission JOURNAL Exported May 12, 2021 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..6721 /organism="synthetic DNA construct" /mol_type="other DNA" primer_bind 1289..1305 /label=pBluescriptKS /note="For pBluescript vector" primer_bind 1290..1306 /label=KS primer /note="common sequencing primer, one of multiple similar variants" CDS 1610..1666 /codon_start=1 /product="2A peptide from porcine teschovirus-1 polyprotein" /label=P2A /note="Eukaryotic ribosomes fail to insert a peptide bond between the Gly and Pro residues, yielding separate polypeptides." /translation="ATNFSLLKQAGDVEENPGP" CDS 1673..2392 /codon_start=1 /product="the original enhanced GFP (Yang et al., 1996)" /label=EGFP /note="mammalian codon-optimized" /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL EFVTAAGITLGMDELYK" CDS 1673..2389 /codon_start=1 /product="enhanced GFP" /label=EGFP /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL EFVTAAGITLGMDELYK" primer_bind complement(1718..1739) /label=EGFP-N /note="EGFP, reverse primer" primer_bind complement(1979..1998) /label=EXFP-R /note="For distinguishing EGFP variants, reverse primer" primer_bind 2326..2347 /label=EGFP-C /note="EGFP, forward primer" promoter complement(2409..2426) /label=T7 promoter /note="promoter for bacteriophage T7 RNA polymerase" polyA_signal 2431..2565 /label=SV40 poly(A) signal /note="SV40 polyadenylation signal" primer_bind complement(2455..2474) /label=EBV-rev /note="SV40 polyA terminator, reverse primer" primer_bind 2509..2528 /label=SV40pA-R /note="SV40 polyA, reverse primer" primer_bind complement(3874..3893) /label=T7 /note="T7 promoter, forward primer" promoter complement(3875..3893) /label=T7 promoter /note="promoter for bacteriophage T7 RNA polymerase" primer_bind complement(3900..3917) /label=M13 Forward /note="In lacZ gene. Also called M13-F20 or M13 (-21) Forward" primer_bind complement(3900..3916) /label=M13 fwd /note="common sequencing primer, one of multiple similar variants" primer_bind complement(3909..3931) /label=M13/pUC Forward /note="In lacZ gene" rep_origin 4058..4513 /direction=RIGHT /label=f1 ori /note="f1 bacteriophage origin of replication; arrow indicates direction of (+) strand synthesis" primer_bind complement(4145..4164) /label=F1ori-R /note="F1 origin, reverse primer" primer_bind 4355..4376 /label=F1ori-F /note="F1 origin, forward primer" promoter 4539..4643 /gene="bla" /label=AmpR promoter CDS 4644..5504 /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 complement(4862..4881) /label=Amp-R /note="Ampicillin resistance gene, reverse primer" rep_origin 5675..6263 /direction=RIGHT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" primer_bind 6164..6183 /label=pBR322ori-F /note="pBR322 origin, forward primer" primer_bind 6417..6434 /label=L4440 /note="L4440 vector, forward primer" protein_bind 6551..6572 /label=CAP binding