| 质粒名称: | pLCA.66/2272 |
|---|---|
| 出品公司: | 爱迪基因 |
| 目录编号: | 22733 |
| 存储实验室: | Mark Magnuson |
|
载体骨架基本信息 |
|
| 载体名称: | pBluescript KS+ |
| 载体出品公司: | Stratagene |
| 载体抗性: | 氨苄青霉素 |
| 空载体大小: | 2969 bp |
| 完整质粒大小: | -- |
| 载体修饰: | -- |
| 载体类型: | Cre/Lox；BAC重组 |
| 筛选标记: | 新霉素（G418）,丙氧鸟苷 |
| 高拷贝/低拷贝: | 高拷贝 |
| 生长条件: | 37 ℃ |
|
插入基因信息 |
|
| 插入基因名称: | Lox66-pU(delta)TK-EM7Neo-Lox2272 |
| 物种来源: | -- |
| 插入大小: | 3697 bp |
| 基因突变: | 该载体是为利用BAC重组方法组装基因靶向结构而设计的。该载体包含一个由小鼠磷酸甘油激酶启动子（pUDTK）驱动的嘌呤霉素（delta）胸苷激酶融合基因和一个由细菌EM7启动子（EM7neo）驱动的新霉素抗性基因。这些可选择的标记在pBluescript KS+主干中的lox66和lox2272位点（Cre重组酶识别序列）的两侧，并带有一个修改的多克隆区 |
| GenBank编号 | -- |
| 启动子 | -- |
| 标签/融合蛋白: | -- |
| 克隆方法: | 限制性内切酶 |
| 5' 克隆位点: | Kpn1(未破坏) |
| 3' 克隆位点: | Kpn1(未破坏) |
| 5' 测序引物 | T3 |
| 3' 测序引物 | T7 |
|
文献引用方法 |
|
| 材料和方法部分: | pLCA.66/2272 was a gift from Mark Magnuson (Addgene plasmid # 22733 ; http://n2t.net/addgene:22733 ; RRID:Addgene_22733) |
| 参考文献部分: | Quantification of factors influencing fluorescent protein expression using RMCE to generate an allelic series in the ROSA26 locus in mice. Chen SX, Osipovich AB, Ustione A, Potter LA, Hipkens S, Gangula R, Yuan W, Piston DW, Magnuson MA. Dis Model Mech. 2011 Feb 14. ():. 10.1242/dmm.006569 PubMed 21324933 |
买家导航
pLCA.662272载体基本信息
pLCA.662272载体图谱质粒图谱和多克隆位点信息
pLCA.662272载体简介
pLCA.662272载体序列
LOCUS pLCA.66/2272 6678 bp ds-DNA circular SYN 06-AUG-2018
DEFINITION synthetic circular DNA
ACCESSION .
VERSION .
KEYWORDS pLCA.66/2272
SOURCE synthetic DNA construct
ORGANISM synthetic DNA construct
REFERENCE 1 (bases 1 to 6678)
AUTHORS Chen SX, Osipovich AB, Ustione A, Potter LA, Hipkens S, Gangula R,
Yuan W, Piston DW, Magnuson MA
TITLE Quantification of factors influencing fluorescent protein expression
using RMCE to generate an allelic series in the ROSA26 locus in
mice.
JOURNAL Dis Model Mech. 2011 Feb 14. ():.
PUBMED 21324933
REFERENCE 2 (bases 1 to 6678)
AUTHORS .
