LOCUS       sequence-446903-        7286 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Mammalian expression of mouse CRT-GPI.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7286)
  AUTHORS   Sen Santara S, Lee DJ, Crespo A, Hu JJ, Walker C, Ma X, Zhang Y, 
            Chowdhury S, Meza-Sosa KF, Lewandrowski M, Zhang H, Rowe M, 
            McClelland A, Wu H, Junqueira C, Lieberman J
  TITLE     The NK cell receptor NKp46 recognizes ecto-calreticulin on 
            ER-stressed cells.
  JOURNAL   Nature. 2023 Apr;616(7956):348-356. doi: 10.1038/s41586-023-05912-0.
            Epub 2023 Apr 5.
  PUBMED    37020026
REFERENCE   2  (bases 1 to 7286)
  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: 
            "10.1038/s41586-023-05912-0"; journalName: "Nature"; date: 
            "2023-04"; volume: "616"; issue: "7956"; pages: "348-356"
COMMENT     Sequence Label: pRetroX-mCRT-GPI-IRES-DsRed
FEATURES             Location/Qualifiers
     source          1..7286
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(506..523)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(506..522)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(515..537)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     complement(731..750)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     850..872
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(910..928)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        996..1100
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(1101..1169,1170..1961)
                     /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(1319..1338)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      2132..2720
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     2621..2640
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     2874..2891
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    3008..3029
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(3044..3049,3050..3067,3068..3074)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3082..3098
                     /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     3087..3109
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     3106..3122
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     3106..3122
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     LTR             3138..3731
                     /note="long terminal repeat from Moloney murine leukemia 
                     virus"
     CDS             complement(3784..4461)
                     /codon_start=1
                     /product="rapidly maturing tetrameric variant of DsRed 
                     fluorescent protein (Bevis and Glick, 2002)"
                     /label=DsRed-Express
                     /note="mammalian codon-optimized"
                     /translation="MASSEDVIKEFMRFKVRMEGSVNGHEFEIEGEGEGRPYEGTQTAK
                     LKVTKGGPLPFAWDILSPQFQYGSKVYVKHPADIPDYKKLSFPEGFKWERVMNFEDGGV
                     VTVTQDSSLQDGSFIYKVKFIGVNFPSDGPVMQKKTMGWEASTERLYPRDGVLKGEIHK
                     ALKLKDGGHYLVEFKSIYMAKKPVQLPGYYYVDSKLDITSHNEDYTIVEQYERAEGRHH
                     LFL"
     primer_bind     complement(3846..3869)
                     /label=DsRed1-C
                     /note="DsRed1, forward primer"
     primer_bind     complement(4044..4063)
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     primer_bind     4261..4281
                     /label=DsRed1-N
                     /note="DsRed1, reverse primer"
     primer_bind     4313..4331
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     misc_feature    4461..5012
                     /label=IRES
                     /note="internal ribosome entry site (IRES) of the 
                     encephalomyocarditis virus (EMCV)"
     misc_feature    4463..5035
                     /label=IRES
                     /note="internal ribosome entry site (IRES) of the 
                     encephalomyocarditis virus (EMCV)"
     primer_bind     4851..4868
                     /label=IRES reverse
                     /note="IRES internal ribosome entry site, reverse primer. 