site /bound_moiety="E. coli catabolite activator protein" /note="CAP binding activates transcription in the presence of cAMP." promoter 6587..6617 /label=lac promoter /note="promoter for the E. coli lac operon" protein_bind 6625..6641 /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 6630..6652 /label=M13/pUC Reverse /note="In lacZ gene" primer_bind 6649..6665 /label=M13 rev /note="common sequencing primer, one of multiple similar variants" primer_bind 6649..6665 /label=M13 Reverse /note="In lacZ gene. Also called M13-rev" promoter 6686..6704 /label=T3 promoter /note="promoter for bacteriophage T3 RNA polymerase" ORIGIN 1 cagaggtgta aagtacttga gtaattttac ttgattactg tacttaagta ttatttttgg 61 ggatttttac tttacttgag tacaattaaa aatcaatact tttactttta cttaattaca 121 tttttttaga aaaaaaagta ctttttactc cttacaattt tatttacagt caaaaagtac 181 ttattttttg gagatcactt cattctattt tcccttgcta ttaccaaacc aattgaattg 241 cgctgatgcc cagtttaatt taaatgttat ttattctgcc tatgaaaatc gttttcacat 301 tatatgaaat tggtcagaca tgttcattgg tcctttggaa gtgacgtcat gtcacatcta 361 ttaccacaat gcacagcacc ttgacctgga aattagggaa attataacag tcaatcagtg 421 gaagaaaatg gaggaagtat gtgattcatc agcagctgcg agcagcacag tccaaaatca 481 gccacaggat caagagcacc cgtggccgta tcttcgcgaa ttcttttctt taagtggtgt 541 aaataaagat tcattcaaga tgaaatgtgt cctctgtctc ccgcgggccc tacgaagtta 601 tagatcctct agcgataagc ttgatatcga attcgagttg gcgcgcctgt cattctaaat 661 ctctctttca gcctaaagct ttttccccgt atccccccag gtgtctgcag gctcaaagag 721 cagcgagaag cgttcagagg aaagcgatcc cgtgccacct tccccgtgcc cgggctgtcc 781 ccgcacgctg ccggctcggg gatgcggggg gagcgccgga ccggagcgga gccccgggcg 841 gctcgctgct gccccctagc gggggaggga cgtaattaca tccctggggg ctttgggagg 901 gggctgtccc cgtgagctcc caggcgcgcc tgtcattcta aatctctctt tcagcctaaa 961 gctttttccc cgtatccccc caggtgtctg caggctcaaa gagcagcgag aagcgttcag 1021 aggaaagcga tcccgtgcca ccttccccgt gcccgggctg tccccgcacg ctgccggctc 1081 ggggatgcgg ggggagcgcc ggaccggagc ggagccccgg gcggctcgct gctgccccct 1141 agcgggggag ggacgtaatt acatccctgg gggctttggg ggggggctgt ccccgtgagc 1201 tcaataaaag agcccacaac ccctcactcg gcgcgccagt cctccgatag actgcgtcgc 1261 ccgggtaccc gtgtatccaa taaacccctc gaggtcgacg gtatcgataa gcttgatatc 1321 gaattcctgc agcccggggg atcacgcggc catcaagctt aggcctccac ggtggcttct 1381 aatccgtgag tcctagcggg tgacagccct ccgtcttcac aggcggagga gagtcttccg 1441 tagggttcct cggagtactg tcctccgacg cgtgcaaggg tcgactctag agggtatata 1501 atggatccca tcgcgtctca gcctcacttt gagctcctcc acacgaattc cctcgacctc 1561 gaagacgcgt ggatccaccg gtggatccct cgagcccggg tctagaggag ctactaactt 1621 cagcctgctg aagcaggctg gagacgtgga ggagaaccct ggacctagat ccatggtgag 1681 caagggcgag gagctgttca ccggggtggt gcccatcctg gtcgagctgg acggcgacgt 1741 aaacggccac aagttcagcg tgtccggcga gggcgagggc gatgccacct acggcaagct 1801 gaccctgaag ttcatctgca ccaccggcaa gctgcccgtg ccctggccca ccctcgtgac 1861 caccctgacc tacggcgtgc agtgcttcag