TITLE Direct Submission
JOURNAL Exported Aug 6, 2018 from SnapGene Server 1.1.58
http://www.snapgene.com
FEATURES Location/Qualifiers
source 1..6678
/organism="synthetic DNA construct"
/mol_type="other DNA"
promoter 11..510
/label=PGK promoter
/note="mouse phosphoglycerate kinase 1 promoter"
primer_bind complement(46..69)
/label=MSCV-rev
/note="Murine stem cell virus, reverse primer"
primer_bind 443..462
/label=mPGK-F
/note="Mouse PGK promoter, forward primer"
regulatory 524..533
/regulatory_class="other"
/note="vertebrate consensus sequence for strong initiation
of translation (Kozak, 1987)"
CDS 530..1129
/codon_start=1
/gene="pac from Streptomyces alboniger"
/product="puromycin N-acetyltransferase"
/label=PuroR
/note="confers resistance to puromycin"
/translation="MGTEYKPTVRLATRDDVPRAVRTLAAAFADYPATRHTVDPDRHIE
RVTELQELFLTRVGLDIGKVWVADDGAAVAVWTTPESVEAGAVFAEIGPRMAELSGSRL
AAQQQMEGLLAPHRPKEPAWFLATVGVSPDHQGKGLGSAVVLPGVEAAERAGVPAFLET
SAPRNLPFYERLGFTVTADVEVPEGPRTWCMTRKPGA"
primer_bind 1029..1049
/label=Puro-F
/note="Puromycin resistance gene, forward primer"
CDS 1136..2131
/codon_start=1
/product="truncated herpes simplex virus thymidine kinase
(Salomon et al., 1995)"
/label=-Delta-TK
/note="confers sensitivity to nucleoside analogs such as
acyclovir or ganciclovir"
/translation="MPTLLRVYIDGPHGMGKTTTTQLLVALGSRDDIVYVPEPMTYWQV
LGASETIANIYTTQHRLDQGEISAGDAAVVMTSAQITMGMPYAVTDAVLAPHIGGEAGS
SHAPPPALTLIFDRHPIAALLCYPAARYLMGSMTPQAVLAFVALIPPTLPGTNIVLGAL
PEDRHIDRLAKRQRPGERLDLAMLAAIRRVYGLLANTVRYLQGGGSWREDWGQLSGTAV
PPQGAEPQSNAGPRPHIGDTLFTLFRAPELLAPNGDLYNVFAWALDVLAKRLRPMHVFI
LDYDQSPAGCRDALLQLTSGMVQTHVTTPGSIPTICDLARTFAREMGEAN"
polyA_signal 2159..2383
/label=bGH poly(A) signal
/note="bovine growth hormone polyadenylation signal"
promoter 2429..2476
/label=EM7 promoter
/note="synthetic bacterial promoter "
CDS 2495..3298
/codon_start=1
/gene="aph(3')-II (or nptII)"
/product="aminoglycoside phosphotransferase from Tn5"
/label=NeoR/KanR
/note="confers resistance to neomycin, kanamycin, and G418
(Geneticin(R))"
/translation="MGSAIEQDGLHAGSPAACVERLFGYDWAQQTIGCSDAAVFRLSAQ
GRPVLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQD
LLSSHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDE
EHQGLAPAELFARLKARMPDGDDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQ
DIALATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
primer_bind complement(2558..2577)
/label=Neo-R
/note="Neomycin resistance gene, reverse primer"
primer_bind 3168..3187
/label=Neo-F
/note="Neomycin resistance gene, forward primer"
polyA_signal 3336..3560
/label=bGH poly(A) signal
/note="bovine growth hormone polyadenylation signal"
protein_bind complement(3626..3659)
/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."
primer_bind complement(3735..3754)
/label=T7
/note="T7 promoter, forward primer"
promoter complement(3736..3754)
/label=T7 promoter
/note="promoter for bacteriophage T7 RNA polymerase"
primer_bind complement(3761..3778)
/label=M13 Forward
/note="In lacZ gene. Also called M13-F20 or M13 (-21)
Forward"
primer_bind complement(3761..3777)
/label=M13 fwd
/note="common sequencing primer, one of multiple similar
variants"
primer_bind complement(3770..3792)
/label=M13/pUC Forward
/note="In lacZ gene"
rep_origin 3919..4374
/direction=RIGHT
/label=f1 ori
/note="f1 bacteriophage origin of replication; arrow
indicates direction of (+) strand synthesis"
primer_bind complement(4006..4025)
/label=F1ori-R
/note="F1 origin, reverse primer"
primer_bind 4216..4237
/label=F1ori-F
/note="F1 origin, forward primer"
promoter 4400..4504
/gene="bla"
/label=AmpR promoter
CDS 4505..5365
/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(4723..4742)
/label=Amp-R
/note="Ampicillin resistance gene, reverse primer"
rep_origin 5536..6124
/direction=RIGHT
/label=ori
/note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
replication"
primer_bind 6025..6044
/label=pBR322ori-F
/note="pBR322 origin, forward primer"
primer_bind 6278..6295
/label=L4440
/note="L4440 vector, forward primer"
protein_bind 6412..6433
/label=CAP binding site
/bound_moiety="E. coli catabolite activator protein"
/note="CAP binding activates transcription in the presence
of cAMP."
promoter 6448..6478
/label=lac promoter
/note="promoter for the E. coli lac operon"
protein_bind 6486..6502
/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 6491..6513
/label=M13/pUC Reverse
/note="In lacZ gene"
primer_bind 6510..6526
/label=M13 rev
/note="common sequencing primer, one of multiple similar
variants"
primer_bind 6510..6526
/label=M13 Reverse
/note="In lacZ gene. Also called M13-rev"
promoter 6547..6565
/label=T3 promoter
/note="promoter for bacteriophage T3 RNA polymerase"
protein_bind complement(6623..6656)
/label=lox66
/bound_moiety="Cre recombinase"
/note="loxP mutant. Cre-mediated recombination occurs in
the 8-bp core sequence (GCATACAT)."
