LOCUS       sequence-488329-        7568 bp    DNA     circular SYN 03-OCT-2025
DEFINITION  Mammalian expression of myc-tagged full-length SHP2.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7568)
  AUTHORS   van Vlimmeren AE, Giglio RM, Jiang Z, Lee M, McFaline-Figueroa JL, 
            Shah NH
  TITLE     Multiplexed single-cell transcriptomics reveals diverse phenotypic 
            outcomes for pathogenic SHP2 variants.
  JOURNAL   bioRxiv [Preprint]. 2025 Jul 2:2025.06.30.662374. doi: 
            10.1101/2025.06.30.662374.
  PUBMED    40631144
REFERENCE   2  (bases 1 to 7568)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Oct 3, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "bioRxiv 
            [Preprint]. 2025 Jul 2:2025.06.30.662374. doi: 
            10.1101/2025.06.30.662374."
COMMENT     Sequence Label: pEF myc-SHP2(E139D)
FEATURES             Location/Qualifiers
     source          1..7568
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(44..63)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     promoter        351..1529
                     /label=EF-1-alpha promoter
                     /note="strong constitutive promoter for human elongation 
                     factor EF-1-alpha"
     intron          582..1520
                     /label=EF-1-alpha intron A
                     /note="intron upstream of the start codon of human 
                     EF-1-alpha"
     primer_bind     1477..1497
                     /label=EF1a-F
                     /note="Human elongation factor-1a promoter, forward primer"
     primer_bind     1546..1565
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        1546..1564
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             1686..1715
                     /codon_start=1
                     /product="Myc (human c-Myc proto-oncogene) epitope tag"
                     /label=Myc
                     /translation="EQKLISEEDL"
     primer_bind     complement(3535..3552)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    3541..3765
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      3811..4239
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3898..3917)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4108..4129
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(4248..4268)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        4253..4582
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      4433..4568
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     4495..4514
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     promoter        4630..4677
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             4696..5094
                     /codon_start=1
                     /gene="Aspergillus terreus BSD"
                     /product="blasticidin S deaminase"
                     /label=BSD
                     /note="confers resistance to blasticidin"
                     /translation="MAKPLSQEESTLIERATATINSIPISEDYSVASAALSSDGRIFTG
                     VNVYHFTGGPCAELVVLGTAAAAAAGNLTCIVAIGNENRGILSPCGRCRQVLLDLHPGI
                     KAIVKDSDGQPTAVGIRELLPSGYVWEG"
     polyA_signal    5252..5373
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5289..5308)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     5343..5362
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(5422..5438)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5422..5438)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(5435..5457)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5446..5462
                     /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(5470..5476,5477..5494,5495..5500))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5515..5536
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5653..5670)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5813..6401)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5893..5912)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6572..7363,7364..7432))
