LOCUS       sequence-380232-        7057 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  sgRNA targeting alpha-satellites on human chromosome 9.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7057)
  AUTHORS   Lungu C, Pinter S, Broche J, Rathert P, Jeltsch A
  TITLE     Modular fluorescence complementation sensors for live cell detection
            of epigenetic signals at endogenous genomic sites.
  JOURNAL   Nat Commun. 2017 Sep 21;8(1):649. doi: 10.1038/s41467-017-00457-z.
  PUBMED    28935858
REFERENCE   2  (bases 1 to 7057)
  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"; journalName: "Nat 
            Commun."; date: "2017-09-21"; volume: "8(1):649. doi"; pages: " 
            10.1038/s41467-017-00457-z"
COMMENT     Sequence Label: pSIN-U6-tracer_C9_1_-EF1a-Thy1.1-P2A-Neo
FEATURES             Location/Qualifiers
     source          1..7057
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     58..77
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(296..400)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     468..486
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     enhancer        586..965
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        966..1169
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     1120..1140
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     LTR             1170..1345
                     /label=5' LTR (truncated)
                     /note="truncated long terminal repeat from Moloney murine 
                     sarcoma virus"
     misc_feature    1408..1607
                     /label=MMLV Psi
                     /note="packaging signal of Moloney murine leukemia virus 
                     (MMLV)"
     CDS             1808..2223
                     /label=gag (truncated)
                     /note="truncated Moloney murine leukemia virus (MMLV) gag 
                     gene lacking the start codon"
     primer_bind     2153..2175
                     /label=pLXSN 5'
                     /note="Murine stem cell virus, forward primer. Also called 
                     MSCV"
     primer_bind     2191..2207
                     /label=pBABE 5'
                     /note="Psi packaging signal, forward primer for pBABE 
                     vectors"
     promoter        2270..2518
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA 
                     (Domitrovich & Kunkel, 2003)"
     primer_bind     2270..2290
                     /label=hU6-F
                     /note="Human U6 promoter, forward primer"
     primer_bind     2441..2460
                     /label=LKO.1 5'
                     /note="Human U6 promoter, forward primer"
     misc_RNA        2540..2615
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
     promoter        2722..2933
                     /label=EF-1-alpha core promoter
                     /note="core promoter for human elongation factor 
                     EF-1-alpha"
     CDS             2953..3438
                     /codon_start=1
                     /product="mouse GPI-anchored cell surface glycoprotein of 
                     the immunoglobulin superfamily"
                     /label=Thy1.1
                     /note="This allele of Thy1 is also known as CD90.1."
                     /translation="MNPAISVALLLSVLQVSRGQKVTSLTACLVNQNLRLDCRHENNTK
                     DNSIQHEFSLTREKRKHVLSGTLGIPEHTYRSRVTLSNQPYIKVLTLANFTTKDEGDYF
                     CELRVSGANPMSSNKSISVYRDKLVKCGGISLLVQNTSWMLLLLLSLSLLQALDFISL"
     CDS             3448..3504
                     /codon_start=1
                     /product="2A peptide from porcine teschovirus-1 
                     polyprotein"
                     /label=P2A
                     /note="Eukaryotic ribosomes fail to insert a peptide bond 
                     between the Gly and Pro residues, yielding separate 
                     polypeptides."
