LOCUS       sequence-211103-        7115 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  empty sgRNA cloning vector with MS2-AIDmono.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7115)
  AUTHORS   Liu LD, Huang M, Dai P, Liu T, Fan S, Cheng X, Zhao Y, Yeap LS, Meng
            FL
  TITLE     Intrinsic Nucleotide Preference of Diversifying Base Editors Guides 
            Antibody Ex Vivo Affinity Maturation.
  JOURNAL   Cell Rep. 2018 Oct 23;25(4):884-892.e3. doi: 
            10.1016/j.celrep.2018.09.090.
  PUBMED    30355495
REFERENCE   2  (bases 1 to 7115)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Dec 15, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; doi: 
            "10.1016/j.celrep.2018.09"; journalName: "Cell Rep"; date: 
            "2018-10-23- 23"; volume: "25"; issue: "4"; pages: "884-892.e3"
COMMENT     Sequence Label: pLL1 sgRNA(MS2)-MS2-AIDmono
FEATURES             Location/Qualifiers
     source          1..7115
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        1..249
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA 
                     (Domitrovich & Kunkel, 2003)"
     primer_bind     1..21
                     /label=hU6-F
                     /note="Human U6 promoter, forward primer"
     primer_bind     172..191
                     /label=LKO.1 5'
                     /note="Human U6 promoter, forward primer"
     promoter        341..445
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      1291..1879
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     misc_RNA        2093..2111
                     /label=MS2 stem loop
                     /note="stem loop that binds the bacteriophage MS2 coat 
                     protein"
     misc_RNA        2163..2181
                     /label=MS2 stem loop
                     /note="stem loop that binds the bacteriophage MS2 coat 
                     protein"
     enhancer        2309..2594
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer; 
                     contains an 18-bp deletion relative to the standard CMV 
                     enhancer"
     promoter        2596..2873
                     /label=chicken beta-actin promoter
     intron          2874..3102
                     /label=hybrid intron
                     /note="hybrid between chicken beta-actin (CBA) and minute 
                     virus of mice (MMV) introns (Gray et al., 2011)"
     CDS             3122..3508
                     /codon_start=1
                     /product="bacteriophage MS2 coat protein"
                     /label=MS2-N55K
                     /note="binds with high affinity to a specific stem-loop 
                     structure in the viral RNA (Lim et al., 1994)"
                     /translation="ASNFTQFVLVDNGGTGDVTVAPSNFANGVAEWISSNSRSQAYKVT
                     CSVRQSSAQKRKYTIKVEVPKVATQTVGGVELPVAAWRSYLNMELTIPIFATNSDCELI
                     VKAMQGLLKDGNPIPSAIAANSGIY"
     CDS             3563..3583
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     primer_bind     complement(4156..4173)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    4162..4369
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     repeat_region   4378..4518
