LOCUS pSIN-PAmCherry-KFERQ-NE 8368 bp ds-DNA circular SYN 12-MAY-2021
DEFINITION Lentiviral overexpression of PA-mCherry-KFERQ fusion.
ACCESSION .
VERSION .
KEYWORDS pSIN-PAmCherry-KFERQ-NE
SOURCE synthetic DNA construct
ORGANISM synthetic DNA construct
REFERENCE 1 (bases 1 to 8368)
AUTHORS Ho PW, Leung CT, Liu H, Pang SY, Lam CS, Xian J, Li L, Kung MH,
Ramsden DB, Ho SL
TITLE Age-dependent accumulation of oligomeric SNCA/alpha-synuclein from
impaired degradation in mutant LRRK2 knockin mouse model of
Parkinson disease: role for therapeutic activation of
chaperone-mediated autophagy (CMA).
JOURNAL Autophagy. 2019 Apr 14:1-24. doi: 10.1080/15548627.2019.1603545.
PUBMED 30983487
REFERENCE 2 (bases 1 to 8368)
AUTHORS .
TITLE Direct Submission
JOURNAL Exported May 12, 2021 from SnapGene Server 1.1.58
http://www.snapgene.com
FEATURES Location/Qualifiers
source 1..8368
/organism="synthetic DNA construct"
/mol_type="other DNA"
promoter 1..1179
/label=EF-1-alpha promoter
/note="strong constitutive promoter for human elongation
factor EF-1-alpha"
intron 232..1170
/label=EF-1-alpha intron A
/note="intron upstream of the start codon of human
EF-1-alpha"
primer_bind 1127..1147
/label=EF1a-F
/note="Human elongation factor-1a promoter, forward primer"
regulatory 1187..1196
/regulatory_class="other"
/note="vertebrate consensus sequence for strong initiation
of translation (Kozak, 1987)"
CDS 1193..1900
/codon_start=1
/product="photoactivatable monomeric derivative of DsRed
fluorescent protein (Subach et al., 2009)"
/label=PAmCherry1
/note="mammalian codon-optimized"
/translation="MVSKGEEDNMAIIKEFMRFKVHMEGSVNGHVFEIEGEGEGRPYEG
TQTAKLKVTKGGPLPFTWDILSPQFMYGSNAYVKHPADIPDYFKLSFPEGFKWERVMKF
EDGGVVTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEALSERMYPEDGALK
GEVKPRVKLKDGGHYDAEVKTTYKAKKPVQLPGAYNVNRKLDITSHNEDYTIVEQYERA
EGRHSTGGMDELYK"
primer_bind complement(1338..1356)
/label=mCherry-R
/note="mCherry, reverse primer"
misc_feature 2186..2648
/label=IRES
/note="internal ribosome entry site (IRES) of the
encephalomyocarditis virus (EMCV)"
primer_bind complement(2242..2259)
/label=IRES reverse
/note="IRES internal ribosome entry site, reverse primer.
Also called pCDH-rev"
primer_bind 2469..2488
/label=IRES-F
/note="IRES internal ribosome entry site, forward primer"
CDS 2681..3280
/codon_start=1
/gene="pac from Streptomyces alboniger"
/product="puromycin N-acetyltransferase"
/label=PuroR
/note="confers resistance to puromycin"
/translation="MTEYKPTVRLATRDDVPRAVRTLAAAFADYPATRHTVDPDRHIER
VTELQELFLTRVGLDIGKVWVADDGAAVAVWTTPESVEAGAVFAEIGPRMAELSGSRLA
AQQQMEGLLAPHRPKEPAWFLATVGVSPDHQGKGLGSAVVLPGVEAAERAGVPAFLETS
APRNLPFYERLGFTVTADVEVPEGPRTWCMTRKPGA"
primer_bind complement(2681..2700)
/label=Puro-R
/note="Puromycin resistance gene, reverse primer. Also
called puro-variant-R"
primer_bind 3177..3197
/label=Puro-F
/note="Puromycin resistance gene, forward primer"
LTR 3483..3716
/label=3' LTR (Delta-U3)
/note="self-inactivating 3' long terminal repeat (LTR) from
HIV-1"
primer_bind complement(3867..3884)
/label=BGH-rev
/note="Bovine growth hormone terminator, reverse primer.
