LOCUS       sequence-431290-        5787 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Empty backbone to express gene specific gRNA from the Plasmodium 
            berghei U6 promoter for traditional CRISPR editing..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5787)
  AUTHORS   Jonsdottir TK, Paoletta MS, Ishizaki T, Hernandez S, Ivanova M, 
            Curbelo AH, Saiki PA, Selinger M, Das D, Henriksson J, Bushell ESC
  TITLE     A scalable CRISPR-Cas9 gene editing system facilitates CRISPR 
            screens in the malaria parasite Plasmodium berghei
  JOURNAL   bioRxiv 2024.04.20.590404
REFERENCE   2  (bases 1 to 5787)
  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: "bioRxiv 
            2024.04.20.590404"
COMMENT     Sequence Label: pPbU6_hdhfr/yfcu
FEATURES             Location/Qualifiers
     source          1..5787
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     misc_RNA        11..86
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
     promoter        126..230
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(231..299,300..1091)
                     /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(449..468)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      1262..1850
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     1751..1770
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     2004..2021
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    2138..2159
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(2174..2179,2180..2197,2198..2204)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    2212..2228
                     /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     2217..2239
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     2236..2252
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     2236..2252
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(5056..5078)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
ORIGIN
        1 cctcgtctca gttttagagc tagaaatagc aagttaaaat aaggctagtc cgttatcaac
       61 ttgaaaaagt ggcaccgagt cggtgctttt tttggacgtc aggtggcact tttcggggaa
      121 atgtgcgcgg aacccctatt tgtttatttt tctaaataca ttcaaatatg tatccgctca
      181 tgagacaata accctgataa atgcttcaat aatattgaaa aaggaagagt atgagtattc
      241 aacatttccg tgtcgccctt attccctttt ttgcggcatt ttgccttcct gtttttgctc
      301 acccagaaac gctggtgaaa gtaaaagatg ctgaagatca gttgggtgca cgagtgggtt
      361 acatcgaact ggatctcaac agcggtaaga tccttgagag ttttcgcccc gaagaacgtt
      421 ttccaatgat gagcactttt aaagttctgc tatgtggcgc ggtattatcc cgtattgacg
      481 ccgggcaaga gcaactcggt cgccgcatac actattctca gaatgacttg gttgagtact
      541 caccagtcac agaaaagcat cttacggatg gcatgacagt aagagaatta tgcagtgctg
      601 ccataaccat gagtgataac actgcggcca acttacttct gacaacgatc ggaggaccga
      661 aggagctaac cgcttttttg cacaacatgg gggatcatgt aactcgcctt gatcgttggg
      721 aaccggagct gaatgaagcc ataccaaacg acgagcgtga caccacgatg cctgtagcaa
      781 tggcaacaac gttgcgcaaa ctattaactg gcgaactact tactctagct tcccggcaac
      841 aattaataga ctggatggag gcggataaag ttgcaggacc acttctgcgc tcggcccttc
