pLH-sgRNA1-PP7-boxB

LOCUS       pLH-sgRNA1-PP7-boxB                8307 bp ds-DNA     circular SYN 11-FEB-2020
DEFINITION  sgRNA1-PP7-boxB.
ACCESSION   .
VERSION     .
KEYWORDS    pLH-sgRNA1-PP7-boxB
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8307)
  AUTHORS   Ma H, Tu LC, Naseri A, Huisman M, Zhang S, Grunwald D, Pederson T
  TITLE     Multiplexed labeling of genomic loci with dCas9 and engineered 
            sgRNAs using CRISPRainbow.
  JOURNAL   Nat Biotechnol. 2016 Apr 18. doi: 10.1038/nbt.3526.
  PUBMED    27088723
REFERENCE   2  (bases 1 to 8307)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Feb 11, 2020 from SnapGene Server 1.1.58
            http://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..8307
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     primer_bind     123..142
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     376..393
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    510..531
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        546..576
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    584..600
                     /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     589..611
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     608..624
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     608..624
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     643..663
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        645..663
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     promoter        691..917
                     /label=RSV promoter
                     /note="Rous sarcoma virus enhancer/promoter"
     LTR             918..1098
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     misc_feature    1145..1270
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     misc_feature    1763..1996
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     promoter        2523..2763
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA"
     primer_bind     2523..2543
                     /label=hU6-F
                     /note="Human U6 promoter, forward primer"
     primer_bind     2694..2713
                     /label=LKO.1 5'
                     /note="Human U6 promoter, forward primer"
     CDS             3112..3417
                     /codon_start=1
                     /gene="ccdB"
                     /product="CcdB, a bacterial toxin that poisons DNA gyrase"
                     /label=ccdB
                     /note="Plasmids containing the ccdB gene cannot be 
                     propagated in standard E. coli strains."
                     /translation="MQFKVYTYKRESRYRLFVDVQSDIIDTPGRRMVIPLASARLLSDK
                     VSRELYPVVHIGDESWRMMTTDMASVPVSVIGEEVADLSHRENDIKNAINLMFWGI"
     primer_bind     3296..3315
                     /label=ccdB-fwd
                     /note="ccdB gene, forward primer"
     misc_feature    3602..3719
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
     promoter        3768..4278
                     /label=hPGK promoter
                     /note="human phosphoglycerate kinase 1 promoter"
     primer_bind     4197..4215
                     /label=hPGK-F
                     /note="Human PGK promoter, forward primer"