Also called BGH reverse"
polyA_signal 3873..4097
/label=bGH poly(A) signal
/note="bovine growth hormone polyadenylation signal"
regulatory 4140..4149
/regulatory_class="other"
/note="vertebrate consensus sequence for strong initiation
of translation (Kozak, 1987)"
promoter 4193..4297
/gene="bla"
/label=AmpR promoter
CDS 4298..5158
/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(4516..4535)
/label=Amp-R
/note="Ampicillin resistance gene, reverse primer"
rep_origin 5329..5917
/direction=RIGHT
/label=ori
/note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
replication"
primer_bind 5818..5837
/label=pBR322ori-F
/note="pBR322 origin, forward primer"
LTR 6076..6709
/label=3' LTR
/note="3' long terminal repeat (LTR) from HIV-1"
misc_feature 6756..6881
/label=HIV-1 Psi
/note="packaging signal of human immunodeficiency virus
type 1"
misc_feature 7374..7607
/label=RRE
/note="The Rev response element (RRE) of HIV-1 allows for
Rev-dependent mRNA export from the nucleus to the
cytoplasm."
CDS 7792..7836
/codon_start=1
/product="antigenic peptide corresponding to amino acids
655 to 669 of the HIV envelope protein gp41 (Lutje Hulsik
et al., 2013)"
/label=gp41 peptide
/note="recognized by the 2H10 single-chain llama nanobody"
/translation="KNEQELLELDKWASL"
misc_feature 8123..8240
/label=cPPT/CTS
/note="central polypurine tract and central termination
sequence of HIV-1"
ORIGIN
1 ggctccggtg cccgtcagtg ggcagagcgc acatcgccca cagtccccga gaagttgggg
61 ggaggggtcg gcaattgaac cggtgcctag agaaggtggc gcggggtaaa ctgggaaagt
121 gatgtcgtgt actggctccg cctttttccc gagggtgggg gagaaccgta tataagtgca
181 gtagtcgccg tgaacgttct ttttcgcaac gggtttgccg ccagaacaca ggtaagtgcc
241 gtgtgtggtt cccgcgggcc tggcctcttt acgggttatg gcccttgcgt gccttgaatt
301 acttccacct ggctgcagta cgtgattctt gatcccgagc ttcgggttgg aagtgggtgg
361 gagagttcga ggccttgcgc ttaaggagcc ccttcgcctc gtgcttgagt tgaggcctgg
421 cctgggcgct ggggccgccg cgtgcgaatc tggtggcacc ttcgcgcctg tctcgctgct
481 ttcgataagt ctctagccat ttaaaatttt tgatgacctg ctgcgacgct ttttttctgg
541 caagatagtc ttgtaaatgc gggccaagat ctgcacactg gtatttcggt ttttggggcc
601 gcgggcggcg acggggcccg tgcgtcccag cgcacatgtt cggcgaggcg gggcctgcga
661 gcgcggccac cgagaatcgg acgggggtag tctcaagctg gccggcctgc tctggtgcct
721 ggcctcgcgc cgccgtgtat cgccccgccc tgggcggcaa ggctggcccg gtcggcacca
781 gttgcgtgag cggaaagatg gccgcttccc ggccctgctg cagggagctc aaaatggagg
841 acgcggcgct cgggagagcg ggcgggtgag tcacccacac aaaggaaaag ggcctttccg
901 tcctcagccg tcgcttcatg tgactccacg gagtaccggg cgccgtccag gcacctcgat
961 tagttctcga gcttttggag tacgtcgtct ttaggttggg gggaggggtt ttatgcgatg
1021 gagtttcccc acactgagtg ggtggagact gaagttaggc cagcttggca cttgatgtaa
1081 ttctccttgg aatttgccct ttttgagttt ggatcttggt tcattctcaa gcctcagaca
1141 gtggttcaaa gtttttttct tccatttcag gtgtcgtgag gaattcgcca ccatggtgag