      901 cggctggctg gtttattgct gataaatctg gagccggtga gcgtgggtct cgcggtatca
      961 ttgcagcact ggggccagat ggtaagccct cccgtatcgt agttatctac acgacgggga
     1021 gtcaggcaac tatggatgaa cgaaatagac agatcgctga gataggtgcc tcactgatta
     1081 agcattggta actgtcagac caagtttact catatatact ttagattgat ttaaaacttc
     1141 atttttaatt taaaaggatc taggtgaaga tcctttttga taatctcatg accaaaatcc
     1201 cttaacgtga gttttcgttc cactgagcgt cagaccccgt agaaaagatc aaaggatctt
     1261 cttgagatcc tttttttctg cgcgtaatct gctgcttgca aacaaaaaaa ccaccgctac
     1321 cagcggtggt ttgtttgccg gatcaagagc taccaactct ttttccgaag gtaactggct
     1381 tcagcagagc gcagatacca aatactgttc ttctagtgta gccgtagtta ggccaccact
     1441 tcaagaactc tgtagcaccg cctacatacc tcgctctgct aatcctgtta ccagtggctg
     1501 ctgccagtgg cgataagtcg tgtcttaccg ggttggactc aagacgatag ttaccggata
     1561 aggcgcagcg gtcgggctga acggggggtt cgtgcacaca gcccagcttg gagcgaacga
     1621 cctacaccga actgagatac ctacagcgtg agctatgaga aagcgccacg cttcccgaag
     1681 ggagaaaggc ggacaggtat ccggtaagcg gcagggtcgg aacaggagag cgcacgaggg
     1741 agcttccagg gggaaacgcc tggtatcttt atagtcctgt cgggtttcgc cacctctgac
     1801 ttgagcgtcg atttttgtga tgctcgtcag gggggcggag cctatggaaa aacgccagca
     1861 acgcggcctt tttacggttc ctggcctttt gctggccttt tgctcacatg ttctttcctg
     1921 cgttatcccc tgattctgtg gataaccgta ttaccgcctt tgagtgagct gataccgctc
     1981 gccgcagccg aacgaccgag cgcagcgagt cagtgagcga ggaagcggaa gagcgcccaa
     2041 tacgcaaacc gcctctcccc gcgcgttggc cgattcatta atgcagctgg cacgacaggt
     2101 ttcccgactg gaaagcgggc agtgagcgca acgcaattaa tgtgagttag ctcactcatt
     2161 aggcacccca ggctttacac tttatgcttc cggctcgtat gttgtgtgga attgtgagcg
     2221 gataacaatt tcacacagga aacagctatg accatgatta cgccaagctt ggtacccatg
     2281 ccatggtccc cgcggccgct cgaggaattc cttaagttta caatttaata ttcatacttt
     2341 aagtattttt tgtagtatcc tagatattgt gctttaaatg ctcacccctc aaagcaccag
     2401 taatattttc atccactgaa ataccattaa attttcaaaa aaatactatg catataatgt
     2461 tatacatata aacataaaac gccatgtaaa tcaaaaaata tataaaaata tgtataaaaa
     2521 taaatatgca ctaaatataa gctaattatg cataaaaatt aaagtgccct ttattaacta
     2581 gtcgtaatta tttatatttc tatgttataa aaaaatcctc atataataat ataattaata
     2641 tatgtaatgt tttttttatt ttataatttt aatataaaat aatatgtaaa ttaattcaaa
     2701 aaataaatat aattgttgtg aaacaaaaaa cgtaattttt tcatttgcct tcaaaattta
     2761 aatttatttt aatatttcct aaaatatata tactttgtgt ataaatatat aaaaatatat
     2821 atttgcttat aaataaataa aaaattttat aaaacatagg gggatctatg gttggttcgc
     2881 taaactgcat cgtcgctgtg tcccagaaca tgggcatcgg caagaacggg gacctgccct
     2941 ggccaccgct caggaacgaa tttagatatt tccagagaat gaccacaacc tcttcagtag
     3001 aaggtaaaca gaatctggtg attatgggta agaagacctg gttctccatt cctgagaaga
     3061 atcgaccttt aaagggtaga attaatttag ttctcagcag agaactcaag gaacctccac
     3121 aaggagctca ttttctttcc agaagtctag atgatgcctt aaaacttact gaacaaccag
     3181 aattagcaaa taaagtagac atggtctgga tagttggtgg cagttctgtt tataaggaag
     3241 ccatgaatca cccaggccat cttaaactat ttgtgacaag gatcatgcaa gactttgaaa
     3301 gtgacacgtt ttttccagaa attgatttgg agaaatataa acttctgcca gaatacccag