     CDS             4368..5393
                     /codon_start=1
                     /gene="aph(4)-Ia"
                     /product="aminoglycoside phosphotransferase from E. coli"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="MKKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRG
                     YVLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLP
                     ETELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQ
                     TVMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMF
                     GDSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDG
                     NFDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAK
                     E"
     primer_bind     complement(5417..5433)
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5418..5434)
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     LTR             5516..5749
                     /label=3' LTR (Delta-U3)
                     /note="self-inactivating 3' long terminal repeat (LTR) from
                     HIV-1"
     polyA_signal    5821..5942
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5858..5877)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     5912..5931
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      5982..6117
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     6044..6063
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     primer_bind     complement(6137..6156)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(6138..6156)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(6166..6183)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(6166..6182)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(6175..6197)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      6324..6779
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(6411..6430)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     6621..6642
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        6805..6909
                     /gene="bla"
                     /label=AmpR promoter
     CDS             6910..7770
                     /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(7128..7147)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      7941..222
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
ORIGIN
        1 gaactgagat acctacagcg tgagctatga gaaagcgcca cgcttcccga agggagaaag
       61 gcggacaggt atccggtaag cggcagggtc ggaacaggag agcgcacgag ggagcttcca
      121 gggggaaacg cctggtatct ttatagtcct gtcgggtttc gccacctctg acttgagcgt
      181 cgatttttgt gatgctcgtc aggggggcgg agcctatgga aaaacgccag caacgcggcc
      241 tttttacggt tcctggcctt ttgctggcct tttgctcaca tgttctttcc tgcgttatcc
      301 cctgattctg tggataaccg tattaccgcc tttgagtgag ctgataccgc tcgccgcagc
      361 cgaacgaccg agcgcagcga gtcagtgagc gaggaagcgg aagagcgccc aatacgcaaa
      421 ccgcctctcc ccgcgcgttg gccgattcat taatgcagct ggcacgacag gtttcccgac
      481 tggaaagcgg gcagtgagcg caacgcaatt aatgtgagtt agctcactca ttaggcaccc
      541 caggctttac actttatgct tccggctcgt atgttgtgtg gaattgtgag cggataacaa
      601 tttcacacag gaaacagcta tgaccatgat tacgccaagc gcgcaattaa ccctcactaa