                     /translation="ATNFSLLKQAGDVEENPGP"
     CDS             3499..4299
                     /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="GPMIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQG
                     RPVLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDL
                     LSSHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEE
                     HQGLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQD
                     IALATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(3559..3578)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4169..4188
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     LTR             4420..4845
                     /label=3' LTR (Delta-U3)
                     /note="self-inactivating 3' long terminal repeat (LTR) from
                     Moloney murine leukemia virus"
     primer_bind     complement(4948..4970)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(5106..5126)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        5111..5440
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      5291..5426
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     5462..5481
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     complement(5562..5579)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5733..6321)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5813..5832)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6492..7057,1..226,227..295))
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRVDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPAAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
ORIGIN
        1 cattctgaga atagtgtatg cggcgaccga gttgctcttg cccggcgtca acacgggata
       61 ataccgcgcc acatagcaga actttaaaag tgctcatcat tggaaaacgt tcttcggggc
      121 gaaaactctc aaggatctta ccgctgttga gatccagttc gatgtaaccc actcgtgcac
      181 ccaactgatc ttcagcatct tttactttca ccagcgtttc tgggtgagca aaaacaggaa
      241 ggcaaaatgc cgcaaaaaag ggaataaggg cgacacggaa atgttgaata ctcatactct
      301 tcctttttca atattattga agcatttatc agggttattg tctcatgagc ggatacatat
      361 ttgaatgtat ttagaaaaat aaacaaatag gggttccgcg cacatttccc cgaaaagtgc
      421 cacctgacgt ctaagaaacc attattatca tgacattaac ctataaaaat aggcgtatca
      481 cgaggccctt tcgtgttcaa gaattagctt ggccattgca tacgttgtat ccatatcata
      541 atatgtacat ttatattggc tcatgtccaa cattaccgcc atgttgacat tgattattga
      601 ctagttatta atagtaatca attacggggt cattagttca tagcccatat atggagttcc
      661 gcgttacata acttacggta aatggcccgc ctggctgacc gcccaacgac ccccgcccat
      721 tgacgtcaat aatgacgtat gttcccatag taacgccaat agggactttc cattgacgtc
      781 aatgggtgga gtatttacgg taaactgccc acttggcagt acatcaagtg tatcatatgc
      841 caagtacgcc ccctattgac gtcaatgacg gtaaatggcc cgcctggcat tatgcccagt
      901 acatgacctt atgggacttt cctacttggc agtacatcta cgtattagtc atcgctatta
      961 ccatggtgat gcggttttgg cagtacatca atgggcgtgg atagcggttt gactcacggg
     1021 gatttccaag tctccacccc attgacgtca atgggagttt gttttggcac caaaatcaac
     1081 gggactttcc aaaatgtcgt aacaactccg ccccattgac gcaaatgggc ggtaggcgtg
     1141 tacggtggga ggtctatata agcagagctc aataaaagag cccacaaccc ctcactcggc
     1201 gcgccagtcc tccgatagac tgcgtcgccc gggtacccgt attcccaata aagcctcttg
     1261 ctgtttgcat ccgaatcgtg gtctcgctgt tccttgggag ggtctcctct gagtgattga
     1321 ctacccacga cgggggtctt tcatttgggg gctcgtccgg gatttggaga cccctgccca
     1381 gggaccaccg acccaccacc gggaggtaag ctggccagca acttatctgt gtctgtccga
     1441 ttgtctagtg tctatgtttg atgttatgcg cctgcgtctg tactagttag ctaactagct