                     /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     7195..7214
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7433..7537)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 gacggatcgg gagatctccc gatcccctat ggtgcactct cagtacaatc tgctctgatg
       61 ccgcatagtt aagccagtat ctgctccctg cttgtgtgtt ggaggtcgct gagtagtgcg
      121 cgagcaaaat ttaagctaca acaaggcaag gcttgaccga caattgcatg aagaatctgc
      181 ttagggttag gcgttttgcg ctgcttcgcg atgtacgggc cagatatacg cgttgacatt
      241 gattattgac taggcttttg caaaaagctt tgcaaagatg gataaagttt taaacagaga
      301 ggaatctttg cagctaatgg accttctagg tcttgaaagg agtgggaatt ggctccggtg
      361 cccgtcagtg ggcagagcgc acatcgccca cagtccccga gaagttgggg ggaggggtcg
      421 gcaattgaac cggtgcctag agaaggtggc gcggggtaaa ctgggaaagt gatgtcgtgt
      481 actggctccg cctttttccc gagggtgggg gagaaccgta tataagtgca gtagtcgccg
      541 tgaacgttct ttttcgcaac gggtttgccg ccagaacaca ggtaagtgcc gtgtgtggtt
      601 cccgcgggcc tggcctcttt acgggttatg gcccttgcgt gccttgaatt acttccacct
      661 ggctgcagta cgtgattctt gatcccgagc ttcgggttgg aagtgggtgg gagagttcga
      721 ggccttgcgc ttaaggagcc ccttcgcctc gtgcttgagt tgaggcctgg cctgggcgct
      781 ggggccgccg cgtgcgaatc tggtggcacc ttcgcgcctg tctcgctgct ttcgataagt
      841 ctctagccat ttaaaatttt tgatgacctg ctgcgacgct ttttttctgg caagatagtc
      901 ttgtaaatgc gggccaagat ctgcacactg gtatttcggt ttttggggcc gcgggcggcg
      961 acggggcccg tgcgtcccag cgcacatgtt cggcgaggcg gggcctgcga gcgcggccac
     1021 cgagaatcgg acgggggtag tctcaagctg gccggcctgc tctggtgcct ggcctcgcgc
     1081 cgccgtgtat cgccccgccc tgggcggcaa ggctggcccg gtcggcacca gttgcgtgag
     1141 cggaaagatg gccgcttccc ggccctgctg cagggagctc aaaatggagg acgcggcgct
     1201 cgggagagcg ggcgggtgag tcacccacac aaaggaaaag ggcctttccg tcctcagccg
     1261 tcgcttcatg tgactccacg gagtaccggg cgccgtccag gcacctcgat tagttctcga
     1321 gcttttggag tacgtcgtct ttaggttggg gggaggggtt ttatgcgatg gagtttcccc
     1381 acactgagtg ggtggagact gaagttaggc cagcttggca cttgatgtaa ttctccttgg
     1441 aatttgccct ttttgagttt ggatcttggt tcattctcaa gcctcagaca gtggttcaaa
     1501 gtttttttct tccatttcag gtgtcgtgag gaattagctt ggtactaata cgactcacta
     1561 tagggagacc caagctggct aggtaagctt ggtaccgagc tcggatccac tagtccagtg
     1621 tggtggaatt ctgcagatat ccagcacagt ggcggccgct cgagtctaga gggcccttca
     1681 ccatggaaca aaaactcatc tcagaagagg atctgggagg gcagtctggg cagatgacat
     1741 cgcggagatg gtttcaccca aatatcactg gtgtggaggc agaaaaccta ctgttgacaa
     1801 gaggagttga tggcagtttt ttggcaaggc ctagtaaaag taaccctgga gacttcacac
     1861 tttccgttag aagaaatgga gctgtcaccc acatcaagat tcagaacact ggtgattact
     1921 atgacctgta tggaggggag aaatttgcca ctttggctga gttggtccag tattacatgg
     1981 aacatcacgg gcaattaaaa gagaagaatg gagatgtcat tgagcttaaa tatcctctga
     2041 actgtgcaga tcctacctct gaaaggtggt ttcatggaca tctctctggg aaagaagcag
     2101 agaaattatt aactgaaaaa ggaaaacatg gtagttttct tgtacgagat agccagagcc
     2161 accctggaga ttttgttctt tctgtgcgca ctggtgatga caaaggggag agcaatgacg
     2221 gcaagtctaa agtgacccat gttatgattc gctgtcagga actgaaatac gacgttggtg
     2281 gaggagaacg gtttgattct ttgacagatc ttgtggaaca ttataagaag aatcctatgg
     2341 tggaaacatt gggtacagta ctacaactca agcagcccct taacacgact cgtataaatg
     2401 ctgctgaaat agaaagcaga gttcgagaac taagcaaatt agctgagacc acagataaag
     2461 tcaaacaagg cttttgggaa gaatttgaga cactacaaca acaggagtgc aaacttctct
     2521 acagccgaaa agagggtcaa aggcaagaaa acaaaaacaa aaatagatat aaaaacatcc
     2581 tgccctttga tcataccagg gttgtcctac acgatggtga tcccaatgag cctgtttcag
     2641 attacatcaa tgcaaatatc atcatgcctg aatttgaaac caagtgcaac aattcaaagc
     2701 ccaaaaagag ttacattgcc acacaaggct gcctgcaaaa cacggtgaat gacttttggc
     2761 ggatggtgtt ccaagaaaac tcccgagtga ttgtcatgac aacgaaagaa gtggagagag
     2821 gaaagagtaa atgtgtcaaa tactggcctg atgagtatgc tctaaaagaa tatggcgtca
     2881 tgcgtgttag gaacgtcaaa gaaagcgccg ctcatgacta tacgctaaga gaacttaaac
