LOCUS AAV pCAG-FLEX-EGFP-WPRE 6555 bp ds-DNA circular SYN 12-MAY-2021
DEFINITION Can be used to generate AAV virus that will express EGFP in the
presence of Cre.
ACCESSION .
VERSION .
KEYWORDS AAV pCAG-FLEX-EGFP-WPRE
SOURCE synthetic DNA construct
ORGANISM synthetic DNA construct
REFERENCE 1 (bases 1 to 6555)
AUTHORS Oh SW, Harris JA, Ng L, Winslow B, Cain N, Mihalas S, Wang Q, Lau C,
Kuan L, Henry AM, Mortrud MT, Ouellette B, Nguyen TN, Sorensen SA,
Slaughterbeck CR, Wakeman W, Li Y, Feng D, Ho A, Nicholas E,
Hirokawa KE, Bohn P, Joines KM, Peng H, Hawrylycz MJ, Phillips JW,
Hohmann JG, Wohnoutka P, Gerfen CR, Koch C, Bernard A, Dang C, Jones
AR, Zeng H
TITLE A mesoscale connectome of the mouse brain.
JOURNAL Nature. 2014 Apr 10;508(7495):207-14. doi: 10.1038/nature13186. Epub
2014 Apr 2.
PUBMED 24695228
REFERENCE 2 (bases 1 to 6555)
AUTHORS .
TITLE Direct Submission
JOURNAL Exported May 12, 2021 from SnapGene Server 1.1.58
http://www.snapgene.com
FEATURES Location/Qualifiers
source 1..6555
/organism="synthetic DNA construct"
/mol_type="other DNA"
repeat_region 1..130
/label=AAV2 ITR (alternate)
/note="Functional equivalent of wild-type AAV2 ITR"
enhancer 223..526
/label=CMV enhancer
/note="human cytomegalovirus immediate early enhancer"
promoter 528..805
/label=chicken beta-actin promoter
intron 807..1824
/label=chimeric intron
/note="chimera between introns from chicken beta-actin and
rabbit beta-globin"
primer_bind 1748..1770
/label=pCAGGS-5
/note="Chimeric intron in CAG promoter, forward primer"
primer_bind 1832..1851
/label=pCAG-F
/note="Rabbit beta-globin intron, for pCAG plasmids,
forward primer"
protein_bind 1928..1961
/label=loxP
/bound_moiety="Cre recombinase"
/note="Cre-mediated recombination occurs in the 8-bp core
sequence (GCATACAT)."
protein_bind 1972..2005
/label=lox2272
/bound_moiety="Cre recombinase"
/note="Cre-mediated recombination occurs in the 8-bp core
sequence (GGATACTT). lox2272 sites are compatible with each
other, but incompatible with loxP or loxN sites (Lee and
Saito, 1988)."
CDS complement(2061..2780)
/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 complement(2064..2780)
/codon_start=1
/product="enhanced GFP"
/label=EGFP
/translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
EFVTAAGITLGMDELYK"
primer_bind complement(2106..2127)
/label=EGFP-C
/note="EGFP, forward primer"
primer_bind 2455..2474
/label=EXFP-R
/note="For distinguishing EGFP variants, reverse primer"
primer_bind 2714..2735
/label=EGFP-N
/note="EGFP, reverse primer"
regulatory 2777..2786
/regulatory_class="other"
/note="vertebrate consensus sequence for strong initiation
of translation (Kozak, 1987)"
protein_bind complement(2858..2891)
/label=loxP
/bound_moiety="Cre recombinase"
/note="Cre-mediated recombination occurs in the 8-bp core
sequence (GCATACAT)."
protein_bind complement(2902..2935)
/label=lox2272
/bound_moiety="Cre recombinase"
/note="Cre-mediated recombination occurs in the 8-bp core
sequence (GGATACTT). lox2272 sites are compatible with each
other, but incompatible with loxP or loxN sites (Lee and
Saito, 1988)."
misc_feature 2959..3547
/label=WPRE
/note="woodchuck hepatitis virus posttranscriptional
regulatory element"
primer_bind complement(3012..3032)
/label=WPRE-R
/note="WPRE, reverse primer"
CDS complement(3430..3441)
/codon_start=1
/product="Factor Xa recognition and cleavage site"
/label=Factor Xa site
/translation="IEGR"
primer_bind complement(3582..3599)
/label=BGH-rev
/note="Bovine growth hormone terminator, reverse primer.
