Catalog No. PVT10876
Packing 2ug
Function Mammal Editing plasmids
Plasmid classification: lactation serial plasmids, lactation editing plasmids, suckling transposable plasmid.
Screening markers: green fluorescent protein CopGFP and puromycin Puro (10ug/ml).
PiggyBac Dual promoter (PB513B-1) is a plasmid for a mammalian cell transposing experiment, and a polyclonal site (MCS) located downstream of the CMV promoter, making the target gene or microRNA more convenient to clone. The downstream EF-1 alpha core promoter starts the expression of GFP.
The PiggyBac (PB) transposon is a mobile genetic element that efficiently transposes between vectors and chromosomes via a "cut and paste" mechanism. During transposition, the PB transposase recognizes transposon-specific inverted terminal repeat sequences (ITRs) located on both ends of the transposon vector and efficiently moves the contents from the original sites and efficiently integrates them into TTAA chromosomal sites. The powerful activity of the piggyBac transposon system enables genes of interest between the two ITRs in the PB vector to be easily mobilized into target genomes.
The unique features of piggyBac transposons are that there is NO Cargo Limit and it is also Reversible. Genomes containing an inserted piggyBac vector can be transiently re-transfected with the PB transposase expression vector. The PB transposase will remove the transposons from the genome, footprint-free.The Super PiggyBac transposase transient expression vector and PB513B-1 were co-transfected into HeLa cells and puromycin selection applied for 10 days (10ug/ml). Cells efficiently transposed were Puro resistant and GFP positive.
LOCUS Exported File 7258 bp ds-DNA circular SYN 24-5-2015
KEYWORDS PiggyBac Dual promoter(PB513B-1)
SOURCE synthetic DNA construct
ORGANISM synthetic DNA construct
REFERENCE 1 (bases 1 to 7258)
TITLE Direct Submission
JOURNAL Exported 2015-5-24
FEATURES Location/Qualifiers
source 1..7258
/organism="synthetic DNA construct"
/mol_type="other DNA"
promoter complement(1..105)
/gene="bla"
/note="AmpR promoter"
rep_origin complement(131..586)
/direction=LEFT
/note="f1 ori"
/note="f1 bacteriophage origin of replication; arrow
indicates direction of (+) strand synthesis"
primer_bind 728..744
/note="M13 fwd"
/note="common sequencing primer, one of multiple similar
variants"
promoter 1442..1645
/note="CMV promoter"
/note="human cytomegalovirus (CMV) immediate early
promoter"
CDS 3068..3121
/codon_start=1
/product="2A peptide from?Thosea asigna virus capsid
protein"
/note="T2A"
/note="Eukaryotic ribosomes fail to insert a peptide bond
between the Gly and Pro residues, yielding separate
polypeptides."
/protein_id="
"
/translation="EGRGSLLTCGDVEENPGP"
CDS 3122..3721
/codon_start=1
/gene="pac from Streptomyces alboniger"
/product="puromycin N-acetyltransferase"
/note="PuroR"
/note="confers resistance to puromycin"
/translation="MTEYKPTVRLATRDDVPRAVRTLAAAFADYPATRHTVDPDRHIER
VTELQELFLTRVGLDIGKVWVADDGAAVAVWTTPESVEAGAVFAEIGPRMAELSGSRLA
AQQQMEGLLAPHRPKEPAWFLATVGVSPDHQGKGLGSAVVLPGVEAAERAGVPAFLETS
APRNLPFYERLGFTVTADVEVPEGPRTWCMTRKPGA"
polyA_signal 3829..3950
/note="SV40 poly(A) signal"
/note="SV40 polyadenylation signal"
primer_bind complement(5237..5253)
/note="M13 rev"
/note="common sequencing primer, one of multiple similar
variants"
protein_bind 5261..5277
/bound_moiety="lac repressor encoded by lacI"
/note="lac operator"
/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(5285..5315)
/note="lac promoter"
/note="promoter for the E. coli lac operon"
protein_bind 5330..5351
/bound_moiety="E. coli catabolite activator protein"
/note="CAP binding site"
/note="CAP binding activates transcription in the presence
of cAMP."