1201 caagggcgag gaggataaca tggccatcat taaggagttc atgcgcttca aggtgcacat
1261 ggaggggtcc gtgaacggcc acgtgttcga gatcgagggc gagggcgagg gccgccccta
1321 cgagggcacc cagaccgcca agctgaaggt gaccaagggc ggccccctgc ccttcacctg
1381 ggacatcctg agccctcagt tcatgtacgg ctccaatgcc tacgtgaagc accccgccga
1441 catccccgac tactttaagc tgtccttccc cgagggcttc aagtgggagc gcgtgatgaa
1501 attcgaggac ggcggcgtgg tgaccgtgac ccaggactcc tccctgcagg acggcgagtt
1561 catctacaag gtgaagctgc gcggcaccaa cttcccctcc gacggccccg tgatgcagaa
1621 gaagaccatg ggctgggagg ccctctccga gcggatgtac cccgaggacg gcgccctgaa
1681 gggcgaggtc aagcccagag tgaagctgaa ggacggcggc cactacgacg ctgaggtcaa
1741 gaccacctac aaggccaaga agcccgtgca gctgcccggc gcctacaacg tcaaccgcaa
1801 gctggacatc accagccaca acgaggacta caccatcgtg gagcagtacg agagagccga
1861 gggccgccac tccaccggcg gcatggacga gctgtacaag gctagcaagg aatcccgggc
1921 caagaaattc cagcggcagc atatggactc agacagttcc cccagcacca aagaaaaccc
1981 tcgtagcaac caggaagaaa gctatgatga taacgaaagc taagtttaaa caatcactag
2041 ttcgaaggat ccgcatgcat ctagggcggc caattccgcc cctctccccc ccccctaacg
2101 ttactggccg aagccgcttg gaataaggcc ggtgtgcgtt tgtctatatg ttattttcca
2161 ccatattgcc gtcttttggc aatgtgaggg cccggaaacc tggccctgtc ttcttgacga
2221 gcattcctag gggtctttcc cctctcgcca aaggaatgca aggtctgttg aatgtcgtga
2281 aggaagcagt tcctctggaa gcttcttgaa gacaaacaac gtctgtagcg accctttgca
2341 ggcagcggaa ccccccacct ggcgacaggt gcctctgcgg ccaaaagcca cgtgtataag
2401 atacacctgc aaaggcggca caaccccagt gccacgttgt gagttggata gttgtggaaa
2461 gagtcaaatg gctctcctca agcgtattca acaaggggct gaaggatgcc cagaaggtac
2521 cccattgtat gggatctgat ctggggcctc ggtgcacatg ctttacatgt gtttagtcga
2581 ggttaaaaaa acgtctaggc cccccgaacc acggggacgt ggttttcctt tgaaaaacac
2641 gatgataagc ttgccacaac ccacaaggag acgaccttcc atgaccgagt acaagcccac
2701 ggtgcgcctc gccacccgcg acgacgtccc ccgggccgta cgcaccctcg ccgccgcgtt
2761 cgccgactac cccgccacgc gccacaccgt cgacccggac cgccacatcg agcgggtcac
2821 cgagctgcaa gaactcttcc tcacgcgcgt cgggctcgac atcggcaagg tgtgggtcgc
2881 ggacgacggc gccgcggtgg cggtctggac cacgccggag agcgtcgaag cgggggcggt
2941 gttcgccgag atcggcccgc gcatggccga gttgagcggt tcccggctgg ccgcgcagca
3001 acagatggaa ggcctcctgg cgccgcaccg gcccaaggag cccgcgtggt tcctggccac
3061 cgtcggcgtc tcgcccgacc accagggcaa gggtctgggc agcgccgtcg tgctccccgg
3121 agtggaggcg gccgagcgcg ccggggtgcc cgccttcctg gagacctccg cgccccgcaa
3181 cctccccttc tacgagcggc tcggcttcac cgtcaccgcc gacgtcgagg tgcccgaagg
3241 accgcgcacc tggtgcatga cccgcaagcc cggtgcctga taataggcgg ccgctcgaga
3301 cctagaaaaa catggagcaa tcacaagtag caatacagca gctaccaatg ctgattgtgc
3361 ctggctagaa gcacaagagg aggaggaggt gggttttcca gtcacacctc aggtaccttt
3421 aagaccaatg acttacaagg cagctgtaga tcttagccac tttttaaaag aaaagggggg
3481 actggaaggg ctaattcact cccaacgaag acaagatctg ctttttgctt gtactgggtc
3541 tctctggtta gaccagatct gagcctggga gctctctggc taactaggga acccactgct