Also called BGH reverse"
polyA_signal 3588..3795
/label=bGH poly(A) signal
/note="bovine growth hormone polyadenylation signal"
repeat_region 3818..3958
/label=AAV2 ITR
/note="inverted terminal repeat of adeno-associated virus
serotype 2"
repeat_region 3818..3947
/label=AAV2 ITR
rep_origin 4033..4488
/direction=RIGHT
/label=f1 ori
/note="f1 bacteriophage origin of replication; arrow
indicates direction of (+) strand synthesis"
primer_bind complement(4120..4139)
/label=F1ori-R
/note="F1 origin, reverse primer"
primer_bind 4330..4351
/label=F1ori-F
/note="F1 origin, forward primer"
primer_bind complement(4505..4524)
/label=pRS-marker
/note="pRS vectors, use to sequence yeast selectable
marker"
primer_bind 4624..4646
/label=pGEX 3'
/note="pGEX vectors, reverse primer"
primer_bind complement(4684..4702)
/label=pBRforEco
/note="pBR322 vectors, upsteam of EcoRI site, forward
primer"
promoter 4770..4874
/gene="bla"
/label=AmpR promoter
CDS 4875..5735
/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(5093..5112)
/label=Amp-R
/note="Ampicillin resistance gene, reverse primer"
rep_origin 5906..6494
/direction=RIGHT
/label=ori
/note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
replication"
primer_bind 6395..6414
/label=pBR322ori-F
/note="pBR322 origin, forward primer"
ORIGIN
1 cctgcaggca gctgcgcgct cgctcgctca ctgaggccgc ccgggcgtcg ggcgaccttt
61 ggtcgcccgg cctcagtgag cgagcgagcg cgcagagagg gagtggccaa ctccatcact
121 aggggttcct gcggccgcac gcgaaacaat tctgcaggaa tctagttatt aatagtaatc
181 aattacgggg tcattagttc atagcccata tatggagttc cgcgttacat aacttacggt
241 aaatggcccg cctggctgac cgcccaacga cccccgccca ttgacgtcaa taatgacgta
301 tgttcccata gtaacgccaa tagggacttt ccattgacgt caatgggtgg agtatttacg
361 gtaaactgcc cacttggcag tacatcaagt gtatcatatg ccaagtacgc cccctattga
421 cgtcaatgac ggtaaatggc ccgcctggca ttatgcccag tacatgacct tatgggactt
481 tcctacttgg cagtacatct acgtattagt catcgctatt accatggtcg aggtgagccc
541 cacgttctgc ttcactctcc ccatctcccc cccctcccca cccccaattt tgtatttatt
601 tattttttaa ttattttgtg cagcgatggg ggcggggggg gggggggggc gcgcgccagg
661 cggggcgggg cggggcgagg ggcggggcgg ggcgaggcgg agaggtgcgg cggcagccaa
721 tcagagcggc gcgctccgaa agtttccttt tatggcgagg cggcggcggc ggcggcccta
781 taaaaagcga agcgcgcggc gggcggggag tcgctgcgac gctgccttcg ccccgtgccc
841 cgctccgccg ccgcctcgcg ccgcccgccc cggctctgac tgaccgcgtt actcccacag
901 gtgagcgggc gggacggccc ttctcctccg ggctgtaatt agcgcttggt ttaatgacgg
961 cttgtttctt ttctgtggct gcgtgaaagc cttgaggggc tccgggaggg ccctttgtgc
1021 ggggggagcg gctcgggggg tgcgtgcgtg tgtgtgtgcg tggggagcgc cgcgtgcggc
1081 tccgcgctgc ccggcggctg tgagcgctgc gggcgcggcg cggggctttg tgcgctccgc
1141 agtgtgcgcg aggggagcgc ggccgggggc ggtgccccgc ggtgcggggg gggctgcgag
1201 gggaacaaag gctgcgtgcg gggtgtgtgc gtgggggggt gagcaggggg tgtgggcgcg
1261 tcggtcgggc tgcaaccccc cctgcacccc cctccccgag ttgctgagca cggcccggct
1321 tcgggtgcgg ggctccgtac ggggcgtggc gcggggctcg ccgtgccggg cggggggtgg
1381 cggcaggtgg gggtgccggg cggggcgggg ccgcctcggg ccggggaggg ctcgggggag