rep_origin complement(5639..6227)
/direction=LEFT
/note="ori"
/note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
replication"
CDS complement(6398..7258)
/codon_start=1
/gene="bla"
/product="beta-lactamase"
/note="AmpR"
/note="confers resistance to ampicillin, carbenicillin, and
related antibiotics"
/translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
LIKHW"
ORIGIN
1 actcttcctt tttcaatatt attgaagcat ttatcagggt tattgtctca tgagcggata
61 catatttgaa tgtatttaga aaaataaaca aataggggtt ccgcgcacat ttccccgaaa
121 agtgccacct aaattgtaag cgttaatatt ttgttaaaat tcgcgttaaa tttttgttaa
181 atcagctcat tttttaacca ataggccgaa atcggcaaaa tcccttataa atcaaaagaa
241 tagaccgaga tagggttgag tgttgttcca gtttggaaca agagtccact attaaagaac
301 gtggactcca acgtcaaagg gcgaaaaacc gtctatcagg gcgatggccc actacgtgaa
361 ccatcaccct aatcaagttt tttggggtcg aggtgccgta aagcactaaa tcggaaccct
421 aaagggagcc cccgatttag agcttgacgg ggaaagccgg cgaacgtggc gagaaaggaa
481 gggaagaaag cgaaaggagc gggcgctagg gcgctggcaa gtgtagcggt cacgctgcgc
541 gtaaccacca cacccgccgc gcttaatgcg ccgctacagg gcgcgtccca ttcgccattc
601 aggctgcgca actgttggga agggcgatcg gtgcgggcct cttcgctatt acgccagctg
661 gcgaaagggg gatgtgctgc aaggcgatta agttgggtaa cgccagggtt ttcccagtca
721 cgacgttgta aaacgacggc cagtgagcgc gcctcgttca ttcacgtttt tgaacccgtg
781 gaggacgggc agactcgcgg tgcaaatgtg ttttacagcg tgatggagca gatgaagatg
841 ctcgacacgc tgcagaacac gcagctagat taaccctaga aagataatca tattgtgacg
901 tacgttaaag ataatcatgc gtaaaattga cgcatgtgtt ttatcggtct gtatatcgag
961 gtttatttat taatttgaat agatattaag ttttattata tttacactta catactaata
1021 ataaattcaa caaacaattt atttatgttt atttatttat taaaaaaaaa caaaaactca
1081 aaatttcttc tataaagtaa caaaactttt atgagggaca gccccccccc aaagccccca
1141 gggatgtaat tacgtccctc ccccgctagg gggcagcagc gagccgcccg gggctccgct
1201 ccggtccggc gctccccccg catccccgag ccggcagcgt gcggggacag cccgggcacg
1261 gggaaggtgg cacgggatcg ctttcctctg aacgcttctc gctgctcttt gagcctgcag
1321 acacctgggg ggatacgggg aaaaggcctc caaggcctac tagtattatg cccagtacat
1381 gaccttatgg gactttccta cttggcagta catctacgta ttagtcatcg ctattaccat
1441 ggtgatgcgg ttttggcagt acatcaatgg gcgtggatag cggtttgact cacggggatt
1501 tccaagtctc caccccattg acgtcaatgg gagtttgttt tggcaccaaa atcaacggga
1561 ctttccaaaa tgtcgtaaca actccgcccc attgacgcaa atgggcggta ggcgtgtacg
1621 gtgggaggtc tatataagca gagctcgttt agtgaaccgt cagatcgcct ggagacgcca
1681 tccacgctgt tttgacctcc atagaagatt ctagagctag cgaattcgaa tttaaatcgg
1741 atccgcggcc gcaaggatct gcgatcgctc cggtgcccgt cagtgggcag agcgcacatc
1801 gcccacagtc cccgagaagt tggggggagg ggtcggcaat tgaacgggtg cctagagaag
1861 gtggcgcggg gtaaactggg aaagtgatgt cgtgtactgg ctccgccttt ttcccgaggg
1921 tgggggagaa ccgtatataa gtgcagtagt cgccgtgaac gttctttttc gcaacgggtt
1981 tgccgccaga acacagctga agcttcgagg ggctcgcatc tctccttcac gcgcccgccg
2041 ccctacctga ggccgccatc cacgccggtt gagtcgcgtt ctgccgcctc ccgcctgtgg
2101 tgcctcctga actgcgtccg ccgtctaggt aagtttaaag ctcaggtcga gaccgggcct
2161 ttgtccggcg ctcccttgga gcctacctag actcagccgg ctctccacgc tttgcctgac