      661 agggaacaaa agctggagct gcaagcttaa tgtagtctta tgcaatactc ttgtagtctt
      721 gcaacatggt aacgatgagt tagcaacatg ccttacaagg agagaaaaag caccgtgcat
      781 gccgattggt ggaagtaagg tggtacgatc gtgccttatt aggaaggcaa cagacgggtc
      841 tgacatggat tggacgaacc actgaattgc cgcattgcag agatattgta tttaagtgcc
      901 tagctcgata cataaacggg tctctctggt tagaccagat ctgagcctgg gagctctctg
      961 gctaactagg gaacccactg cttaagcctc aataaagctt gccttgagtg cttcaagtag
     1021 tgtgtgcccg tctgttgtgt gactctggta actagagatc cctcagaccc ttttagtcag
     1081 tgtggaaaat ctctagcagt ggcgcccgaa cagggacttg aaagcgaaag ggaaaccaga
     1141 ggagctctct cgacgcagga ctcggcttgc tgaagcgcgc acggcaagag gcgaggggcg
     1201 gcgactggtg agtacgccaa aaattttgac tagcggaggc tagaaggaga gagatgggtg
     1261 cgagagcgtc agtattaagc gggggagaat tagatcgcga tgggaaaaaa ttcggttaag
     1321 gccaggggga aagaaaaaat ataaattaaa acatatagta tgggcaagca gggagctaga
     1381 acgattcgca gttaatcctg gcctgttaga aacatcagaa ggctgtagac aaatactggg
     1441 acagctacaa ccatcccttc agacaggatc agaagaactt agatcattat ataatacagt
     1501 agcaaccctc tattgtgtgc atcaaaggat agagataaaa gacaccaagg aagctttaga
     1561 caagatagag gaagagcaaa acaaaagtaa gaccaccgca cagcaagcgg ccgctgatct
     1621 tcagacctgg aggaggagat atgagggaca attggagaag tgaattatat aaatataaag
     1681 tagtaaaaat tgaaccatta ggagtagcac ccaccaaggc aaagagaaga gtggtgcaga
     1741 gagaaaaaag agcagtggga ataggagctt tgttccttgg gttcttggga gcagcaggaa
     1801 gcactatggg cgcagcgtca atgacgctga cggtacaggc cagacaatta ttgtctggta
     1861 tagtgcagca gcagaacaat ttgctgaggg ctattgaggc gcaacagcat ctgttgcaac
     1921 tcacagtctg gggcatcaag cagctccagg caagaatcct ggctgtggaa agatacctaa
     1981 aggatcaaca gctcctgggg atttggggtt gctctggaaa actcatttgc accactgctg
     2041 tgccttggaa tgctagttgg agtaataaat ctctggaaca gatttggaat cacacgacct
     2101 ggatggagtg ggacagagaa attaacaatt acacaagctt aatacactcc ttaattgaag
     2161 aatcgcaaaa ccagcaagaa aagaatgaac aagaattatt ggaattagat aaatgggcaa
     2221 gtttgtggaa ttggtttaac ataacaaatt ggctgtggta tataaaatta ttcataatga
     2281 tagtaggagg cttggtaggt ttaagaatag tttttgctgt actttctata gtgaatagag
     2341 ttaggcaggg atattcacca ttatcgtttc agacccacct cccaaccccg aggggacccg
     2401 acaggcccga aggaatagaa gaagaaggtg gagagagaga cagagacaga tccattcgat
     2461 tagtgaacgg atctcgacgg tatcgatcac gagactagcc tcgagcggcc gcccccttca
     2521 ccgagggcct atttcccatg attccttcat atttgcatat acgatacaag gctgttagag
     2581 agataattgg aattaatttg actgtaaaca caaagatatt agtacaaaat acgtgacgta
     2641 gaaagtaata atttcttggg tagtttgcag ttttaaaatt atgttttaaa atggactatc
     2701 atatgcttac cgtaacttga aagtatttcg atttcttggc tttatatatc ttgtggaaag
     2761 gacgaaacac cggtgtcttc gaggcttact aaaagccaga taacagtatg cgtatttgcg
     2821 cgctgatttt tgcggtataa gaatatatac tgatatgtat acccgaagta tgtcaaaaag
     2881 aggtgtgcta tgaagcagcg tattacagtg acagttgaca gcgacagcta tcagttgctc
     2941 aaggcatata tgatgtcaat atctccggtc tggtaagcac aaccatgcag aatgaagccc
     3001 gtcgtctgcg tgccgaacgc tggaaagcgg aaaatcagga agggatggct gaggtcgccc
     3061 ggtttattga aatgaacggc tcttttgctg acgagaacag ggactggtga aatgcagttt
     3121 aaggtttaca cctataaaag agagagccgt tatcgtctgt ttgtggatgt acagagtgat
     3181 attattgaca cgcccgggcg acggatggtg atccccctgg ccagtgcacg tctgctgtca