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   4378..4507
                     /label=AAV2 ITR
     rep_origin      4593..5048
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4680..4699)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4890..4911
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(5065..5084)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     5184..5206
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     promoter        5330..5434
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5435..5503,5504..6295)
                     /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"
     rep_origin      6466..7054
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
ORIGIN
        1 gagggcctat ttcccatgat tccttcatat ttgcatatac gatacaaggc tgttagagag
       61 ataattggaa ttaatttgac tgtaaacaca aagatattag tacaaaatac gtgacgtaga
      121 aagtaataat ttcttgggta gtttgcagtt ttaaaattat gttttaaaat ggactatcat
      181 atgcttaccg taacttgaaa gtatttcgat ttcttggctt tatatatctt gtggaaagga
      241 cgaaacaccg ggtcttctcg tgatacgcct atttttatag gttaatgtca tgataataat
      301 ggtttcttag acgtcaggtg gcacttttcg gggaaatgtg cgcggaaccc ctatttgttt
      361 atttttctaa atacattcaa atatgtatcc gctcatgaga caataaccct gataaatgct
      421 tcaataatat tgaaaaagga agagtatgag tattcaacat ttccgtgtcg cccttattcc
      481 cttttttgcg gcattttgcc ttcctgtttt tgctcaccca gaaacgctgg tgaaagtaaa
      541 agatgctgaa gatcagttgg gtgcacgagt gggttacatc gaactggatc tcaacagcgg
      601 taagatcctt gagagttttc gccccgaaga acgttttcca atgatgagca cttttaaagt
      661 tctgctatgt ggcgcggtat tatcccgtat tgacgccggg caagagcaac tcggtcgccg
      721 catacactat tctcagaatg acttggttga gtactcacca gtcacagaaa agcatcttac
      781 ggatggcatg acagtaagag aattatgcag tgctgccata accatgagtg ataacactgc
      841 ggccaactta cttctgacaa cgatcggagg accgaaggag ctaaccgctt ttttgcccaa
      901 catgggggat catgtaactc gccttgatcg ttgggaaccg gagctgaatg aagtaatttg
      961 gagccggtga gcgtggtctc gcggtatcat tgcagcactg gggccagatg gtaagccctc
     1021 ccgtatcgta gttatctaca cgacggggag tcaggcaact atggatgaac gaaatagaca
     1081 gatcgctgag ataggtgcct cactgattaa gcattggtaa ctgtcagacc aagtttactc
     1141 atatatactt tagattgatt taaaacttca tttttaattt aaaaggatct aggtgaagat
     1201 cctttttgat aatctcatga ccaaaatccc ttaacgtgag ttttcgttcc actgagcgtc
     1261 agaccccgta gaaaagatca aaggatcttc ttgagatcct ttttttctgc gcgtaatctg
     1321 ctgcttgcaa acaaaaaaac caccgctacc agcggtggtt tgtttgccgg atcaagagct
     1381 accaactctt tttccgaagg taactggctt cagcagagcg cagataccaa atactgttct
     1441 tctagtgtag ccgtagttag gccaccactt caagaactct gtagcaccgc ctacatacct
     1501 cgctctgcta atcctgttac cagtggctgc tgccagtggc gataagtcgt gtcttaccgg
     1561 gttggactca agacgatagt taccggataa ggcgcagcgg tcgggctgaa cggggggttc
     1621 gtgcacacag cccagcttgg agcgaacgac ctacaccgaa ctgagatacc tacagcgtga
     1681 gctatgagaa agcgccacgc ttcccgaagg gagaaaggcg gacaggtatc cggtaagcgg
     1741 cagggtcgga acaggagagc gcacgaggga gcttccaggg ggaaacgcct ggtatcttta
     1801 tagtcctgtc gggtttcgcc acctctgact tgagcgtcga tttttgtgat gctcgtcagg
     1861 ggggcggagc ctatggaaaa acgccagcaa cgcggccttt ttacggttcc tggccttttg
     1921 ctggcctttt gctcacatgt tctttcctgc gttatcccct gattctgtgg ataaccgtat