ccgctacccc gaccacatga agcagcacga 1921 cttcttcaag tccgccatgc ccgaaggcta cgtccaggag cgcaccatct tcttcaagga 1981 cgacggcaac tacaagaccc gcgccgaggt gaagttcgag ggcgacaccc tggtgaaccg 2041 catcgagctg aagggcatcg acttcaagga ggacggcaac atcctggggc acaagctgga 2101 gtacaactac aacagccaca acgtctatat catggccgac aagcagaaga acggcatcaa 2161 ggtgaacttc aagatccgcc acaacatcga ggacggcagc gtgcagctcg ccgaccacta 2221 ccagcagaac acccccatcg gcgacggccc cgtgctgctg cccgacaacc actacctgag 2281 cacccagtcc gccctgagca aagaccccaa cgagaagcgc gatcacatgg tcctgctgga 2341 gttcgtgacc gccgccggga tcactctcgg catggacgag ctgtacaagt aatcgattct 2401 agctagaact atagtgagtc gtattacgta gatccagaca tgataagata cattgatgag 2461 tttggacaaa ccacaactag aatgcagtga aaaaaatgct ttatttgtga aatttgtgat 2521 gctattgctt tatttgtaac cattataagc tgcaataaac aagttaacaa caacaattgc 2581 attcatttta tgtttcaggt tcagggggag gtgtgggagg ttttttaatt cgcggccgcg 2641 gcgactagtg ggtttattgg atacacgggt acccgggcga cgcagtctat cggaggactg 2701 gcgcgccgag tgaggggttg tgggctcttt tattgagctc acggggacag ccccccccca 2761 aagcccccag ggatgtaatt acgtccctcc cccgctaggg ggcagcagcg agccgcccgg 2821 ggctccgctc cggtccggcg ctccccccgc atccccgagc cggcagcgtg cggggacagc 2881 ccgggcacgg ggaaggtggc acgggatcgc tttcctctga acgcttctcg ctgctctttg 2941 agcctgcaga cacctggggg gatacgggga aaaagcttta ggctgaaaga gagatttaga 3001 atgacaggcg cgcctgggag ctcacgggga cagccccctc ccaaagcccc cagggatgta 3061 attacgtccc tcccccgcta gggggcagca gcgagccgcc cggggctccg ctccggtccg 3121 gcgctccccc cgcatccccg agccggcagc gtgcggggac agcccgggca cggggaaggt 3181 ggcacgggat cgctttcctc tgaacgcttc tcgctgctct ttgagcctgc agacacctgg 3241 ggggatacgg ggaaaaagct ttaggctgaa agagagattt agaatgacag gcgcgccaac 3301 tcgaattcga tatcaagctt atcgctagag gatctataac ttcgtaagat ctgtctctgc 3361 tcacgtttcc tgctatttgc agcctctcta tcaagactaa tacacctctt cccgcatcgg 3421 ctgcctgtga gaggcttttc agcactgcag gattgctttt cagccccaaa agagctaggc 3481 ttgacactaa caattttgag aatcagcttc tactgaagtt aaatctgagg ttttacaact 3541 ttgagtagcg tgtactggca ttagattgtc tgtcttatag tttgataatt aaatacaaac 3601 agttctaaag caggataaaa ccttgtatgc atttcattta atgttttttg agattaaaag 3661 cttaaacaag aatctctagt tttctttctt gcttttactt ttacttcctt aatactcaag 3721 tacaatttta atggagtact tttttacttt tactcaagta agattctagc cagatacttt 3781 tacttttaat tgagtaaaat tttccctaag tacttgtact ttcacttgag taaaattttt 3841 gagtactttt tacacctctg gagctccaat tcgccctata gtgagtcgta ttacaattca 3901 ctggccgtcg ttttacaacg tcgtgactgg gaaaaccctg gcgttaccca acttaatcgc 3961 cttgcagcac atcccccttt cgccagctgg cgtaatagcg aagaggcccg caccgatcgc 4021 ccttcccaac agttgcgcag cctgaatggc gaatgggacg cgccctgtag cggcgcatta 4081 agcgcggcgg gtgtggtggt tacgcgcagc gtgaccgcta cacttgccag cgccctagcg 4141 cccgctcctt tcgctttctt cccttccttt ctcgccacgt tcgccggctt tccccgtcaa 4201 gctctaaatc gggggctccc tttagggttc cgatttagtg ctttacggca cctcgacccc 4261 aaaaaacttg attagggtga tggttcacgt agtgggccat cgccctgata