LOCUS       sequence-181632-        4690 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses sgRNA for targeting CrPEPC1 gene, paromomycin resistance.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4690)
  AUTHORS   Kao PH, Ng IS
  TITLE     CRISPRi mediated phosphoenolpyruvate carboxylase regulation to 
            enhance the production of lipid in Chlamydomonas reinhardtii.
  JOURNAL   Bioresour Technol. 2017 May 4. pii: S0960-8524(17)30619-3. doi: 
            10.1016/j.biortech.2017.04.111.
  PUBMED    28501380
REFERENCE   2  (bases 1 to 4690)
  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: "Bioresour 
            Technol. 2017 May 4. pii: S0960-8524(17)30619-3. doi: 
            10.1016/j.biortech.2017.04.111."
COMMENT     Sequence Label: pIKU801-CrPEPC1
FEATURES             Location/Qualifiers
     source          1..4690
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     misc_RNA        21..96
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
     promoter        complement(2130..2148)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(2169..2185)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2169..2185)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(2182..2204)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    2193..2209
                     /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)."
     promoter        complement(join(2217..2223,2224..2241,2242..2247))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    2262..2283
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(2400..2417)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(2571..3159)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2651..2670)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(3330..4121,4122..4190))
                     /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     3953..3972
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
ORIGIN
        1 ggacgcggtg accacctacc gttttagagc tagaaatagc aagttaaaat aaggctagtc
       61 cgttatcaac ttgaaaaagt ggcaccgagt cggtgctttt ttttgcaaaa ttttccagat
      121 cgatttcttc ttcctctgtt cttcggcgtt caatttctgg ggttttctct tcgttttctg
      181 taactgaaac ctaaaatttg acctaaaaaa aatctcaaat aatatgattc agtggttttg
      241 tacttttcag ttagttgagt tttgcagttc cgatgagata aaccaataga attcacgcgt
      301 catatggagc tcgctgaggc ttgacatgat tggtgcgtat gtttgtatga agctacagga
      361 ctgatttggc gggctatgag ggcgggggaa gctctggaag ggccgcgatg gggcgcgcgg
      421 cgtccagaag gcgccatacg gcccgctggc ggcacccatc cggtataaaa gcccgcgacc
      481 ccgaacggtg acctccactt tcagcgacaa acgagcactt atacatacgc gactattctg
      541 ccgctataca taaccactca gctagcttaa gatcccatca agcttgcatg ccgggcgcgc
      601 cagaaggagc gcagccaaac caggatgatg tttgatgggg tatttgagca cttgcaaccc
      661 ttatccggaa gccccctggc ccacaaaggc taggcgccaa tgcaagcagt tcgcatgcag
      721 cccctggagc ggtgccctcc tgataaaccg gccagggggc ctatgttctt tactttttta
      781 caagagaagt cactcaacat cttaaaatgg ccaggtgagt cgacgagcaa gcccggcgga
      841 tcaggcagcg tgcttgcaga tttgacttgc aacgcccgca ttgtgtcgac gaaggctttt
      901 ggctcctctg tcgctgtctc aagcagcatc taaccctgcg tcgccgtttc catttgcagg
      961 atggccactc cgccctcccc ggtgctgaag aatttcgaag catggacgat gcgttgcgtg
     1021 cactgcgggg tcggtatccc ggttgtgagt gggttgttgt ggaggatggg gcctcggggg
     1081 ctggtgttta tcggcttcgg ggtggtgggc gggagttgtt tgtcaaggtg gcagctctgg
     1141 gggccggggt gggcttgttg ggtgaggctg agcggctggt gtggttggcg gaggtgggga
     1201 ttcccgtacc tcgtgttgtg gagggtggtg gggacgagag ggtcgcctgg ttggtcaccg
     1261 aagcggttcc ggggcgtccg gccagtgcgc ggtggccgcg ggagcagcgg ctggacgtgg
     1321 cggtggcgct cgcggggctc gctcgttcgc tgcacgcgct ggactgggag cggtgtccgt
     1381 tcgatcgcag tctcgcggtg acggtgccgc aggcggcccg tgctgtcgct gaagggagcg
     1441 tcgacttgga ggatctggac gaggagcgga aggggtggtc gggggagcgg cttctcgccg
     1501 agctggagcg gactcggcct gcggacgagg atctggcggt ttgccacggt gacctgtgcc