     1981 taccgccttt gagtgagctg ataccgctcg ccgcagccga acgaccgagc gcagcgagtc
     2041 agtgagcgag gaagcggaag agcgcgagaa gacctgtttt agagctaggc caacatgagg
     2101 atcacccatg tctgcagggc ctagcaagtt aaaataaggc tagtccgtta tcaacttggc
     2161 caacatgagg atcacccatg tctgcagggc caagtggcac cgagtcggtg ctttttttgt
     2221 tttagagcta gaaatagcaa gttaaaataa ggctagtccg tttttagcgc gtgcgccaat
     2281 tctgcagaca aatggctcta gaggtacccg ttacataact tacggtaaat ggcccgcctg
     2341 gctgaccgcc caacgacccc cgcccattga cgtcaatagt aacgccaata gggactttcc
     2401 attgacgtca atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt
     2461 atcatatgcc aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt
     2521 gtgcccagta catgacctta tgggactttc ctacttggca gtacatctac gtattagtca
     2581 tcgctattac catggtcgag gtgagcccca cgttctgctt cactctcccc atctcccccc
     2641 cctccccacc cccaattttg tatttattta ttttttaatt attttgtgca gcgatggggg
     2701 cggggggggg gggggggcgc gcgccaggcg gggcggggcg gggcgagggg cggggcgggg
     2761 cgaggcggag aggtgcggcg gcagccaatc agagcggcgc gctccgaaag tttcctttta
     2821 tggcgaggcg gcggcggcgg cggccctata aaaagcgaag cgcgcggcgg gcgggagtcg
     2881 ctgcgacgct gccttcgccc cgtgccccgc tccgccgccg cctcgcgccg cccgccccgg
     2941 ctctgactga ccgcgttact cccacaggtg agcgggcggg acggcccttc tcctccgggc
     3001 tgtaattagc tgagcaagag gtaagggttt aagggatggt tggttggtgg ggtattaatg
     3061 tttaattacc tggagcacct gcctgaaatc actttttttc aggttggacc ggtccaccat
     3121 ggcttcaaac tttactcagt tcgtgctcgt ggacaatggt gggacagggg atgtgacagt
     3181 ggctccttct aatttcgcta atggggtggc agagtggatc agctccaact cacggagcca
     3241 ggcctacaag gtgacatgca gcgtcaggca gtctagtgcc cagaagagaa agtataccat
     3301 caaggtggag gtccccaaag tggctaccca gacagtgggc ggagtcgaac tgcctgtcgc
     3361 cgcttggagg tcctacctga acatggagct cactatccca attttcgcta ccaattctga
     3421 ctgtgaactc atcgtgaagg caatgcaggg gctcctcaaa gacggtaatc ctatcccttc
     3481 cgccatcgcc gctaactcag gtatctacag cgctggagga ggtggaagcg gaggaggagg
     3541 aagcggagga ggaggtagcg gacctaagaa aaagaggaag gtggcggccg ctggaatgga
     3601 ccccgctacc ttcacatacc aattcaaaaa tgtccgctgg gctaagggtc ggcgtgagac
     3661 ctacctgtgc tacgtagtga agaggcgtga cagtgctaca tccttttcac tggactttgg
     3721 ttatcttcgc aataagaacg gctgccacgt ggaattgctc ttcctccgct acatctcgga
     3781 ctgggaccta gaccctggcc gctgctaccg cgtcacctgg ttcacctcct ggagcccctg
     3841 ctacgactgt gcccgacatg tggccgactt tctgcgaggg aaccccaacc tcagtctgag
     3901 gatcttcacc gcgcgcctct acttctgtga ggaccgcaag gctgagcccg aggggctgcg
     3961 gcggctggca gaagccgggg tgcaaatagc catcatgacc ttcaaagatt atttttactg
     4021 ctggaatact tttgtagaaa accatgaaag aactttcaaa gcctgggaag ggctgcatga
     4081 aaattcagtt cgtctctcca gacagcttcg gcgcatcctt cagtaagatc ctaattccta
     4141 gagctcgctg atcagcctcg actgtgcctt ctagttgcca gccatctgtt gtttgcccct
     4201 cccccgtgcc ttccttgacc ctggaaggtg ccactcccac tgtcctttcc taataaaatg
     4261 aggaaattgc atcgcattgt ctgagtaggt gtcattctat tctggggggt ggggtggggc
     4321 aggacagcaa gggggaggat tgggaagaga atagcaggca tgctggggag cggccgcagg
     4381 aacccctagt gatggagttg gccactccct ctctgcgcgc tcgctcgctc actgaggccg
     4441 ggcgaccaaa ggtcgcccga cgcccgggct ttgcccgggc ggcctcagtg agcgagcgag
     4501 cgcgcagctg cctgcagggg cgcctgatgc ggtattttct ccttacgcat ctgtgcggta