2221 cctgcttgct caactctacg tctttgtttc gttttctgtt ctgcgccgtt acagatccaa
2281 gctgtgaccg gcgcctacgc tagacgccac catggagagc gacgagagcg gcctgcccgc
2341 catggagatc gagtgccgca tcaccggcac cctgaacggc gtggagttcg agctggtggg
2401 cggcggagag ggcaccccca agcagggccg catgaccaac aagatgaaga gcaccaaagg
2461 cgccctgacc ttcagcccct acctgctgag ccacgtgatg ggctacggct tctaccactt
2521 cggcacctac cccagcggct acgagaaccc cttcctgcac gccatcaaca acggcggcta
2581 caccaacacc cgcatcgaga agtacgagga cggcggcgtg ctgcacgtga gcttcagcta
2641 ccgctacgag gccggccgcg tgatcggcga cttcaaggtg gtgggcaccg gcttccccga
2701 ggacagcgtg atcttcaccg acaagatcat ccgcagcaac gccaccgtgg agcacctgca
2761 ccccatgggc gataacgtgc tggtgggcag cttcgcccgc accttcagcc tgcgcgacgg
2821 cggctactac agcttcgtgg tggacagcca catgcacttc aagagcgcca tccaccccag
2881 catcctgcag aacgggggcc ccatgttcgc cttccgccgc gtggaggagc tgcacagcaa
2941 caccgagctg ggcatcgtgg agtaccagca cgccttcaag acccccatcg ccttcgccag
3001 atcccgcgct cagtcgtcca attctgccgt ggacggcacc gccggacccg gctccaccgg
3061 atctcgcgag ggcagaggaa gtcttctaac atgcggtgac gtggaggaga atcccggccc
3121 tatgaccgag tacaagccca cggtgcgcct cgccacccgc gacgacgtcc ccagggccgt
3181 acgcaccctc gccgccgcgt tcgccgacta ccccgccacg cgccacaccg tcgatccgga
3241 ccgccacatc gagcgggtca ccgagctgca agaactcttc ctcacgcgcg tcgggctcga
3301 catcggcaag gtgtgggtcg cggacgacgg cgccgcggtg gcggtctgga ccacgccgga
3361 gagcgtcgaa gcgggggcgg tgttcgccga gatcggcccg cgcatggccg agttgagcgg
3421 ttcccggctg gccgcgcagc aacagatgga aggcctcctg gcgccgcacc ggcccaagga
3481 gcccgcgtgg ttcctggcca ccgtcggcgt ctcgcccgac caccagggca agggtctggg
3541 cagcgccgtc gtgctccccg gagtggaggc ggccgagcgc gccggggtgc ccgccttcct
3601 ggagacctcc gcgccccgca acctcccctt ctacgagcgg ctcggcttca ccgtcaccgc
3661 cgacgtcgag gtgcccgaag gaccgcgcac ctggtgcatg acccgcaagc ccggtgcctg
3721 aaatcaacct ctggattaca aaatttgtga aagattgact ggtattctta actatgttgc
3781 tccttttacg ctatgtggat acgctgcttt aatgcctttg tatcagttaa cttgtttatt
3841 gcagcttata atggttacaa ataaagcaat agcatcacaa atttcacaaa taaagcattt
3901 ttttcactgc attctagttg tggtttgtcc aaactcatca atgtatctta tcatgtctgg
3961 aattgactca aatgatgtca attagtctat cagaagctat ctggtctccc ttccggggga
4021 caagacatcc ctgtttaata tttaaacagc agtgttccca aactgggttc ttatatccct
4081 tgctctggtc aaccaggttg cagggtttcc tgtcctcaca ggaacgaagt ccctaaagaa
4141 acagtggcag ccaggtttag ccccggaatt gactggattc cttttttagg gcccattggt
4201 atggcttttt ccccgtatcc ccccaggtgt ctgcaggctc aaagagcagc gagaagcgtt
4261 cagaggaaag cgatcccgtg ccaccttccc cgtgcccggg ctgtccccgc acgctgccgg
4321 ctcggggatg cggggggagc gccggaccgg agcggagccc cgggcggctc gctgctgccc
4381 cctagcgggg gagggacgta attacatccc tgggggcttt gggggggggc tgtccctgat
4441 atctataaca agaaaatata tatataataa gttatcacgt aagtagaaca tgaaataaca
4501 atataattat cgtatgagtt aaatcttaaa agtcacgtaa aagataatca tgcgtcattt
4561 tgactcacgc ggtcgttata gttcaaaatc agtgacactt accgcattga caagcacgcc
4621 tcacgggagc tccaagcggc gactgagatg tcctaaatgc acagcgacgg attcgcgcta
4681 tttagaaaga gagagcaata tttcaagaat gcatgcgtca attttacgca gactatcttt