ORIGIN
1 taattctacc gggtagggga ggcgcttttc ccaaggcagt ctggagcatg cgctttagca
61 gccccgctgg gcacttggcg ctacacaagt ggcctctggc ctcgcacaca ttccacatcc
121 accggtaggc gccaaccggc tccgttcttt ggtggcccct tcgcgccacc ttctactcct
181 cccctagtca ggaagttccc ccccgccccg cagctcgcgt cgtgcaggac gtgacaaatg
241 gaagtagcac gtctcactag tctcgtgcag atggacagca ccgctgagca atggaagcgg
301 gtaggccttt ggggcagcgg ccaatagcag ctttgctcct tcgctttctg ggctcagagg
361 ctgggaaggg gtgggtccgg gggcgggctc aggggcgggc tcaggggcgg ggcgggcgcc
421 cgaaggtcct ccggaggccc ggcattctgc acgcttcaaa agcgcacgtc tgccgcgctg
481 ttctcctctt cctcatctcc gggcctttcg acctgcagcc aacgccacca tggggaccga
541 gtacaagccc acggtgcgcc tcgccacccg cgacgacgtc ccccgggccg tacgcaccct
601 cgccgccgcg ttcgccgact accccgccac gcgccacacc gtcgacccgg accgccacat
661 cgagcgggtc accgagctgc aagaactctt cctcacgcgc gtcgggctcg acatcggcaa
721 ggtgtgggtc gcggacgacg gcgccgcggt ggcggtctgg accacgccgg agagcgtcga
781 agcgggggcg gtgttcgccg agatcggccc gcgcatggcc gagttgagcg gttcccggct
841 ggccgcgcag caacagatgg aaggcctcct ggcgccgcac cggcccaagg agcccgcgtg
901 gttcctggcc accgtcggcg tctcgcccga ccaccagggc aagggtctgg gcagcgccgt
961 cgtgctcccc ggagtggagg cggccgagcg cgccggggtg cccgccttcc tggagacctc
1021 cgcgccccgc aacctcccct tctacgagcg gctcggcttc accgtcaccg ccgacgtcga
1081 ggtgcccgaa ggaccgcgca cctggtgcat gacccgcaag cccggtgccg gatccatgcc
1141 cacgctactg cgggtttata tagacggtcc tcacgggatg gggaaaacca ccaccacgca
1201 actgctggtg gccctgggtt cgcgcgacga tatcgtctac gtacccgagc cgatgactta
1261 ctggcaggtg ctgggggctt ccgagacaat cgcgaacatc tacaccacac aacaccgcct
1321 cgaccagggt gagatatcgg ccggggacgc ggcggtggta atgacaagcg cccagataac
1381 aatgggcatg ccttatgccg tgaccgacgc cgttctggct cctcatatcg ggggggaggc
1441 tgggagctca catgccccgc ccccggccct caccctcatc ttcgaccgcc atcccatcgc
1501 cgccctcctg tgctacccgg ccgcgcgata ccttatgggc agcatgaccc cccaggccgt
1561 gctggcgttc gtggccctca tcccgccgac cttgcccggc acaaacatcg tgttgggggc
1621 ccttccggag gacagacaca tcgaccgcct ggccaaacgc cagcgccccg gcgagcggct
1681 tgacctggct atgctggccg cgattcgccg cgtttacggg ctgcttgcca atacggtgcg
1741 gtatctgcag ggcggcgggt cgtggcggga ggattgggga cagctttcgg ggacggccgt
1801 gccgccccag ggtgccgagc cccagagcaa cgcgggccca cgaccccata tcggggacac
1861 gttatttacc ctgtttcggg cccccgagtt gctggccccc aacggcgacc tgtataacgt
1921 gtttgcctgg gccttggacg tcttggccaa acgcctccgt cccatgcacg tctttatcct
1981 ggattacgac caatcgcccg ccggctgccg ggacgccctg ctgcaactta cctccgggat
2041 ggtccagacc cacgtcacca cccccggctc cataccgacg atctgcgacc tggcgcgcac
2101 gtttgcccgg gagatggggg aggctaactg agctctagag ctcgctgatc agcctcgact
2161 gtgccttcta gttgccagcc atctgttgtt tgcccctccc ccgtgccttc cttgaccctg
2221 gaaggtgcca ctcccactgt cctttcctaa taaaatgagg aaattgcatc gcattgtctg