     3241 gataaagtct cccgtgaact ttacccggtg gtgcatatcg gggatgaaag ctggcgcatg
     3301 atgaccaccg atatggccag tgtgccggtc tccgttatcg gggaagaagt ggctgatctc
     3361 agccaccgcg aaaatgacat caaaaacgcc attaacctga tgttctgggg aatataaacc
     3421 ctgcaggacg aagacccgtt tgagagctac cggagcagac gatatggcgt cgctccggta
     3481 gcaagttcaa ataaggctag tccgttatca acttgggccc tgaagaaggg cccaagtggc
     3541 accgagtcgg tgcttttttt gaattctcga cctcgagaca aatggcagta ttcatccaca
     3601 attttaaaag aaaagggggg attggggggt acagtgcagg ggaaagaata gtagacataa
     3661 tagcaacaga catacaaact aaagaattac aaaaacaaat tacaaaaatt caaaattttc
     3721 gggtttatta cagggacagc agagatccac tttggccgcg gctcgagggg gttggggttg
     3781 cgccttttcc aaggcagccc tgggtttgcg cagggacgcg gctgctctgg gcgtggttcc
     3841 gggaaacgca gcggcgccga ccctgggtct cgcacattct tcacgtccgt tcgcagcgtc
     3901 acccggatct tcgccgctac ccttgtgggc cccccggcga cgcttcctgc tccgccccta
     3961 agtcgggaag gttccttgcg gttcgcggcg tgccggacgt gacaaacgga agccgcacgt
     4021 ctcactagta ccctcgcaga cggacagcgc cagggagcaa tggcagcgcg ccgaccgcga
     4081 tgggctgtgg ccaatagcgg ctgctcagca gggcgcgccg agagcagcgg ccgggaaggg
     4141 gcggtgcggg aggcggggtg tggggcggta gtgtgggccc tgttcctgcc cgcgcggtgt
     4201 tccgcattct gcaagcctcc ggagcgcacg tcggcagtcg gctccctcgt tgaccgaatc
     4261 accgacctct ctccccaggg ggatccaccg gatcaagctt gccagctggg gcgccctctg
     4321 gtaaggttgg gaagccctgc agatctgatc ccggggggca atgagatatg aaaaagcctg
     4381 aactcaccgc gacgtctgtc gagaagtttc tgatcgaaaa gttcgacagc gtctccgacc
     4441 tgatgcagct ctcggagggc gaagaatctc gtgctttcag cttcgatgta ggagggcgtg
     4501 gatatgtcct gcgggtaaat agctgcgccg atggtttcta caaagatcgt tatgtttatc
     4561 ggcactttgc atcggccgcg ctcccgattc cggaagtgct tgacattggg gaatttagcg
     4621 agagcctgac ctattgcatc tcccgccgtg cacagggtgt cacgttgcaa gatctgcctg
     4681 aaaccgaact gcccgctgtt ctgcagccgg tcgcggaggc catggatgcg atcgctgcgg
     4741 ccgatcttag ccagacgagc gggttcggcc cattcggacc gcaaggaatc ggtcaataca
     4801 ctacatggcg tgatttcata tgcgcgattg ctgatcccca tgtgtatcac tggcaaactg
     4861 tgatggacga caccgtcagt gcgtccgtcg cgcaggctct cgatgagctg atgctttggg
     4921 ccgaggactg ccccgaagtc cggcacctcg tgcacgcgga tttcggctcc aacaatgtcc
     4981 tgacggacaa tggccgcata acagcggtca ttgactggag cgaggcgatg ttcggggatt
     5041 cccaatacga ggtcgccaac atcttcttct ggaggccgtg gttggcttgt atggagcagc
     5101 agacgcgcta cttcgagcgg aggcatccgg agcttgcagg atcgccgcgg ctccgggcgt
     5161 atatgctccg cattggtctt gaccaactct atcagagctt ggttgacggc aatttcgatg
     5221 atgcagcttg ggcgcagggt cgatgcgacg caatcgtccg atccggagcc gggactgtcg
     5281 ggcgtacaca aatcgcccgc agaagcgcgg ccgtctggac cgatggctgt gtagaagtac
     5341 tcgccgatag tggaaaccga cgccccagca ctcgtccgag ggcaaaggaa tagagtagat
     5401 gccgaccggg atctatcgat accgtcgacc tcgagggggg gcccggtacc tttaagacca
     5461 atgacttaca aggcagctgt agatcttagc cactttttaa aagaaaaggg gggactggaa
     5521 gggctaattc actcccaacg aagaaaagat ctgctttttg cttgtactgg gtctctctgg
     5581 ttagaccaga tctgagcctg ggagctctct ggctaactag ggaacccact gcttaagcct
     5641 caataaagct tgccttgagt gcttcaagta gtgtgtgccc gtctgttgtg tgactctggt
     5701 aactagagat ccctcagacc cttttagtca gtgtggaaaa tctctagcag tagtagttca