     3361 gtgttctctc tgatgtccag gaggagaaag gcattaagta caaatttgaa gtatatgaga
     3421 agaatgatgc aagcggagga ggtggatctg gtggaggtgg aagtgcaagc gtgacagggg
     3481 gaatggctag caagtgggat cagaagggta tggacattgc ctatgaggag gcggccttag
     3541 gttacaagga gggtggtgtt cctattggcg gatgtcttat caataacaaa gacggaagtg
     3601 ttctcggtcg tggtcacaac atgagatttc aaaagggatc cgccacacta catggtgaga
     3661 tctccacttt ggaaaactgt gggagattag agggcaaagt gtacaaagat accactttgt
     3721 atacgacgct gtctccatgc gacatgtgta caggtgccat catcatgtat ggtattccac
     3781 gctgtgttgt cggtgagaac gttaatttca aaagtaaggg cgagaaatat ttacaaacta
     3841 gaggtcacga ggttgttgtt gttgacgatg agaggtgtaa aaagatcatg aaacaattta
     3901 tcgatgaaag acctcaggat tggtttgaag atattggtga ggcttcggaa ccatttaaga
     3961 acgtctactt gctacctcaa acaaaccaat tgctgggttt gtacaccatc atcagaaata
     4021 agaatacaac tagacctgat ttcattttct actccgatag aatcatcaga ttgttggttg
     4081 aagaaggttt gaaccatcta cctgtgcaaa agcaaattgt ggaaactgac accaacgaaa
     4141 acttcgaagg tgtctcattc atgggtaaaa tctgtggtgt ttccattgtc agagctggtg
     4201 aatcgatgga gcaaggatta agagactgtt gtaggtctgt gcgtatcggt aaaattttaa
     4261 ttcaaaggga cgaggagact gctttaccaa agttattcta cgaaaaatta ccagaggata
     4321 tatctgaaag gtatgtcttc ctattagacc caatgctggc caccggtggt agtgctatca
     4381 tggctacaga agtcttgatt aagagaggtg ttaagccaga gagaatttac ttcttaaacc
     4441 taatctgtag taaggaaggg attgaaaaat accatgccgc cttcccagag gtcagaattg
     4501 ttactggtgc cctcgacaga ggtctagatg aaaacaagta tctagttcca gggttgggtg
     4561 actttggtga cagatactac tgtgtttagg gatccgatcc cgtttttctt acttatatat
     4621 ttataccaat tgattgtatt tataactgta aaaatgtgta tgttgtgtgc atattttttt
     4681 tgtgcatgca catgcatgta aatagctaaa attatgaaca ttttattttt tgttcagaaa
     4741 aaaaaaactt tacacacata aaatggctag tatgaatagc catattttat ataaattaaa
     4801 tcctatgaat ttatgaccat attaaaaatt tagatattta tggaacataa tatgtttgaa
     4861 acaataagac aaaattatta ttattattat tatttttact gttataatta tgttgtctct
     4921 tcaatgattc ataaatagtt ggacttgatt tttaaaatgt ttataatatg attagcatag
     4981 ttaaataaaa aaagttgaaa aattaaaaaa aaacatataa acacaaatga tgttttttcc
     5041 ttcaatttcg gccgtcgttt tacaacgtcg tgactgggaa aaccctggcg ttacccaact
     5101 taatcgcctt gcagcacatc cccctttcgc cagctggcgt aatagcgaag aggcccgcac
     5161 cgatcgccct tcccaacagt tgcgcagcct gaatggcgaa tgatcgagaa ctattgtttc
     5221 cttttttgtt tttttatttt taattaaaat gtaaatataa actcacaaaa tgaatgaata
     5281 aataaataaa taaataaata aaaaatgtta aatcgtgtta aaccgcgtgc ttcatgtgat
     5341 atggcaagcg attaatggcc attttatcca tttttataat ttgataaaaa attaaaacaa
     5401 ataaatgtaa tgttaattat attttgtact ataaatatta ttctctactt cacgcaaatg
     5461 taacactatc caactacact cataatgttt aaagaaaatt ggcattgttt taccattttt
     5521 ttaaaacaaa aaaagaataa aatactttcg tattatttag ctaaaattgt actaaatttc
     5581 ataccaaaaa aaaatatata tatgtatgca gctaatattt atttgataag tttttatgta
     5641 aataatattt aattaaaaaa aaaattacat agcatgccga atgcactatt cattttatgg
     5701 gggtaatttt ttgagtaacc atttttactt tttttttaca tatgcgcata cttcgagtta
     5761 tacaatatta ttggagacgc gtagagg
//