1441 gggcgcggcg gcccccggag cgccggcggc tgtcgaggcg cggcgagccg cagccattgc
1501 cttttatggt aatcgtgcga gagggcgcag ggacttcctt tgtcccaaat ctgtgcggag
1561 ccgaaatctg ggaggcgccg ccgcaccccc tctagcgggc gcggggcgaa gcggtgcggc
1621 gccggcagga aggaaatggg cggggagggc cttcgtgcgt cgccgcgccg ccgtcccctt
1681 ctccctctcc agcctcgggg ctgtccgcgg ggggacggct gccttcgggg gggacggggc
1741 agggcggggt tcggcttctg gcgtgtgacc ggcggctcta gagcctctgc taaccatgtt
1801 catgccttct tctttttcct acagctcctg ggcaacgtgc tggttattgt gctgtctcat
1861 cattttggca aagaattgat ttgataccgc gggccccgcg tggacggtac ccttcaggtc
1921 gacgtcgata acttcgtata gcatacatta tacgaagtta tccgaatcgc aataacttcg
1981 tataggatac tttatacgaa gttatttggc gcgccaaatt taaataacgc aacgtacctt
2041 cgagggggcc caaggaattc ttacttgtac agctcgtcca tgccgagagt gatcccggcg
2101 gcggtcacga actccagcag gaccatgtga tcgcgcttct cgttggggtc tttgctcagg
2161 gcggactggg tgctcaggta gtggttgtcg ggcagcagca cggggccgtc gccgatgggg
2221 gtgttctgct ggtagtggtc ggcgagctgc acgctgccgt cctcgatgtt gtggcggatc
2281 ttgaagttca ccttgatgcc gttcttctgc ttgtcggcca tgatatagac gttgtggctg
2341 ttgtagttgt actccagctt gtgccccagg atgttgccgt cctccttgaa gtcgatgccc
2401 ttcagctcga tgcggttcac cagggtgtcg ccctcgaact tcacctcggc gcgggtcttg
2461 tagttgccgt cgtccttgaa gaagatggtg cgctcctgga cgtagccttc gggcatggcg
2521 gacttgaaga agtcgtgctg cttcatgtgg tcggggtagc ggctgaagca ctgcacgccg
2581 taggtcaggg tggtcacgag ggtgggccag ggcacgggca gcttgccggt ggtgcagatg
2641 aacttcaggg tcagcttgcc gtaggtggca tcgccctcgc cctcgccgga cacgctgaac
2701 ttgtggccgt ttacgtcgcc gtccagctcg accaggatgg gcaccacccc ggtgaacagc
2761 tcctcgccct tgctcaccat ggtggcgaat tcccccaagc ttggaacgta ctaaccttga
2821 tagctagcta gttggcgcgc caacctggcc ggccttgata acttcgtata atgtatgcta
2881 tacgaagtta tccgaatcgc aataacttcg tataaagtat cctatacgaa gttatcgaag
2941 atcatcaagc ttatcgataa tcaacctctg gattacaaaa tttgtgaaag attgactggt
3001 attcttaact atgttgctcc ttttacgcta tgtggatacg ctgctttaat gcctttgtat
3061 catgctattg cttcccgtat ggctttcatt ttctcctcct tgtataaatc ctggttgctg
3121 tctctttatg aggagttgtg gcccgttgtc aggcaacgtg gcgtggtgtg cactgtgttt
3181 gctgacgcaa cccccactgg ttggggcatt gccaccacct gtcagctcct ttccgggact
3241 ttcgctttcc ccctccctat tgccacggcg gaactcatcg ccgcctgcct tgcccgctgc
3301 tggacagggg ctcggctgtt gggcactgac aattccgtgg tgttgtcggg gaaatcatcg
3361 tcctttcctt ggctgctcgc ctatgttgcc acctggattc tgcgcgggac gtccttctgc
3421 tacgtccctt cggccctcaa tccagcggac cttccttccc gcggcctgct gccggctctg
3481 cggcctcttc cgcgtcttcg ccttcgccct cagacgagtc ggatctccct ttgggccgcc
3541 tccccgcatc gataccgagc gctgctcgag agatcgatct gcctcgactg tgccttctag
3601 ttgccagcca tctgttgttt gcccctcccc cgtgccttcc ttgaccctgg aaggtgccac
3661 tcccactgtc ctttcctaat aaaatgagga aattgcatcg cattgtctga gtaggtgtca
3721 ttctattctg gggggtgggg tggggcagga cagcaagggg gaggattggg aagacaatag
3781 caggcatgct ggggacacgt gcggaccgag cggccgcagg aacccctagt gatggagttg
3841 gccactccct ctctgcgcgc tcgctcgctc actgaggccg ggcgaccaaa ggtcgcccga
3901 cgcccgggct ttgcccgggc ggcctcagtg agcgagcgag cgcgcagctg cctgcagggg
3961 cgcctgatgc ggtattttct ccttacgcat ctgtgcggta tttcacaccg catacgtcaa