     5761 tgtcatctta ttattcagta tttataactt gcaaagaaat gaatatcaga gagtgagagg
     5821 aacttgttta ttgcagctta taatggttac aaataaagca atagcatcac aaatttcaca
     5881 aataaagcat ttttttcact gcattctagt tgtggtttgt ccaaactcat caatgtatct
     5941 tatcatgtct ggctctagct atcccgcccc taactccgcc catcccgccc ctaactccgc
     6001 ccagttccgc ccattctccg ccccatggct gactaatttt ttttatttat gcagaggccg
     6061 aggccgcctc ggcctctgag ctattccaga agtagtgagg aggctttttt ggaggcctag
     6121 ggacgtaccc aattcgccct atagtgagtc gtattacgcg cgctcactgg ccgtcgtttt
     6181 acaacgtcgt gactgggaaa accctggcgt tacccaactt aatcgccttg cagcacatcc
     6241 ccctttcgcc agctggcgta atagcgaaga ggcccgcacc gatcgccctt cccaacagtt
     6301 gcgcagcctg aatggcgaat gggacgcgcc ctgtagcggc gcattaagcg cggcgggtgt
     6361 ggtggttacg cgcagcgtga ccgctacact tgccagcgcc ctagcgcccg ctcctttcgc
     6421 tttcttccct tcctttctcg ccacgttcgc cggctttccc cgtcaagctc taaatcgggg
     6481 gctcccttta gggttccgat ttagtgcttt acggcacctc gaccccaaaa aacttgatta
     6541 gggtgatggt tcacgtagtg ggccatcgcc ctgatagacg gtttttcgcc ctttgacgtt
     6601 ggagtccacg ttctttaata gtggactctt gttccaaact ggaacaacac tcaaccctat
     6661 ctcggtctat tcttttgatt tataagggat tttgccgatt tcggcctatt ggttaaaaaa
     6721 tgagctgatt taacaaaaat ttaacgcgaa ttttaacaaa atattaacgc ttacaattta
     6781 ggtggcactt ttcggggaaa tgtgcgcgga acccctattt gtttattttt ctaaatacat
     6841 tcaaatatgt atccgctcat gagacaataa ccctgataaa tgcttcaata atattgaaaa
     6901 aggaagagta tgagtattca acatttccgt gtcgccctta ttcccttttt tgcggcattt
     6961 tgccttcctg tttttgctca cccagaaacg ctggtgaaag taaaagatgc tgaagatcag
     7021 ttgggtgcac gagtgggtta catcgaactg gatctcaaca gcggtaagat ccttgagagt
     7081 tttcgccccg aagaacgttt tccaatgatg agcactttta aagttctgct atgtggcgcg
     7141 gtattatccc gtattgacgc cgggcaagag caactcggtc gccgcataca ctattctcag
     7201 aatgacttgg ttgagtactc accagtcaca gaaaagcatc ttacggatgg catgacagta
     7261 agagaattat gcagtgctgc cataaccatg agtgataaca ctgcggccaa cttacttctg
     7321 acaacgatcg gaggaccgaa ggagctaacc gcttttttgc acaacatggg ggatcatgta
     7381 actcgccttg atcgttggga accggagctg aatgaagcca taccaaacga cgagcgtgac
     7441 accacgatgc ctgtagcaat ggcaacaacg ttgcgcaaac tattaactgg cgaactactt
     7501 actctagctt cccggcaaca attaatagac tggatggagg cggataaagt tgcaggacca
     7561 cttctgcgct cggcccttcc ggctggctgg tttattgctg ataaatctgg agccggtgag
     7621 cgtgggtctc gcggtatcat tgcagcactg gggccagatg gtaagccctc ccgtatcgta
     7681 gttatctaca cgacggggag tcaggcaact atggatgaac gaaatagaca gatcgctgag
     7741 ataggtgcct cactgattaa gcattggtaa ctgtcagacc aagtttactc atatatactt
     7801 tagattgatt taaaacttca tttttaattt aaaaggatct aggtgaagat cctttttgat
     7861 aatctcatga ccaaaatccc ttaacgtgag ttttcgttcc actgagcgtc agaccccgta
     7921 gaaaagatca aaggatcttc ttgagatcct ttttttctgc gcgtaatctg ctgcttgcaa
     7981 acaaaaaaac caccgctacc agcggtggtt tgtttgccgg atcaagagct accaactctt
     8041 tttccgaagg taactggctt cagcagagcg cagataccaa atactgttct tctagtgtag
     8101 ccgtagttag gccaccactt caagaactct gtagcaccgc ctacatacct cgctctgcta
     8161 atcctgttac cagtggctgc tgccagtggc gataagtcgt gtcttaccgg gttggactca
     8221 agacgatagt taccggataa ggcgcagcgg tcgggctgaa cggggggttc gtgcacacag
     8281 cccagcttgg agcgaacgac ctacacc
//