LOCUS       sequence-443042-        5674 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses CD55 variant in mammalian cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5674)
  TITLE     CRISPR/Cas9 screening reveals a distinct class of pMHC-I binders 
            with precise HLA-peptide recognition
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 5674)
  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: 
            "Unpublished"
COMMENT     Sequence Label: CD55-Delta4
FEATURES             Location/Qualifiers
     source          1..5674
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        64..267
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     218..238
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     misc_feature    join(724..1298,1299..1301,1302..1310)
                     /label=IRES2
                     /note="internal ribosome entry site (IRES) of the 
                     encephalomyocarditis virus (EMCV)"
     primer_bind     complement(891..908)
                     /label=IRES reverse
                     /note="IRES internal ribosome entry site, reverse primer. 
                     Also called pCDH-rev"
     primer_bind     1118..1137
                     /label=IRES-F
                     /note="IRES internal ribosome entry site, forward primer"
     regulatory      1314..1323
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(1320..1322,1323..1325,1326..2039)
                     /codon_start=1
                     /product="enhanced GFP with monomerizing A206K mutation 
                     (Zacharias et al., 2002)"
                     /label=mEGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSKLSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(1365..1386)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(1626..1645)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1973..1994
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     polyA_signal    2161..2282
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2198..2217)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     2252..2271
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(2289..2744)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2426..2447)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     2638..2657
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        2771..2875
                     /gene="bla"
                     /label=AmpR promoter
     promoter        2877..3234
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        2877..3068
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(2900..2920)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      3085..3220
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             3269..4063
                     /codon_start=1
                     /product="aminoglycoside phosphotransferase"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     CDS             3269..4063
                     /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="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(3323..3342)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     3933..3952
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(4251..4270)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    4295..4342
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      4671..5259
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5160..5179
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     enhancer        join(5434..5674,1..63)
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
ORIGIN