3601 taagcctcaa taaagcttgc cttgagtgct tcaagtagtg tgtgcccgtc tgttgtgtga
3661 ctctggtaac tagagatccc tcagaccctt ttagtcagtg tggaaaatct ctagcagtag
3721 tagttcatgt catcttatta ttcagtattt ataacttgca aagaaatgaa tatcagagag
3781 tgagaggcct tgacattata atagatttag caggaattga actaggagtg gagcacacag
3841 gcaaagttct agagctcgct gatcagcctc gactgtgcct tctagttgcc agccatctgt
3901 tgtttgcccc tcccccgtgc cttccttgac cctggaaggt gccactccca ctgtcctttc
3961 ctaataaaat gaggaaattg catcgcattg tctgagtagg tgtcattcta ttctgggggg
4021 tggggtgggg caggacagca agggggagga ttgggaagac aatagcaggc atgctgggga
4081 tgcggtgggc tctatggctt ctgaggcgga aagaaccagc tgggggcgcg cccctcgagg
4141 ccgccatggt catagctgtt tgacgtcagg tggcactttt cggggaaatg tgcgcggaac
4201 ccctatttgt ttatttttct aaatacattc aaatatgtat ccgctcacga gacaataacc
4261 ctgataaatg cttcaataat attgaaaaag gaagagtatg agtattcaac atttccgtgt
4321 cgcccttatt cccttttttg cggcattttg ccttcctgtt tttgctcacc cagaaacgct
4381 ggtgaaagta aaagatgctg aagatcagtt gggtgcacga gtgggttaca tcgaactgga
4441 tctcaacagc ggtaagatcc ttgagagttt tcgccccgaa gaacgttttc caatgatgag
4501 cacttttaaa gttctgctat gtggcgcggt attatcccgt attgacgccg ggcaagagca
4561 actcggtcgc cgcatacact attctcagaa tgacttggtt gagtactcac cagtcacaga
4621 aaagcatctt acggatggca tgacagtaag agaattatgc agtgctgcca taaccatgag
4681 tgataacact gcggccaact tacttctgac aacgatcgga ggaccgaagg agctaaccgc
4741 ttttttgcac aacatggggg atcatgtaac tcgccttgat cgttgggaac cggagctgaa
4801 tgaagccata ccaaacgacg agcgtgacac cacgatgcct gtagcaatgg caacaacgtt
4861 gcgcaaacta ttaactggcg aactacttac tctagcttcc cggcaacaat taatagactg
4921 gatggaggcg gataaagttg caggaccact tctgcgctcg gcccttccgg ctggctggtt
4981 tattgctgat aaatctggag ccggtgagcg tgggtctcgc ggtatcattg cagcactggg
5041 gccagatggt aagccctccc gtatcgtagt tatctacacg acggggagtc aggcaactat
5101 ggatgaacga aatagacaga tcgctgagat aggtgcctca ctgattaagc attggtaact
5161 gtcagaccaa gtttactcat atatacttta gattgattta aaacttcatt tttaatttaa
5221 aaggatctag gtgaagatcc tttttgataa tctcatgacc aaaatccctt aacgtgagtt
5281 ttcgttccac tgagcgtcag accccgtaga aaagatcaaa ggatcttctt gagatccttt
5341 ttttctgcgc gtaatctgct gcttgcaaac aaaaaaacca ccgctaccag cggtggtttg
5401 tttgccggat caagagctac caactctttt tccgaaggta actggcttca gcagagcgca
5461 gataccaaat actgttcttc tagtgtagcc gtagttaggc caccacttca agaactctgt
5521 agcaccgcct acatacctcg ctctgctaat cctgttacca gtggctgctg ccagtggcga
5581 taagtcgtgt cttaccgggt tggactcaag acgatagtta ccggataagg cgcagcggtc
5641 gggctgaacg gggggttcgt gcacacagcc cagcttggag cgaacgacct acaccgaact
5701 gagataccta cagcgtgagc tatgagaaag cgccacgctt cccgaaggga gaaaggcgga
5761 caggtatccg gtaagcggca gggtcggaac aggagagcgc acgagggagc ttccaggggg
5821 aaacgcctgg tatctttata gtcctgtcgg gtttcgccac ctctgacttg agcgtcgatt
5881 tttgtgatgc tcgtcagggg ggcggagcct atggaaaaac gccagcaacg cggccttttt
5941 acggttctgg ccttttgcac ggaccgccac tgccaattac ctgtggtttc atttactcta