     1501 ctgtatctgg cggacccgtg gtggaactga cgagttctga acacccggcc gcaaccctgg
     1561 gagacgtccc agggactttg ggggccgttt ttgtggcccg acctgaggaa gggagtcgat
     1621 gtggaatccg accccgtcag gatatgtggt tctggtagga gacgagaacc taaaacagtt
     1681 cccgcctccg tctgaatttt tgctttcggt ttggaaccga agccgcgcgt cttgtctgct
     1741 gcagcgctgc agcatcgttc tgtgttgtct ctgtctgact gtgtttctgt atttgtctga
     1801 aaattagggc cagactgtta ccactccctt aagtttgacc ttaggtcact ggaaagatgt
     1861 cgagcggatc gctcacaacc agtcggtaga tgtcaagaag agacgttggt taccttctgc
     1921 tctgcagaat ggccaacctt taacgtcgga tggccgcgag acggcacctt taaccgagac
     1981 ctcatcaccc aggttaagat caaggtcttt tcacctggcc cgcatggaca cccagaccag
     2041 gtcccctaca tcgtgacctg ggaagccttg gcttttgacc cccctccctg ggtcaagccc
     2101 tttgtacacc ctaagcctcc gcctcctctt cctccatccg ccccgtctct cccccttgaa
     2161 cctcctcgtt cgaccccgcc tcgatcctcc ctttatccag ccctcactcc ttctctaggc
     2221 gccggaattg aagatcttta attaacgatc tatcgattct agatctagag agggcctatt
     2281 tcccatgatt ccttcatatt tgcatatacg atacaaggct gttagagaga taattagaat
     2341 taatttgact gtaaacacaa agatattagt acaaaatacg tgacgtagaa agtaataatt
     2401 tcttgggtag tttgcagttt taaaattatg ttttaaaatg gactatcata tgcttaccgt
     2461 aacttgaaag tatttcgatt tcttggcttt atatatcttg tggaaaggac gaaacaccgt
     2521 ggaatggaat ggaatggaag ttttagagct agaaatagca agttaaaata aggctagtcc
     2581 gttatcaact tgaaaaagtg gcaccgagtc ggtgcttttt tgaatttaga tcttgagaca
     2641 aggtgccggg ccgggaattc ggcgaattgg gcccgacgtc gcatgctccc ggcccctcga
     2701 gtggctccgg tgcccgtcag tgggcagagc gcacatcgcc cacagtcccc gagaagttgg
     2761 ggggaggggt cggcaattga accggtgcct agagaaggtg gcgcggggta aactgggaaa
     2821 gtgatgtcgt gtactggctc cgcctttttc ccgagggtgg gggagaaccg tatataagtg
     2881 cagtagtcgc cgtgaacgtt ctttttcgca acgggtttgc cgccagaaca caggtgtcgt
     2941 gacgcggcca ccatgaaccc agccatcagc gtcgctctcc tgctctcagt cttgcaggtg
     3001 tcccgagggc agaaggtgac cagcctgaca gcctgcctgg tgaaccaaaa ccttcgcctg
     3061 gactgccgcc atgagaataa caccaaggat aactccatcc agcatgagtt cagcctgacc
     3121 cgagagaaga ggaagcacgt gctctcaggc accctcggga tacccgagca cacgtaccgc
     3181 tcccgcgtca ccctctccaa ccagccctat atcaaggtcc ttaccctagc caacttcacc
     3241 accaaggatg agggcgacta cttttgtgag cttcgagtct cgggcgcgaa tcccatgagc
     3301 tccaataaaa gtatcagtgt gtatagagac aaactggtca agtgtggcgg cataagcctg
     3361 ctggttcaga acacatcctg gatgctgctg ctgctgcttt ccctctccct cctccaagcc
     3421 ctggacttca tttctctggg cagcggcgcc accaacttca gcctgctgaa gcaggccggc
     3481 gacgtggagg agaaccccgg ccccatgatt gaacaagatg gattgcacgc aggttctccg
     3541 gccgcttggg tggagaggct attcggctat gactgggcac aacagacaat cggctgctct
     3601 gatgccgccg tgttccggct gtcagcgcag gggcgcccgg ttctttttgt caagaccgac
     3661 ctgtccggtg ccctgaatga actgcaggac gaggcagcgc ggctatcgtg gctggccacg
     3721 acgggcgttc cttgcgcagc tgtgctcgac gttgtcactg aagcgggaag ggactggctg
     3781 ctattgggcg aagtgccggg gcaggatctc ctgtcatctc accttgctcc tgccgagaaa
     3841 gtatccatca tggctgatgc aatgcggcgg ctgcatacgc ttgatccggc tacctgccca
     3901 ttcgaccacc aagcgaaaca tcgcatcgag cgagcacgta ctcggatgga agccggtctt
     3961 gtcgatcagg atgatctgga cgaagagcat caggggctcg cgccagccga actgttcgcc
     4021 aggctcaagg cgcgcatgcc cgacggcgag gatctcgtcg tgacccatgg cgatgcctgc
     4081 ttgccgaata tcatggtgga aaatggccgc ttttctggat tcatcgactg tggccggctg
     4141 ggtgtggcgg accgctatca ggacatagcg ttggctaccc gtgatattgc tgaagagctt
     4201 ggcggcgaat gggctgaccg cttcctcgtg ctttacggta tcgccgctcc cgattcgcag