gacggttttt 4321 cgccctttga cgttggagtc cacgttcttt aatagtggac tcttgttcca aactggaaca 4381 acactcaacc ctatctcggt ctattctttt gatttataag ggattttgcc gatttcggcc 4441 tattggttaa aaaatgagct gatttaacaa aaatttaacg cgaattttaa caaaatatta 4501 acgcttacaa tttaggtggc acttttcggg gaaatgtgcg cggaacccct atttgtttat 4561 ttttctaaat acattcaaat atgtatccgc tcatgagaca ataaccctga taaatgcttc 4621 aataatattg aaaaaggaag agtatgagta ttcaacattt ccgtgtcgcc cttattccct 4681 tttttgcggc attttgcctt cctgtttttg ctcacccaga aacgctggtg aaagtaaaag 4741 atgctgaaga tcagttgggt gcacgagtgg gttacatcga actggatctc aacagcggta 4801 agatccttga gagttttcgc cccgaagaac gttttccaat gatgagcact tttaaagttc 4861 tgctatgtgg cgcggtatta tcccgtattg acgccgggca agagcaactc ggtcgccgca 4921 tacactattc tcagaatgac ttggttgagt actcaccagt cacagaaaag catcttacgg 4981 atggcatgac agtaagagaa ttatgcagtg ctgccataac catgagtgat aacactgcgg 5041 ccaacttact tctgacaacg atcggaggac cgaaggagct aaccgctttt ttgcacaaca 5101 tgggggatca tgtaactcgc cttgatcgtt gggaaccgga gctgaatgaa gccataccaa 5161 acgacgagcg tgacaccacg atgcctgtag caatggcaac aacgttgcgc aaactattaa 5221 ctggcgaact acttactcta gcttcccggc aacaattaat agactggatg gaggcggata 5281 aagttgcagg accacttctg cgctcggccc ttccggctgg ctggtttatt gctgataaat 5341 ctggagccgg tgagcgtggg tctcgcggta tcattgcagc actggggcca gatggtaagc 5401 cctcccgtat cgtagttatc tacacgacgg ggagtcaggc aactatggat gaacgaaata 5461 gacagatcgc tgagataggt gcctcactga ttaagcattg gtaactgtca gaccaagttt 5521 actcatatat actttagatt gatttaaaac ttcattttta atttaaaagg atctaggtga 5581 agatcctttt tgataatctc atgaccaaaa tcccttaacg tgagttttcg ttccactgag 5641 cgtcagaccc cgtagaaaag atcaaaggat cttcttgaga tccttttttt ctgcgcgtaa 5701 tctgctgctt gcaaacaaaa aaaccaccgc taccagcggt ggtttgtttg ccggatcaag 5761 agctaccaac tctttttccg aaggtaactg gcttcagcag agcgcagata ccaaatactg 5821 ttcttctagt gtagccgtag ttaggccacc acttcaagaa ctctgtagca ccgcctacat 5881 acctcgctct gctaatcctg ttaccagtgg ctgctgccag tggcgataag tcgtgtctta 5941 ccgggttgga ctcaagacga tagttaccgg ataaggcgca gcggtcgggc tgaacggggg 6001 gttcgtgcac acagcccagc ttggagcgaa cgacctacac cgaactgaga tacctacagc 6061 gtgagctatg agaaagcgcc acgcttcccg aagggagaaa ggcggacagg tatccggtaa 6121 gcggcagggt cggaacagga gagcgcacga gggagcttcc agggggaaac gcctggtatc 6181 tttatagtcc tgtcgggttt cgccacctct gacttgagcg tcgatttttg tgatgctcgt 6241 caggggggcg gagcctatgg aaaaacgcca gcaacgcggc ctttttacgg ttcctggcct 6301 tttgctggcc ttttgctcac atgttctttc ctgcgttatc ccctgattct gtggataacc 6361 gtattaccgc ctttgagtga gctgataccg ctcgccgcag ccgaacgacc gagcgcagcg 6421 agtcagtgag cgaggaagcg gaagagcgcc caatacgcaa accgcctctc cccgcgcgtt 6481 ggccgattca ttaatgcagc tggcacgaca ggtttcccga ctggaaagcg ggcagtgagc 6541 gcaacgcaat taatgtgagt tagctcactc attaggcacc ccaggcttta cactttatgc 6601 ttccggctcg tatgttgtgt ggaattgtga gcggataaca atttcacaca ggaaacagct 6661 atgaccatga ttacgccaag ctcgaaatta accctcacta aagggaacaa aagctggtac 6721 c //