2281 agtaggtgtc attctattct ggggggtggg gtggggcagg acagcaaggg ggaggattgg
2341 gaagacaata gcaggcatgc tggggatgcg gtgggctcta tggcttctga ggcggaaaga
2401 accatctggg gctgaattcc tgcagcctgt tgacaattaa tcatcggcat agtatatcgg
2461 catagtataa tacgacaagg tgaggaacta aaccatggga tcggccattg aacaagatgg
2521 attgcacgca ggttctccgg ccgcttgcgt ggagaggcta ttcggctatg actgggcaca
2581 acagacaatc ggctgctctg atgccgccgt gttccggctg tcagcgcagg ggcgcccggt
2641 tctttttgtc aagaccgacc tgtccggtgc cctgaatgaa ctgcaggacg aggcagcgcg
2701 gctatcgtgg ctggccacga cgggcgttcc ttgcgcagct gtgctcgacg ttgtcactga
2761 agcgggaagg gactggctgc tattgggcga agtgccgggg caggatctcc tgtcatctca
2821 ccttgctcct gccgagaaag tatccatcat ggctgatgca atgcggcggc tgcatacgct
2881 tgatccggct acctgcccat tcgaccacca agcgaaacat cgcatcgagc gagcacgtac
2941 tcggatggaa gccggtcttg tcgatcagga tgatctggac gaagagcatc aggggctcgc
3001 gccagccgaa ctgttcgcca ggctcaaggc gcgcatgccc gacggcgatg atctcgtcgt
3061 gacccatggc gatgcctgct tgccgaatat catggtggaa aatggccgct tttctggatt
3121 catcgactgt ggccggctgg gtgtggcgga ccgctatcag gacatagcgt tggctacccg
3181 tgatattgct gaagagcttg gcggcgaatg ggctgaccgc ttcctcgtgc tttacggtat
3241 cgccgctccc gattcgcagc gcatcgcctt ctatcgcctt cttgacgagt tcttctgagg
3301 ggatcaattc tctagagctc gctgatcagc ctcgactgtg ccttctagtt gccagccatc
3361 tgttgtttgc ccctcccccg tgccttcctt gaccctggaa ggtgccactc ccactgtcct
3421 ttcctaataa aatgaggaaa ttgcatcgca ttgtctgagt aggtgtcatt ctattctggg
3481 gggtggggtg gggcaggaca gcaaggggga ggattgggaa gacaatagca ggcatgctgg
3541 ggatgcggtg ggctctatgg cttctgaggc ggaaagaacc agctggggct cgactagagc
3601 ttgcggaacc cttgaattcc tgcagataac ttcgtataaa gtatcctata ggaagttatg
3661 gatccaggta ccttcgaaac gcgtcctagg catatgatct agagcggccg ccaccgcggt
3721 ggagctccaa ttcgccctat agtgagtcgt attacaattc actggccgtc gttttacaac
3781 gtcgtgactg ggaaaaccct ggcgttaccc aacttaatcg ccttgcagca catccccctt
3841 tcgccagctg gcgtaatagc gaagaggccc gcaccgatcg cccttcccaa cagttgcgca
3901 gcctgaatgg cgaatgggac gcgccctgta gcggcgcatt aagcgcggcg ggtgtggtgg
3961 ttacgcgcag cgtgaccgct acacttgcca gcgccctagc gcccgctcct ttcgctttct
4021 tcccttcctt tctcgccacg ttcgccggct ttccccgtca agctctaaat cgggggctcc
4081 ctttagggtt ccgatttagt gctttacggc acctcgaccc caaaaaactt gattagggtg
4141 atggttcacg tagtgggcca tcgccctgat agacggtttt tcgccctttg acgttggagt
4201 ccacgttctt taatagtgga ctcttgttcc aaactggaac aacactcaac cctatctcgg
4261 tctattcttt tgatttataa gggattttgc cgatttcggc ctattggtta aaaaatgagc
4321 tgatttaaca aaaatttaac gcgaatttta acaaaatatt aacgcttaca atttaggtgg
4381 cacttttcgg ggaaatgtgc gcggaacccc tatttgttta tttttctaaa tacattcaaa
4441 tatgtatccg ctcatgagac aataaccctg ataaatgctt caataatatt gaaaaaggaa