     4261 cgcatcgcct tctatcgcct tcttgacgag ttcttctgat tagatatcat cagtggtcca
     4321 ggctctagtt ttgactcaac aatatcacca gctgaagcct atagagtacg agccatagat
     4381 aaaataaaag attttattta gtctccagaa aaagggggga atgaaagacc ccacctgtag
     4441 gtttggcaag ctagcttaag taacgccatt ttgcaaggca tggaaaaata cataactgag
     4501 aatagagaag ttcagatcaa ggtcaggaac agatggaaca gggtcgaccc tagagaacca
     4561 tcagatgttt ccagggtgcc ccaaggacct gaaatgaccc tgtgccttat ttgaactaac
     4621 caatcagttc gcttctcgct tctgttcgcg cgcttctgct ccccgagctc aataaaagag
     4681 cccacaaccc ctcactcggg gcgccagtcc tccgattgac tgagtcgccc gggtacccgt
     4741 gtatccaata aaccctcttg cagttgcatc cgacttgtgg tctcgctgtt ccttgggagg
     4801 gtctcctctg agtgattgac tacccgtcag cgggggtctt tcatttgggg gctcgtccgg
     4861 gatcgggaga cccctgccca gggaccaccg acccaccacc gggaggtaag ctggctgcct
     4921 cgcgcgtttc ggtgatgacg gtgaaaacct ctgacacatg cagctcccgg agacggtcac
     4981 agcttgtctg taagcggatg ccgggagcag acaagcccgt cagggcgcgt cagcgggtgt
     5041 tggcgggtgt cggggcgcag ccatgaccca gtcacgtagc gatagcggag tgtagatccg
     5101 gctgtggaat gtgtgtcagt tagggtgtgg aaagtcccca ggctccccag caggcagaag
     5161 tatgcaaagc atgcatctca attagtcagc aaccaggtgt ggaaagtccc caggctcccc
     5221 agcaggcaga agtatgcaaa gcatgcatct caattagtca gcaaccatag tcccgcccct
     5281 aactccgccc atcccgcccc taactccgcc cagttccgcc cattctccgc cccatggctg
     5341 actaattttt tttatttatg cagaggccga ggccgcctcg gcctctgagc tattccagaa
     5401 gtagtgagga ggcttttttg gaggcctagg cttttgcaaa aagcttactg gcttaactat
     5461 gcggcatcag agcagattgt actgagagtg caccatatgc ggtgtgaaat accgcacaga
     5521 tgcgtaagga gaaaataccg catcaggcgc tcttccgctt cctcgctcac tgactcgctg
     5581 cgctcggtcg ttcggctgcg gcgagcggta tcagctcact caaaggcggt aatacggtta
     5641 tccacagaat caggggataa cgcaggaaag aacatgtgag caaaaggcca gcaaaaggcc
     5701 aggaaccgta aaaaggccgc gttgctggcg tttttccata ggctccgccc ccctgacgag
     5761 catcacaaaa atcgacgctc aagtcagagg tggcgaaacc cgacaggact ataaagatac
     5821 caggcgtttc cccctggaag ctccctcgtg cgctctcctg ttccgaccct gccgcttacc
     5881 ggatacctgt ccgcctttct cccttcggga agcgtggcgc tttctcatag ctcacgctgt
     5941 aggtatctca gttcggtgta ggtcgttcgc tccaagctgg gctgtgtgca cgaacccccc
     6001 gttcagcccg accgctgcgc cttatccggt aactatcgtc ttgagtccaa cccggtaaga
     6061 cacgacttat cgccactggc agcagccact ggtaacagga ttagcagagc gaggtatgta
     6121 ggcggtgcta cagagttctt gaagtggtgg cctaactacg gctacactag aaggacagta
     6181 tttggtatct gcgctctgct gaagccagtt accttcggaa aaagagttgg tagctcttga
     6241 tccggcaaac aaaccaccgc tggtagcggt ggtttttttg tttgcaagca gcagattacg
     6301 cgcagaaaaa aaggatctca agaagatcct ttgatctttt ctacggggtc tgacgctcag
     6361 tggaacgaaa actcacgtta agggattttg gtcatgagat tatcaaaaag gatcttcacc
     6421 tagatccttt taaattaaaa atgaagtttt aaatcaatct aaagtatata tgagtaaact
     6481 tggtctgaca gttaccaatg cttaatcagt gaggcaccta tctcagcgat ctgtctattt
     6541 cgttcatcca tagttgcctg actccccgtc gtgtagataa ctacgatacg ggagggctta
     6601 ccatctggcc ccagtgctgc aatgataccg cgagacccac gctcaccggc tccagattta
     6661 tcagcaataa accagccagc cggaagggcc gagcgcagaa gtggtcctgc aactttatcc
     6721 gcctccatcc agtctattaa ttgttgccgg gaagctagag taagtagttc gccagttaat
     6781 agtttgcgca acgttgttgc cattgctgca ggcatcgtgg tgtcacgctc gtcgtttggt
     6841 atggcttcat tcagctccgg ttcccaacga tcaaggcgag ttacatgatc ccccatgttg
     6901 tgcaaaaaag cggttagctc cttcggtcct ccgatcgttg tcagaagtaa gttggccgca
     6961 gtgttatcac tcatggttat ggcagcactg cataattctc ttactgtcat gccatccgta
     7021 agatgctttt ctgtgactgg tgagtactca accaagt
//