        1 atgaccttat gggactttcc tacttggcag tacatctacg tattagtcat cgctattacc
       61 atggtgatgc ggttttggca gtacatcaat gggcgtggat agcggtttga ctcacgggga
      121 tttccaagtc tccaccccat tgacgtcaat gggagtttgt tttggcacca aaatcaacgg
      181 gactttccaa aatgtcgtaa caactccgcc ccattgacgc aaatgggcgg taggcgtgta
      241 cggtgggagg tctatataag cagagctggt ttagtgaacc gtcagatccg ctagcgctac
      301 cggactcaga tctcgaggcc accatgaccg tcgcgcggcc gagcgtgccc gcggcgctgc
      361 ccctcctcgg ggagctgccc cggctgctgc tgctggtgct gttgtgcctg ccggccgtgt
      421 ggggtaaatc tctaacttcc aaggtcccac caacagttca gaaacctacc acagtaaatg
      481 ttccaactac agaagtctca ccaacttctc agaaaaccac cacaaaaacc accacaccaa
      541 atgctcaagc aacacggagt acacctgttt ccaggacaac caagcatttt catgaaacaa
      601 ccccaaataa aggaagtgga accacttcag gtactacccg tcttctatct gggcacacgt
      661 gtttcacgtt gacaggtttg cttgggacgc tagtaaccat gggcttgctg acttagggat
      721 ccgcccctct ccctcccccc cccctaacgt tactggccga agccgcttgg aataaggccg
      781 gtgtgcgttt gtctatatgt tattttccac catattgccg tcttttggca atgtgagggc
      841 ccggaaacct ggccctgtct tcttgacgag cattcctagg ggtctttccc ctctcgccaa
      901 aggaatgcaa ggtctgttga atgtcgtgaa ggaagcagtt cctctggaag cttcttgaag
      961 acaaacaacg tctgtagcga ccctttgcag gcagcggaac cccccacctg gcgacaggtg
     1021 cctctgcggc caaaagccac gtgtataaga tacacctgca aaggcggcac aaccccagtg
     1081 ccacgttgtg agttggatag ttgtggaaag agtcaaatgg ctctcctcaa gcgtattcaa
     1141 caaggggctg aaggatgccc agaaggtacc ccattgtatg ggatctgatc tggggcctcg
     1201 gtacacatgc tttacatgtg tttagtcgag gttaaaaaaa cgtctaggcc ccccgaacca
     1261 cggggacgtg gttttccttt gaaaaacacg atgataatat ggccacaacc atggccacca
     1321 tggtgagcaa gggcgaggag ctgttcaccg gggtggtgcc catcctggtc gagctggacg
     1381 gcgacgtaaa cggccacaag ttcagcgtgt ccggcgaggg cgagggcgat gccacctacg
     1441 gcaagctgac cctgaagttc atctgcacca ccggcaagct gcccgtgccc tggcccaccc
     1501 tcgtgaccac cctgacctac ggcgtgcagt gcttcagccg ctaccccgac cacatgaagc
     1561 agcacgactt cttcaagtcc gccatgcccg aaggctacgt ccaggagcgc accatcttct
     1621 tcaaggacga cggcaactac aagacccgcg ccgaggtgaa gttcgagggc gacaccctgg
     1681 tgaaccgcat cgagctgaag ggcatcgact tcaaggagga cggcaacatc ctggggcaca
     1741 agctggagta caactacaac agccacaacg tctatatcat ggccgacaag cagaagaacg
     1801 gcatcaaggt gaacttcaag atccgccaca acatcgagga cggcagcgtg cagctcgccg
     1861 accactacca gcagaacacc cccatcggcg acggccccgt gctgctgccc gacaaccact
     1921 acctgagcac ccagtccaag ctgagcaaag accccaacga gaagcgcgat cacatggtcc
     1981 tgctggagtt cgtgaccgcc gccgggatca ctctcggcat ggacgagctg tacaagtaag
     2041 cggccgcgac tctagatcat aatcagccat accacatttg tagaggtttt acttgcttta
     2101 aaaaacctcc cacacctccc cctgaacctg aaacataaaa tgaatgcaat tgttgttgtt
     2161 aacttgttta ttgcagctta taatggttac aaataaagca atagcatcac aaatttcaca
     2221 aataaagcat ttttttcact gcattctagt tgtggtttgt ccaaactcat caatgtatct
     2281 taaggcgtaa attgtaagcg ttaatatttt gttaaaattc gcgttaaatt tttgttaaat
     2341 cagctcattt tttaaccaat aggccgaaat cggcaaaatc ccttataaat caaaagaata
     2401 gaccgagata gggttgagtg ttgttccagt ttggaacaag agtccactat taaagaacgt
     2461 ggactccaac gtcaaagggc gaaaaaccgt ctatcagggc gatggcccac tacgtgaacc
     2521 atcaccctaa tcaagttttt tggggtcgag gtgccgtaaa gcactaaatc ggaaccctaa
     2581 agggagcccc cgatttagag cttgacgggg aaagccggcg aacgtggcga gaaaggaagg
     2641 gaagaaagcg aaaggagcgg gcgctagggc gctggcaagt gtagcggtca cgctgcgcgt
     2701 aaccaccaca cccgccgcgc ttaatgcgcc gctacagggc gcgtcaggtg gcacttttcg
     2761 gggaaatgtg cgcggaaccc ctatttgttt atttttctaa atacattcaa atatgtatcc