                     Also called pCDH-rev"
     exon            6472..6501
                     /label=EF-1-alpha exon
                     /note="truncated exon 2 from human EF-1-alpha, beginning 
                     with a GT-to-CC mutation that reduces splicing efficiency"
     intron          6502..6705
                     /label=EF-1-alpha intron A
                     /note="truncated intron upstream of the start codon of 
                     human EF-1-alpha"
     primer_bind     complement(6525..6545)
                     /label=EF1a-F
                     /note="Human elongation factor-1a promoter, forward primer"
     misc_feature    6768..7125
                     /label=MMLV Psi
                     /note="packaging signal of Moloney murine leukemia virus 
                     (MMLV)"
     LTR             join(7190..7286,1..373)
                     /note="long terminal repeat from Moloney murine leukemia 
                     virus"
ORIGIN
        1 ggatacacgg gtacccgggc gactcagtca atcggaggac tggcgcgccg agtgaggggt
       61 tgtgggctct tttattgagc ccggggagca gaagcgcgcg aacagaagcg agaagcgaac
      121 tgattggtta gttcaaataa ggcacagggt catttcaggt ccttggggca ccctggaaac
      181 atctgatggt tctctagaaa ctgctgaggg ctggaccgca tctggggacc atctgttctt
      241 ggccctgagc cggggcagga actgcttacc acagatatcc tgtttggccc atattcagct
      301 gttccatctg ttcttggccc tgagccgggg caggaactgc ttaccacaga tatcctgttt
      361 ggcccatatt cagctgttcc atctgttcct gaccttgatc tgaacttttc tattctcagt
      421 tatgtatttt tccatgcctt gcaaaatggc gttacttaag ctagcttgcc aaacctacag
      481 gtggggtctt tcacataagc ttggcactgg ccgtcgtttt acaacgtcgt gactgggaaa
      541 accctggcgt tacccaactt aatcgccttg cagcacatcc ccctttcgcc agctggcgta
      601 atagcgaaga ggcccgcacc gatcgccctt cccaacagtt gcgcagcctg aatggcgaat
      661 ggcgcctgat gcggtatttt ctccttacgc atctgtgcgg tatttcacac cgcatatggt
      721 gcactctcag tacaatctgc tctgatgccg catagttaag ccagccccga cacccgccaa
      781 cacccgctga cgcgccctga cgggcttgtc tgctcccggc atccgcttac agacaagctg
      841 tgaccgtctc cgggagctgc atgtgtcaga ggttttcacc gtcatcaccg aaacgcgcga
      901 gacgaaaggg cctcgtgata cgcctatttt tataggttaa tgtcatgata ataatggttt
      961 cttagacgtc aggtggcact tttcggggaa atgtgcgcgg aacccctatt tgtttatttt
     1021 tctaaataca ttcaaatatg tatccgctca tgagacaata accctgataa atgcttcaat
     1081 aatattgaaa aaggaagagt atgagtattc aacatttccg tgtcgccctt attccctttt
     1141 ttgcggcatt ttgccttcct gtttttgctc acccagaaac gctggtgaaa gtaaaagatg
     1201 ctgaagatca gttgggtgca cgagtgggtt acatcgaact ggatctcaac agcggtaaga
     1261 tccttgagag ttttcgcccc gaagaacgtt ttccaatgat gagcactttt aaagttctgc
     1321 tatgtggcgc ggtattatcc cgtattgacg ccgggcaaga gcaactcggt cgccgcatac
     1381 actattctca gaatgacttg gttgagtact caccagtcac agaaaagcat cttacggatg
     1441 gcatgacagt aagagaatta tgcagtgctg ccataaccat gagtgataac actgcggcca
     1501 acttacttct gacaacgatc ggaggaccga aggagctaac cgcttttttg cacaacatgg
     1561 gggatcatgt aactcgcctt gatcgttggg aaccggagct gaatgaagcc ataccaaacg
     1621 acgagcgtga caccacgatg cctgtagcaa tggcaacaac gttgcgcaaa ctattaactg
     1681 gcgaactact tactctagct tcccggcaac aattaataga ctggatggag gcggataaag
     1741 ttgcaggacc acttctgcgc tcggcccttc cggctggctg gtttattgct gataaatctg
     1801 gagccggtga gcgtgggtct cgcggtatca ttgcagcact ggggccagat ggtaagccct
     1861 cccgtatcgt agttatctac acgacgggga gtcaggcaac tatggatgaa cgaaatagac