     1561 cggacaacgt gctgctcgac cctcgtacct gcgaggtgac cgggctgatc gacgtggggc
     1621 gggtcggccg tgcggaccgg cactccgatc tcgcgctggt gctgcgcgag ctggcccacg
     1681 aggaggaccc gtggttcggg ccggagtgtt ccgcggcgtt cctgcgggag tacgggcgcg
     1741 ggtgggatgg ggcggtatcg gaggaaaagc tggcgtttta ccggctgttg gacgagttct
     1801 tctgagggac ctgatggtgt tggtggctgg gtagggttgc gtcgcgtggg tgacagcaca
     1861 gtgtggacgt tgggatcccc gctccgtgta aatggaggcg ctcgttgatc tgagccttgc
     1921 cccctgacga acggcggtgg atggaagata ctgctctcaa gtgctgaagc ggtagcttag
     1981 ctccccgttt cgtgctgatc agtctttttc aacacgtaaa aagcggagga gttttgcaat
     2041 tttgttggtt gtaacgatcc tccgttgatt ttggcctctt tctccatggg cgggctgggc
     2101 gtatttgaag cgggtaccag cttttgttcc ctttagtgag ggttaatttc gagcttggcg
     2161 taatcatggt catagctgtt tcctgtgtga aattgttatc cgctcacaat tccacacaac
     2221 atacgagccg gaagcataaa gtgtaaagcc tggggtgcct aatgagtgag ctaactcaca
     2281 ttaattgcgt tgcgctcact gcccgctttc cagtcgggaa acctgtcgtg ccagctgcat
     2341 taatgaatcg gccaacgcgc ggggagaggc ggtttgcgta ttgggcgctc ttccgcttcc
     2401 tcgctcactg actcgctgcg ctcggtcgtt cggctgcggc gagcggtatc agctcactca
     2461 aaggcggtaa tacggttatc cacagaatca ggggataacg caggaaagaa catgtgagca
     2521 aaaggccagc aaaaggccag gaaccgtaaa aaggccgcgt tgctggcgtt tttccatagg
     2581 ctccgccccc ctgacgagca tcacaaaaat cgacgctcaa gtcagaggtg gcgaaacccg
     2641 acaggactat aaagatacca ggcgtttccc cctggaagct ccctcgtgcg ctctcctgtt
     2701 ccgaccctgc cgcttaccgg atacctgtcc gcctttctcc cttcgggaag cgtggcgctt
     2761 tctcatagct cacgctgtag gtatctcagt tcggtgtagg tcgttcgctc caagctgggc
     2821 tgtgtgcacg aaccccccgt tcagcccgac cgctgcgcct tatccggtaa ctatcgtctt
     2881 gagtccaacc cggtaagaca cgacttatcg ccactggcag cagccactgg taacaggatt
     2941 agcagagcga ggtatgtagg cggtgctaca gagttcttga agtggtggcc taactacggc
     3001 tacactagaa gaacagtatt tggtatctgc gctctgctga agccagttac cttcggaaaa
     3061 agagttggta gctcttgatc cggcaaacaa accaccgctg gtagcggtgg tttttttgtt
     3121 tgcaagcagc agattacgcg cagaaaaaaa ggatctcaag aagatccttt gatcttttct
     3181 acggggtctg acgctcagtg gaacgaaaac tcacgttaag ggattttggt catgagatta
     3241 tcaaaaagga tcttcaccta gatcctttta aattaaaaat gaagttttaa atcaatctaa
     3301 agtatatatg agtaaacttg gtctgacagt taccaatgct taatcagtga ggcacctatc
     3361 tcagcgatct gtctatttcg ttcatccata gttgcctgac tccccgtcgt gtagataact
     3421 acgatacggg agggcttacc atctggcccc agtgctgcaa tgataccgcg agacccacgc
     3481 tcaccggctc cagatttatc agcaataaac cagccagccg gaagggccga gcgcagaagt
     3541 ggtcctgcaa ctttatccgc ctccatccag tctattaatt gttgccggga agctagagta
     3601 agtagttcgc cagttaatag tttgcgcaac gttgttgcca ttgctacagg catcgtggtg
     3661 tcacgctcgt cgtttggtat ggcttcattc agctccggtt cccaacgatc aaggcgagtt
     3721 acatgatccc ccatgttgtg caaaaaagcg gttagctcct tcggtcctcc gatcgttgtc
     3781 agaagtaagt tggccgcagt gttatcactc atggttatgg cagcactgca taattctctt
     3841 actgtcatgc catccgtaag atgcttttct gtgactggtg agtactcaac caagtcattc
     3901 tgagaatagt gtatgcggcg accgagttgc tcttgcccgg cgtcaatacg ggataatacc
     3961 gcgccacata gcagaacttt aaaagtgctc atcattggaa aacgttcttc ggggcgaaaa
     4021 ctctcaagga tcttaccgct gttgagatcc agttcgatgt aacccactcg tgcacccaac
     4081 tgatcttcag catcttttac tttcaccagc gtttctgggt gagcaaaaac aggaaggcaa
     4141 aatgccgcaa aaaagggaat aagggcgaca cggaaatgtt gaatactcat actcttcctt
     4201 tttcaatatt attgaagcat ttatcagggt tattgtctca tgagcggctg cagagatctc
     4261 tcgagcgact tgccttccgc acaatacatc atttcttctt agcttttttt cttcttcttc
     4321 gttcatacag tttttttttg tttatcagct tacattttct tgaaccgtag ctttcgtttt
     4381 cttcttttta actttccatt cggagttttt gtatcttgtt tcatagtttg tcccaggatt
     4441 agaatgatta ggcatcgaac cttcaagaat ttgattgaat aaaacatctt cattcttaag
     4501 atatgaagat aatcttcaaa aggcccctgg gaatctgaaa gaagagaagc aggcccattt
     4561 atatgggaaa gaacaatagt atttcttata taggcccatt taagttgaaa acaatcttca
     4621 aaagtcccac atcgcttaga taagaaaacg aagctgagtt tatatacagc tagagtcgaa
     4681 gtagtgattg
//