     2821 gctcatgaga caataaccct gataaatgct tcaataatat tgaaaaagga agagtcctga
     2881 ggcggaaaga accagctgtg gaatgtgtgt cagttagggt gtggaaagtc cccaggctcc
     2941 ccagcaggca gaagtatgca aagcatgcat ctcaattagt cagcaaccag gtgtggaaag
     3001 tccccaggct ccccagcagg cagaagtatg caaagcatgc atctcaatta gtcagcaacc
     3061 atagtcccgc ccctaactcc gcccatcccg cccctaactc cgcccagttc cgcccattct
     3121 ccgccccatg gctgactaat tttttttatt tatgcagagg ccgaggccgc ctcggcctct
     3181 gagctattcc agaagtagtg aggaggcttt tttggaggcc taggcttttg caaagatcga
     3241 tcaagagaca ggatgaggat cgtttcgcat gattgaacaa gatggattgc acgcaggttc
     3301 tccggccgct tgggtggaga ggctattcgg ctatgactgg gcacaacaga caatcggctg
     3361 ctctgatgcc gccgtgttcc ggctgtcagc gcaggggcgc ccggttcttt ttgtcaagac
     3421 cgacctgtcc ggtgccctga atgaactgca agacgaggca gcgcggctat cgtggctggc
     3481 cacgacgggc gttccttgcg cagctgtgct cgacgttgtc actgaagcgg gaagggactg
     3541 gctgctattg ggcgaagtgc cggggcagga tctcctgtca tctcaccttg ctcctgccga
     3601 gaaagtatcc atcatggctg atgcaatgcg gcggctgcat acgcttgatc cggctacctg
     3661 cccattcgac caccaagcga aacatcgcat cgagcgagca cgtactcgga tggaagccgg
     3721 tcttgtcgat caggatgatc tggacgaaga gcatcagggg ctcgcgccag ccgaactgtt
     3781 cgccaggctc aaggcgagca tgcccgacgg cgaggatctc gtcgtgaccc atggcgatgc
     3841 ctgcttgccg aatatcatgg tggaaaatgg ccgcttttct ggattcatcg actgtggccg
     3901 gctgggtgtg gcggaccgct atcaggacat agcgttggct acccgtgata ttgctgaaga
     3961 gcttggcggc gaatgggctg accgcttcct cgtgctttac ggtatcgccg ctcccgattc
     4021 gcagcgcatc gccttctatc gccttcttga cgagttcttc tgagcgggac tctggggttc
     4081 gaaatgaccg accaagcgac gcccaacctg ccatcacgag atttcgattc caccgccgcc
     4141 ttctatgaaa ggttgggctt cggaatcgtt ttccgggacg ccggctggat gatcctccag
     4201 cgcggggatc tcatgctgga gttcttcgcc caccctaggg ggaggctaac tgaaacacgg
     4261 aaggagacaa taccggaagg aacccgcgct atgacggcaa taaaaagaca gaataaaacg
     4321 cacggtgttg ggtcgtttgt tcataaacgc ggggttcggt cccagggctg gcactctgtc
     4381 gataccccac cgagacccca ttggggccaa tacgcccgcg tttcttcctt ttccccaccc
     4441 caccccccaa gttcgggtga aggcccaggg ctcgcagcca acgtcggggc ggcaggccct
     4501 gccatagcct caggttactc atatatactt tagattgatt taaaacttca tttttaattt
     4561 aaaaggatct aggtgaagat cctttttgat aatctcatga ccaaaatccc ttaacgtgag
     4621 ttttcgttcc actgagcgtc agaccccgta gaaaagatca aaggatcttc ttgagatcct
     4681 ttttttctgc gcgtaatctg ctgcttgcaa acaaaaaaac caccgctacc agcggtggtt
     4741 tgtttgccgg atcaagagct accaactctt tttccgaagg taactggctt cagcagagcg
     4801 cagataccaa atactgttct tctagtgtag ccgtagttag gccaccactt caagaactct
     4861 gtagcaccgc ctacatacct cgctctgcta atcctgttac cagtggctgc tgccagtggc
     4921 gataagtcgt gtcttaccgg gttggactca agacgatagt taccggataa ggcgcagcgg
     4981 tcgggctgaa cggggggttc gtgcacacag cccagcttgg agcgaacgac ctacaccgaa
     5041 ctgagatacc tacagcgtga gctatgagaa agcgccacgc ttcccgaagg gagaaaggcg
     5101 gacaggtatc cggtaagcgg cagggtcgga acaggagagc gcacgaggga gcttccaggg
     5161 ggaaacgcct ggtatcttta tagtcctgtc gggtttcgcc acctctgact tgagcgtcga
     5221 tttttgtgat gctcgtcagg ggggcggagc ctatggaaaa acgccagcaa cgcggccttt
     5281 ttacggttcc tggccttttg ctggcctttt gctcacatgt tctttcctgc gttatcccct
     5341 gattctgtgg ataaccgtat taccgccatg cattagttat taatagtaat caattacggg
     5401 gtcattagtt catagcccat atatggagtt ccgcgttaca taacttacgg taaatggccc
     5461 gcctggctga ccgcccaacg acccccgccc attgacgtca ataatgacgt atgttcccat
     5521 agtaacgcca atagggactt tccattgacg tcaatgggtg gagtatttac ggtaaactgc
     5581 ccacttggca gtacatcaag tgtatcatat gccaagtacg ccccctattg acgtcaatga
     5641 cggtaaatgg cccgcctggc attatgccca gtac
//