4021 agcaaccata gtacgcgccc tgtagcggcg cattaagcgc ggcgggtgtg gtggttacgc
4081 gcagcgtgac cgctacactt gccagcgcct tagcgcccgc tcctttcgct ttcttccctt
4141 cctttctcgc cacgttcgcc ggctttcccc gtcaagctct aaatcggggg ctccctttag
4201 ggttccgatt tagtgcttta cggcacctcg accccaaaaa acttgatttg ggtgatggtt
4261 cacgtagtgg gccatcgccc tgatagacgg tttttcgccc tttgacgttg gagtccacgt
4321 tctttaatag tggactcttg ttccaaactg gaacaacact caactctatc tcgggctatt
4381 cttttgattt ataagggatt ttgccgattt cggtctattg gttaaaaaat gagctgattt
4441 aacaaaaatt taacgcgaat tttaacaaaa tattaacgtt tacaatttta tggtgcactc
4501 tcagtacaat ctgctctgat gccgcatagt taagccagcc ccgacacccg ccaacacccg
4561 ctgacgcgcc ctgacgggct tgtctgctcc cggcatccgc ttacagacaa gctgtgaccg
4621 tctccgggag ctgcatgtgt cagaggtttt caccgtcatc accgaaacgc gcgagacgaa
4681 agggcctcgt gatacgccta tttttatagg ttaatgtcat gataataatg gtttcttaga
4741 cgtcaggtgg cacttttcgg ggaaatgtgc gcggaacccc tatttgttta tttttctaaa
4801 tacattcaaa tatgtatccg ctcatgagac aataaccctg ataaatgctt caataatatt
4861 gaaaaaggaa gagtatgagt attcaacatt tccgtgtcgc ccttattccc ttttttgcgg
4921 cattttgcct tcctgttttt gctcacccag aaacgctggt gaaagtaaaa gatgctgaag
4981 atcagttggg tgcacgagtg ggttacatcg aactggatct caacagcggt aagatccttg
5041 agagttttcg ccccgaagaa cgttttccaa tgatgagcac ttttaaagtt ctgctatgtg
5101 gcgcggtatt atcccgtatt gacgccgggc aagagcaact cggtcgccgc atacactatt
5161 ctcagaatga cttggttgag tactcaccag tcacagaaaa gcatcttacg gatggcatga
5221 cagtaagaga attatgcagt gctgccataa ccatgagtga taacactgcg gccaacttac
5281 ttctgacaac gatcggagga ccgaaggagc taaccgcttt tttgcacaac atgggggatc
5341 atgtaactcg ccttgatcgt tgggaaccgg agctgaatga agccatacca aacgacgagc
5401 gtgacaccac gatgcctgta gcaatggcaa caacgttgcg caaactatta actggcgaac
5461 tacttactct agcttcccgg caacaattaa tagactggat ggaggcggat aaagttgcag
5521 gaccacttct gcgctcggcc cttccggctg gctggtttat tgctgataaa tctggagccg
5581 gtgagcgtgg gtctcgcggt atcattgcag cactggggcc agatggtaag ccctcccgta
5641 tcgtagttat ctacacgacg gggagtcagg caactatgga tgaacgaaat agacagatcg
5701 ctgagatagg tgcctcactg attaagcatt ggtaactgtc agaccaagtt tactcatata
5761 tactttagat tgatttaaaa cttcattttt aatttaaaag gatctaggtg aagatccttt
5821 ttgataatct catgaccaaa atcccttaac gtgagttttc gttccactga gcgtcagacc
5881 ccgtagaaaa gatcaaagga tcttcttgag atcctttttt tctgcgcgta atctgctgct
5941 tgcaaacaaa aaaaccaccg ctaccagcgg tggtttgttt gccggatcaa gagctaccaa
6001 ctctttttcc gaaggtaact ggcttcagca gagcgcagat accaaatact gttcttctag
6061 tgtagccgta gttaggccac cacttcaaga actctgtagc accgcctaca tacctcgctc
6121 tgctaatcct gttaccagtg gctgctgcca gtggcgataa gtcgtgtctt accgggttgg
6181 actcaagacg atagttaccg gataaggcgc agcggtcggg ctgaacgggg ggttcgtgca
6241 cacagcccag cttggagcga acgacctaca ccgaactgag atacctacag cgtgagctat
6301 gagaaagcgc cacgcttccc gaagggagaa aggcggacag gtatccggta agcggcaggg
6361 tcggaacagg agagcgcacg agggagcttc cagggggaaa cgcctggtat ctttatagtc
6421 ctgtcgggtt tcgccacctc tgacttgagc gtcgattttt gtgatgctcg tcaggggggc
6481 ggagcctatg gaaaaacgcc agcaacgcgg cctttttacg gttcctggcc ttttgctggc
6541 cttttgctca catgt
//