     4561 tttcacaccg catacgtcaa agcaaccata gtacgcgccc tgtagcggcg cattaagcgc
     4621 ggcgggtgtg gtggttacgc gcagcgtgac cgctacactt gccagcgccc tagcgcccgc
     4681 tcctttcgct ttcttccctt cctttctcgc cacgttcgcc ggctttcccc gtcaagctct
     4741 aaatcggggg ctccctttag ggttccgatt tagtgcttta cggcacctcg accccaaaaa
     4801 acttgatttg ggtgatggtt cacgtagtgg gccatcgccc tgatagacgg tttttcgccc
     4861 tttgacgttg gagtccacgt tctttaatag tggactcttg ttccaaactg gaacaacact
     4921 caaccctatc tcgggctatt cttttgattt ataagggatt ttgccgattt cggcctattg
     4981 gttaaaaaat gagctgattt aacaaaaatt taacgcgaat tttaacaaaa tattaacgtt
     5041 tacaatttta tggtgcactc tcagtacaat ctgctctgat gccgcatagt taagccagcc
     5101 ccgacacccg ccaacacccg ctgacgcgcc ctgacgggct tgtctgctcc cggcatccgc
     5161 ttacagacaa gctgtgaccg tctccgggag ctgcatgtgt cagaggtttt caccgtcatc
     5221 accgaaacgc gcgagacgaa agggcctcgt gatacgccta tttttatagg ttaatgtcat
     5281 gataataatg gtttcttaga cgtcaggtgg cacttttcgg ggaaatgtgc gcggaacccc
     5341 tatttgttta tttttctaaa tacattcaaa tatgtatccg ctcatgagac aataaccctg
     5401 ataaatgctt caataatatt gaaaaaggaa gagtatgagt attcaacatt tccgtgtcgc
     5461 ccttattccc ttttttgcgg cattttgcct tcctgttttt gctcacccag aaacgctggt
     5521 gaaagtaaaa gatgctgaag atcagttggg tgcacgagtg ggttacatcg aactggatct
     5581 caacagcggt aagatccttg agagttttcg ccccgaagaa cgttttccaa tgatgagcac
     5641 ttttaaagtt ctgctatgtg gcgcggtatt atcccgtatt gacgccgggc aagagcaact
     5701 cggtcgccgc atacactatt ctcagaatga cttggttgag tactcaccag tcacagaaaa
     5761 gcatcttacg gatggcatga cagtaagaga attatgcagt gctgccataa ccatgagtga
     5821 taacactgcg gccaacttac ttctgacaac gatcggagga ccgaaggagc taaccgcttt
     5881 tttgcacaac atgggggatc atgtaactcg ccttgatcgt tgggaaccgg agctgaatga
     5941 agccatacca aacgacgagc gtgacaccac gatgcctgta gcaatggcaa caacgttgcg
     6001 caaactatta actggcgaac tacttactct agcttcccgg caacaattaa tagactggat
     6061 ggaggcggat aaagttgcag gaccacttct gcgctcggcc cttccggctg gctggtttat
     6121 tgctgataaa tctggagccg gtgagcgtgg aagccgcggt atcattgcag cactggggcc
     6181 agatggtaag ccctcccgta tcgtagttat ctacacgacg gggagtcagg caactatgga
     6241 tgaacgaaat agacagatcg ctgagatagg tgcctcactg attaagcatt ggtaactgtc
     6301 agaccaagtt tactcatata tactttagat tgatttaaaa cttcattttt aatttaaaag
     6361 gatctaggtg aagatccttt ttgataatct catgaccaaa atcccttaac gtgagttttc
     6421 gttccactga gcgtcagacc ccgtagaaaa gatcaaagga tcttcttgag atcctttttt
     6481 tctgcgcgta atctgctgct tgcaaacaaa aaaaccaccg ctaccagcgg tggtttgttt
     6541 gccggatcaa gagctaccaa ctctttttcc gaaggtaact ggcttcagca gagcgcagat
     6601 accaaatact gtccttctag tgtagccgta gttaggccac cacttcaaga actctgtagc
     6661 accgcctaca tacctcgctc tgctaatcct gttaccagtg gctgctgcca gtggcgataa
     6721 gtcgtgtctt accgggttgg actcaagacg atagttaccg gataaggcgc agcggtcggg
     6781 ctgaacgggg ggttcgtgca cacagcccag cttggagcga acgacctaca ccgaactgag
     6841 atacctacag cgtgagctat gagaaagcgc cacgcttccc gaagggagaa aggcggacag
     6901 gtatccggta agcggcaggg tcggaacagg agagcgcacg agggagcttc cagggggaaa
     6961 cgcctggtat ctttatagtc ctgtcgggtt tcgccacctc tgacttgagc gtcgattttt
     7021 gtgatgctcg tcaggggggc ggagcctatg gaaaaacgcc agcaacgcgg cctttttacg
     7081 gttcctggcc ttttgctggc cttttgctca catgt
//