6001 aacctgtgat tcctctgaat tattttcatt ttaaagaaat tgtatttgtt aaatatgtac
6061 tacaaactta gtagttggaa gggctaattc actcccaaag aagacaagat atccttgatc
6121 tgtggatcta ccacacacaa ggctacttcc ctgattagca gaactacaca ccagggccag
6181 gggtcagata tccactgacc tttggatggt gctacaagct agtaccagtt gagccagata
6241 aggtagaaga ggccaataaa ggagagaaca ccagcttgtt acaccctgtg agcctgcatg
6301 ggatggatga cccggagaga gaagtgttag agtggaggtt tgacagccgc ctagcatttc
6361 atcacgtggc ccgagagctg catccggagt acttcaagaa ctgctgatat cgagcttgct
6421 acaagggact ttccgctggg gactttccag ggaggcgtgg cctgggcggg actggggagt
6481 ggcgagccct cagatcctgc atataagcag ctgctttttg cctgtactgg gtctctctgg
6541 ttagaccaga tctgagcctg ggagctctct ggctaactag ggaacccact gcttaagcct
6601 caataaagct tgccttgagt gcttcaagta gtgtgtgccc gtctgttgtg tgactctggt
6661 aactagagat ccctcagacc cttttagtca gtgtggaaaa tctctagcag tggcgcccga
6721 acagggactt gaaagcgaaa gggaaaccag aggagctctc tcgacgcagg actcggcttg
6781 ctgaagcgcg cacggcaaga ggcgaggggc ggcgactggt gagtacgcca aaaattttga
6841 ctagcggagg ctagaaggag agagatgggt gcgagagcgt cagtattaag cgggggagaa
6901 ttagatcgcg atgggaaaaa attcggttaa ggccaggggg aaagaaaaaa tataaattaa
6961 aacatatagt atgggcaagc agggagctag aacgattcgc agttaatcct ggcctgttag
7021 aaacatcaga aggctgtaga caaatactgg gacagctaca accatccctt cagacaggat
7081 cagaagaact tagatcatta tataatacag tagcaaccct ctattgtgtg catcaaagga
7141 tagagataaa agacaccaag gaagctttag acaagataga ggaagagcaa aacaaaagta
7201 agaccaccgc acagcaagcg gccgctgatc ttcagacctg gaggaggaga tatgagggac
7261 aattggagaa gtgaattata taaatataaa gtagtaaaaa ttgaaccatt aggagtagca
7321 cccaccaagg caaagagaag agtggtgcag agagaaaaaa gagcagtggg aataggagct
7381 ttgttccttg ggttcttggg agcagcagga agcactatgg gcgcagcgtc aatgacgctg
7441 acggtacagg ccagacaatt attgtctggt atagtgcagc agcagaacaa tttgctgagg
7501 gctattgagg cgcaacagca tctgttgcaa ctcacagtct ggggcatcaa gcagctccag
7561 gcaagaatcc tggctgtgga aagataccta aaggatcaac agctcctggg gatttggggt
7621 tgctctggaa aactcatttg caccactgct gtgccttgga atgctagttg gagtaataaa
7681 tctctggaac agatttggaa tcacacgacc tggatggagt gggacagaga aattaacaat
7741 tacacaagct taatacactc cttaattgaa gaatcgcaaa accagcaaga aaagaatgaa
7801 caagaattat tggaattaga taaatgggca agtttgtgga attggtttaa cataacaaat
7861 tggctgtggt atataaaatt attcataatg atagtaggag gcttggtagg tttaagaata
7921 gtttttgctg tactttctat agtgaataga gttaggcagg gatattcacc attatcgttt
7981 cagacccacc tcccaacccc gaggggaccc gacaggcccg aaggaataga agaagaaggt
8041 ggagagagag acagagacag atccattcga ttagtgaacg gatctcgacg gtatcgccac
8101 aaatggcagt attcatccac aattttaaaa gaaagggggg gattgggggg tacagtgcag
8161 gggaaagaat agtagacata atagcaacag acatacaaac taaagaatta caaaaacaaa
8221 ttacaaaaat tcaaaatttt cgggtttatt acagggacag cagagatcca ctttggatcg
8281 ataagctttg caaagatgga taaagtttta aacagagagg aatctttgca gctaatggac
8341 cttctaggtc ttgaaaggag tgggaatt
//