     1921 agatcgctga gataggtgcc tcactgatta agcattggta actgtcagac caagtttact
     1981 catatatact ttagattgat ttaaaacttc atttttaatt taaaaggatc taggtgaaga
     2041 tcctttttga taatctcatg accaaaatcc cttaacgtga gttttcgttc cactgagcgt
     2101 cagaccccgt agaaaagatc aaaggatctt cttgagatcc tttttttctg cgcgtaatct
     2161 gctgcttgca aacaaaaaaa ccaccgctac cagcggtggt ttgtttgccg gatcaagagc
     2221 taccaactct ttttccgaag gtaactggct tcagcagagc gcagatacca aatactgttc
     2281 ttctagtgta gccgtagtta ggccaccact tcaagaactc tgtagcaccg cctacatacc
     2341 tcgctctgct aatcctgtta ccagtggctg ctgccagtgg cgataagtcg tgtcttaccg
     2401 ggttggactc aagacgatag ttaccggata aggcgcagcg gtcgggctga acggggggtt
     2461 cgtgcacaca gcccagcttg gagcgaacga cctacaccga actgagatac ctacagcgtg
     2521 agctatgaga aagcgccacg cttcccgaag ggagaaaggc ggacaggtat ccggtaagcg
     2581 gcagggtcgg aacaggagag cgcacgaggg agcttccagg gggaaacgcc tggtatcttt
     2641 atagtcctgt cgggtttcgc cacctctgac ttgagcgtcg atttttgtga tgctcgtcag
     2701 gggggcggag cctatggaaa aacgccagca acgcggcctt tttacggttc ctggcctttt
     2761 gctggccttt tgctcacatg ttctttcctg cgttatcccc tgattctgtg gataaccgta
     2821 ttaccgcctt tgagtgagct gataccgctc gccgcagccg aacgaccgag cgcagcgagt
     2881 cagtgagcga ggaagcggaa gagcgcccaa tacgcaaacc gcctctcccc gcgcgttggc
     2941 cgattcatta atgcagctgg cacgacaggt ttcccgactg gaaagcgggc agtgagcgca
     3001 acgcaattaa tgtgagttag ctcactcatt aggcacccca ggctttacac tttatgcttc
     3061 cggctcgtat gttgtgtgga attgtgagcg gataacaatt tcacacagga aacagctatg
     3121 accatgatta cgaattcaat gaaagacccc cgctgacggg tagtcaatca ctcagaggag
     3181 accctcccaa ggaacagcga gaccacaagt cggatgcaac tgcaagaggg tttattggat
     3241 acacgggtac ccgggcgact cagtcaatcg gaggactggc gccccgagtg aggggttgtg
     3301 ggctctttta ttgagctcgg ggagcagaag cgcgcgaaca gaagcgagaa gcgaactgat
     3361 tggttagttc aaataaggca cagggtcatt tcaggtcctt ggggcaccct ggaaacatct
     3421 gatggttctc tagaaactgc tgagggctgg accgcatctg gggaccatct gttcttggcc
     3481 ctgagccggg gcaggaactg cttaccacag atatcctgtt tggcccatat tcagctgttc
     3541 catctgttct tggccctgag ccggggcagg aactgcttac cacagatatc ctgtttggcc
     3601 catattcagc tgttccatct gttcctgacc ttgatctgaa cttctctatt ctcagttatg
     3661 tatttttcca tgccttgcaa aatggcgtta cttaagctag cttgccaaac ctacaggtgg
     3721 ggtctttcat tccccccttt ttctggagac taaataaaat cttttatttt atcctcgagc
     3781 attctacagg aacaggtggt ggcggccctc ggcgcgctcg tactgctcca cgatggtgta
     3841 gtcctcgttg tgggaggtga tgtccagctt ggagtccacg tagtagtagc cgggcagctg
     3901 cacgggcttc ttggccatat agatagactt gaactccacc aggtagtggc cgccgtcctt
     3961 cagcttcagg gccttgtgga tctcgccctt cagcacgccg tcgcgggggt acaggcgctc
     4021 ggtggaggcc tcccagccca tagtcttctt ctgcattacg gggccgtcgg aggggaagtt
     4081 cacgccgatg aacttcacct tgtagatgaa ggagccgtcc tgcagggagg agtcctgggt
     4141 cacggtcacc acgccgccgt cctcgaagtt catcacgcgc tcccacttga agccctcggg
     4201 gaaggacagc ttcttgtagt cggggatgtc ggcggggtgc ttcacgtaca ccttggagcc
     4261 gtactggaac tggggggaca ggatgtccca ggcgaagggc agggggccgc ccttggtcac
     4321 cttcagcttg gcggtctggg tgccctcgta ggggcggccc tcgccctcgc cctcgatctc
     4381 gaactcgtgg ccgttcacgg agccctccat gcgcaccttg aagcgcatga actccttgat
     4441 gacgtcctcg gaggaggcca tattatcatc gtgtttttca aaggaaaacc acgtccccgt
     4501 ggttcggggg gcctagacgt tttttaacct cgactaaaca catgtaaagc atgtgcaccg
     4561 aggccccaga tcagatccca tacaatgggg taccttctgg gcatccttca gccccttgtt