4501 gagtatgagt attcaacatt tccgtgtcgc ccttattccc ttttttgcgg cattttgcct
4561 tcctgttttt gctcacccag aaacgctggt gaaagtaaaa gatgctgaag atcagttggg
4621 tgcacgagtg ggttacatcg aactggatct caacagcggt aagatccttg agagttttcg
4681 ccccgaagaa cgttttccaa tgatgagcac ttttaaagtt ctgctatgtg gcgcggtatt
4741 atcccgtatt gacgccgggc aagagcaact cggtcgccgc atacactatt ctcagaatga
4801 cttggttgag tactcaccag tcacagaaaa gcatcttacg gatggcatga cagtaagaga
4861 attatgcagt gctgccataa ccatgagtga taacactgcg gccaacttac ttctgacaac
4921 gatcggagga ccgaaggagc taaccgcttt tttgcacaac atgggggatc atgtaactcg
4981 ccttgatcgt tgggaaccgg agctgaatga agccatacca aacgacgagc gtgacaccac
5041 gatgcctgta gcaatggcaa caacgttgcg caaactatta actggcgaac tacttactct
5101 agcttcccgg caacaattaa tagactggat ggaggcggat aaagttgcag gaccacttct
5161 gcgctcggcc cttccggctg gctggtttat tgctgataaa tctggagccg gtgagcgtgg
5221 gtctcgcggt atcattgcag cactggggcc agatggtaag ccctcccgta tcgtagttat
5281 ctacacgacg gggagtcagg caactatgga tgaacgaaat agacagatcg ctgagatagg
5341 tgcctcactg attaagcatt ggtaactgtc agaccaagtt tactcatata tactttagat
5401 tgatttaaaa cttcattttt aatttaaaag gatctaggtg aagatccttt ttgataatct
5461 catgaccaaa atcccttaac gtgagttttc gttccactga gcgtcagacc ccgtagaaaa
5521 gatcaaagga tcttcttgag atcctttttt tctgcgcgta atctgctgct tgcaaacaaa
5581 aaaaccaccg ctaccagcgg tggtttgttt gccggatcaa gagctaccaa ctctttttcc
5641 gaaggtaact ggcttcagca gagcgcagat accaaatact gtccttctag tgtagccgta
5701 gttaggccac cacttcaaga actctgtagc accgcctaca tacctcgctc tgctaatcct
5761 gttaccagtg gctgctgcca gtggcgataa gtcgtgtctt accgggttgg actcaagacg
5821 atagttaccg gataaggcgc agcggtcggg ctgaacgggg ggttcgtgca cacagcccag
5881 cttggagcga acgacctaca ccgaactgag atacctacag cgtgagctat gagaaagcgc
5941 cacgcttccc gaagggagaa aggcggacag gtatccggta agcggcaggg tcggaacagg
6001 agagcgcacg agggagcttc cagggggaaa cgcctggtat ctttatagtc ctgtcgggtt
6061 tcgccacctc tgacttgagc gtcgattttt gtgatgctcg tcaggggggc ggagcctatg
6121 gaaaaacgcc agcaacgcgg cctttttacg gttcctggcc ttttgctggc cttttgctca
6181 catgttcttt cctgcgttat cccctgattc tgtggataac cgtattaccg cctttgagtg
6241 agctgatacc gctcgccgca gccgaacgac cgagcgcagc gagtcagtga gcgaggaagc
6301 ggaagagcgc ccaatacgca aaccgcctct ccccgcgcgt tggccgattc attaatgcag
6361 ctggcacgac aggtttcccg actggaaagc gggcagtgag cgcaacgcaa ttaatgtgag
6421 ttagctcact cattaggcac cccaggcttt acactttatg cttccggctc gtatgttgtg
6481 tggaattgtg agcggataac aatttcacac aggaaacagc tatgaccatg attacgccaa
6541 gctcgaaatt aaccctcact aaagggaaca aaagctgcgg gccccccctc gaagatctct
6601 cgaggctagc atgcatatcg attaccgttc gtataatgta tgctatacga agttatacta
6661 gtaagcttga tatcgaat
//
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