4741 ctagggttaa tctagctgca tcaggatcat atcgtcgggt cttttttccg gctcagtcat
4801 cgcccaagct ggcgctatct gggcatcggg gaggaagaag cccgtgcctt ttcccgcgag
4861 gttgaagcgg catggaaaga gtttgccgag gatgactgct gctgcattga cgttgagcga
4921 aaacgcacgt ttaccatgat gattcgggaa ggtgtggcca tgcacgcctt taacggtgaa
4981 ctgttcgttc aggccacctg ggataccagt tcgtcgcggc ttttccggac acagttccgg
5041 atggtcagcc cgaagcgcat cagcaacccg aacaataccg gcgacagccg gaactgccgt
5101 gccggtgtgc agattaatga cagcggtgcg gcgctgggat attacgtcag cgaggacggg
5161 tatcctggct ggatgccgca gaaatggaca tggatacccc gtgagttacc cggcgggcgc
5221 gcttggcgta atcatggtca tagctgtttc ctgtgtgaaa ttgttatccg ctcacaattc
5281 cacacaacat acgagccgga agcataaagt gtaaagcctg gggtgcctaa tgagtgagct
5341 aactcacatt aattgcgttg cgctcactgc ccgctttcca gtcgggaaac ctgtcgtgcc
5401 agctgcatta atgaatcggc caacgcgcgg ggagaggcgg tttgcgtatt gggcgctctt
5461 ccgcttcctc gctcactgac tcgctgcgct cggtcgttcg gctgcggcga gcggtatcag
5521 ctcactcaaa ggcggtaata cggttatcca cagaatcagg ggataacgca ggaaagaaca
5581 tgtgagcaaa aggccagcaa aaggccagga accgtaaaaa ggccgcgttg ctggcgtttt
5641 tccataggct ccgcccccct gacgagcatc acaaaaatcg acgctcaagt cagaggtggc
5701 gaaacccgac aggactataa agataccagg cgtttccccc tggaagctcc ctcgtgcgct
5761 ctcctgttcc gaccctgccg cttaccggat acctgtccgc ctttctccct tcgggaagcg
5821 tggcgctttc tcatagctca cgctgtaggt atctcagttc ggtgtaggtc gttcgctcca
5881 agctgggctg tgtgcacgaa ccccccgttc agcccgaccg ctgcgcctta tccggtaact
5941 atcgtcttga gtccaacccg gtaagacacg acttatcgcc actggcagca gccactggta
6001 acaggattag cagagcgagg tatgtaggcg gtgctacaga gttcttgaag tggtggccta
6061 actacggcta cactagaagg acagtatttg gtatctgcgc tctgctgaag ccagttacct
6121 tcggaaaaag agttggtagc tcttgatccg gcaaacaaac caccgctggt agcggtggtt
6181 tttttgtttg caagcagcag attacgcgca gaaaaaaagg atctcaagaa gatcctttga
6241 tcttttctac ggggtctgac gctcagtgga acgaaaactc acgttaaggg attttggtca
6301 tgagattatc aaaaaggatc ttcacctaga tccttttaaa ttaaaaatga agttttaaat
6361 caatctaaag tatatatgag taaacttggt ctgacagtta ccaatgctta atcagtgagg
6421 cacctatctc agcgatctgt ctatttcgtt catccatagt tgcctgactc cccgtcgtgt
6481 agataactac gatacgggag ggcttaccat ctggccccag tgctgcaatg ataccgcgag
6541 acccacgctc accggctcca gatttatcag caataaacca gccagccgga agggccgagc
6601 gcagaagtgg tcctgcaact ttatccgcct ccatccagtc tattaattgt tgccgggaag
6661 ctagagtaag tagttcgcca gttaatagtt tgcgcaacgt tgttgccatt gctacaggca
6721 tcgtggtgtc acgctcgtcg tttggtatgg cttcattcag ctccggttcc caacgatcaa
6781 ggcgagttac atgatccccc atgttgtgca aaaaagcggt tagctccttc ggtcctccga
6841 tcgttgtcag aagtaagttg gccgcagtgt tatcactcat ggttatggca gcactgcata
6901 attctcttac tgtcatgcca tccgtaagat gcttttctgt gactggtgag tactcaacca
6961 agtcattctg agaatagtgt atgcggcgac cgagttgctc ttgcccggcg tcaatacggg
7021 ataataccgc gccacatagc agaactttaa aagtgctcat cattggaaaa cgttcttcgg
7081 ggcgaaaact ctcaaggatc ttaccgctgt tgagatccag ttcgatgtaa cccactcgtg
7141 cacccaactg atcttcagca tcttttactt tcaccagcgt ttctgggtga gcaaaaacag
7201 gaaggcaaaa tgccgcaaaa aagggaataa gggcgacacg gaaatgttga atactcat
//
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