     4621 gaatacgctt gaggtgagcc atttgactct ttccacaact atccaactca caacgtggca
     4681 ctggggttgt gccgcctttg caggtgtatc ttatacacgt ggcttttggc cgcagaggca
     4741 cctgtcgcca ggtggggggt tccgctgcct gcaaagggtc gctacagacg ttgtttgtct
     4801 tcaagaagct tccagaggaa ctgcttcctt cacgacattc aacagacctt gcattccttt
     4861 ggcgagaggg gaaagacccc taggaatgct cgtcaagaag acagggccag gtttccgggc
     4921 cctcacattg ccaaaagacg gcaatatggt ggaaaataac atatagacaa acgcacaccg
     4981 gccttattcc aagcggcttc ggccagtaac gttagggggg ggggagggag aggggcacgg
     5041 atccctaggt cagcaggccc atggtcacca gggtgcccag caggccggtc agggtgaagc
     5101 aggtgtggcc gctcagcagc ctggtggtgc cgctggtggt gccgctgccc ttgttagggg
     5161 tggtctcgtg ggcttggcca ggggattctt cctcatcttc ctccttctca tcttcttcat
     5221 cctcatcttc atctctgtca tcatcatcct ctttatcctc agcttcttct tcctctttac
     5281 gcttcttgtc ctcttcttct tccttaagcc tctgctcctc atcctgcttg tccttcatct
     5341 gcttctctgc agccttggta acaccccacg tctcattgcc aaactcctct gcataggcct
     5401 catcattggt gatgaggaaa ttgtcaaaga ttgtcccgga cttgacctgc cagagatcta
     5461 ggcccagtac agcaaaacta tcataggcat agatatttgc atcgggggag tattcagggt
     5521 tgtcaatttc tgggtgtatc caggtaccct tgtaatctgg gttgtcaatt tgacgtggtt
     5581 tccactcgcc cttgtattca ggattttgaa tcactggtgg ttcccactct ccatccatct
     5641 cttcatccca gtcctcaggc ttcttagcat cagggtcagg gatgtgctct ggcttgtccc
     5701 agtcctcagg cttggaatct gtggggtcat cgatcttggc tcgttcatcc cagtcttccg
     5761 gcttggcagc atcagggtcc tttatcttct tgggtggcag aaagtcccaa tcatcctcca
     5821 aggagcctga ctccacctgg ctgttgtcaa ttttcacctc ataggtgttg tctggccgca
     5881 caatcagtgt gtataggtgt gtgaattcat catccttaca ccggatatcc ttgttgatca
     5941 gcacattctt gcccttgtag ttaaagatga catgaacctt cttggtgcca ggaccgcaga
     6001 tgtccggacc aaacatgatg ttatattctg agtctccatg catgtccttc tggtccaagc
     6061 cactcggaaa cagcttcacg tagccgcccc cacagtcgat attctgctca tgcttcaccg
     6121 tgaactgtac caccagtgtc tggcccttat tgctgaaggg ttcgaatttg gcggacagtg
     6181 cgtaaaatcg ggcatcttgg cttgtctgca gccctttatc cttctccagg tccccgtaaa
     6241 atttgccaga actgaggaca aatttgccaa aatcggactt atgtttggat tcgacccagc
     6301 ggttggtcca ggcatctccg tccaagaact gctctttgaa atagatggca gggtctgcgg
     6361 cggccaggcc gaggaggcca agcaggagcg gcaccgaaag gagcatctcg agccgcgggc
     6421 ccacgtgaga tccgagctcg gtaccaagct taagtttggg aattcgattt ttttggcttt
     6481 taggggtagt tttcacgacg gctgaaatgg aagaaaaaaa ctttgaacca ctgtctgagg
     6541 cttgagaatg aaccaagatc caaactcaaa aagggcaaat tccaaggaga attacatcaa
     6601 gtgccaagct ggcctaactt cagtctccac ccactcagtg tggggaaact ccatcgcata
     6661 aaacccctcc ccccaaccta aagacgacgt actccaaaag ctcgacacgc gtattttcag
     6721 acaaatacag aaacacagtc agacagagac aacacagaac gatgctgcag cagacaagac
     6781 gcgcggcgcg gcttcggtcc caaaccgaaa gcaaaaattc agacggaggc gggaactgtt
     6841 ttaggttctc gtctcctacc agaaccacat atccctcctc taaggggggt gcaccaaaga
     6901 gtccaaaacg atcgggattt ttggactcag gtcgggccac aaaaacggcc cccgaagtcc
     6961 ctgggacgtc tcccagggtt gcggccgggt gttccgaact cgtcagttcc accacgggtc
     7021 cgccagatac agagctagtt agctaactag taccgacgca ggcgcataaa atcagtcata
     7081 gacactagac aatcggacag acacagataa gttgctggcc agcttacctc ccggtggtgg
     7141 gtcggtggtc cctgggcagg ggtctcccga tcccggacga gcccccaaat gaaagacccc
     7201 cgctgacggg tagtcaatca ctcagaggag accctcccaa ggaacagcga ggccacaagt
     7261 cggatgcaac tgcaagaggg tttatt
//