     2941 tttcaaaggt tggacaaggg aatacggaga gaacggtctg gcaataccac tttcggacct
     3001 ggccggacca cggcgtgccc agcgaccctg ggggcgtgct ggacttcctg gaggaggtgc
     3061 accataagca ggagagcatc atggatgcag ggccggtcgt ggtgcactgc agtgctggaa
     3121 ttggccggac agggacgttc attgtgattg atattcttat tgacatcatc agagagaaag
     3181 gtgttgactg cgatattgac gttcccaaaa ccatccagat ggtgcggtct cagaggtcag
     3241 ggatggtcca gacagaagca cagtaccgat ttatctatat ggcggtccag cattatattg
     3301 aaacactaca gcgcaggatt gaagaagagc agaaaagcaa gaggaaaggg cacgaatata
     3361 caaatattaa gtattctcta gcggaccaga cgagtggaga tcagagccct ctcccgcctt
     3421 gtactccaac gccaccctgt gcagaaatga gagaagacag tgctagagtc tatgaaaacg
     3481 tgggcctgat gcaacagcag aaaagtttca gataggttta aacccgctga tcagcctcga
     3541 ctgtgccttc tagttgccag ccatctgttg tttgcccctc ccccgtgcct tccttgaccc
     3601 tggaaggtgc cactcccact gtcctttcct aataaaatga ggaaattgca tcgcattgtc
     3661 tgagtaggtg tcattctatt ctggggggtg gggtggggca ggacagcaag ggggaggatt
     3721 gggaagacaa tagcaggcat gctggggatg cggtgggctc tatggcttct gaggcggaaa
     3781 gaaccagctg gggctctagg gggtatcccc acgcgccctg tagcggcgca ttaagcgcgg
     3841 cgggtgtggt ggttacgcgc agcgtgaccg ctacacttgc cagcgcccta gcgcccgctc
     3901 ctttcgcttt cttcccttcc tttctcgcca cgttcgccgg ctttccccgt caagctctaa
     3961 atcgggggct ccctttaggg ttccgattta gtgctttacg gcacctcgac cccaaaaaac
     4021 ttgattaggg tgatggttca cgtagtgggc catcgccctg atagacggtt tttcgccctt
     4081 tgacgttgga gtccacgttc tttaatagtg gactcttgtt ccaaactgga acaacactca
     4141 accctatctc ggtctattct tttgatttat aagggatttt gccgatttcg gcctattggt
     4201 taaaaaatga gctgatttaa caaaaattta acgcgaatta attctgtgga atgtgtgtca
     4261 gttagggtgt ggaaagtccc caggctcccc agcaggcaga agtatgcaaa gcatgcatct
     4321 caattagtca gcaaccaggt gtggaaagtc cccaggctcc ccagcaggca gaagtatgca
     4381 aagcatgcat ctcaattagt cagcaaccat agtcccgccc ctaactccgc ccatcccgcc
     4441 cctaactccg cccagttccg cccattctcc gccccatggc tgactaattt tttttattta
     4501 tgcagaggcc gaggccgcct ctgcctctga gctattccag aagtagtgag gaggcttttt
     4561 tggaggccta ggcttttgca aaaagctccc gggagcttgt atatccattt tcggatctga
     4621 tcagcacgtg ttgacaatta atcatcggca tagtatatcg gcatagtata atacgacaag
     4681 gtgaggaact aaaccatggc caagcctttg tctcaagaag aatccaccct cattgaaaga
     4741 gcaacggcta caatcaacag catccccatc tctgaagact acagcgtcgc cagcgcagct
     4801 ctctctagcg acggccgcat cttcactggt gtcaatgtat atcattttac tgggggacct
     4861 tgtgcagaac tcgtggtgct gggcactgct gctgctgcgg cagctggcaa cctgacttgt
     4921 atcgtcgcga tcggaaatga gaacaggggc atcttgagcc cctgcggacg gtgccgacag
     4981 gtgcttctcg atctgcatcc tgggatcaaa gccatagtga aggacagtga tggacagccg
     5041 acggcagttg ggattcgtga attgctgccc tctggttatg tgtgggaggg ctaagcactt
     5101 cgtggccgag gagcaggact gacacgtgct acgagatttc gattccaccg ccgccttcta
     5161 tgaaaggttg ggcttcggaa tcgttttccg ggacgccggc tggatgatcc tccagcgcgg
     5221 ggatctcatg ctggagttct tcgcccaccc caacttgttt attgcagctt ataatggtta
     5281 caaataaagc aatagcatca caaatttcac aaataaagca tttttttcac tgcattctag
     5341 ttgtggtttg tccaaactca tcaatgtatc ttatcatgtc tgtataccgt cgacctctag
     5401 ctagagcttg gcgtaatcat ggtcatagct gtttcctgtg tgaaattgtt atccgctcac
     5461 aattccacac aacatacgag ccggaagcat aaagtgtaaa gcctggggtg cctaatgagt
     5521 gagctaactc acattaattg cgttgcgctc actgcccgct ttccagtcgg gaaacctgtc
     5581 gtgccagctg cattaatgaa tcggccaacg cgcggggaga ggcggtttgc gtattgggcg
     5641 ctcttccgct tcctcgctca ctgactcgct gcgctcggtc gttcggctgc ggcgagcggt
     5701 atcagctcac tcaaaggcgg taatacggtt atccacagaa tcaggggata acgcaggaaa
     5761 gaacatgtga gcaaaaggcc aggaaccgta aaaaggccgc gttgctggcg tttttccata
     5821 ggctccgccc ccctgacgag catcacaaaa atcgacgctc aagtcagagg tggcgaaacc
     5881 cgacaggact ataaagatac caggcgtttc cccctggaag ctccctcgtg cgctctcctg
     5941 ttccgaccct gccgcttacc ggatacctgt ccgcctttct cccttcggga agcgtggcgc
     6001 tttctcatag ctcacgctgt aggtatctca gttcggtgta ggtcgttcgc tccaagctgg
     6061 gctgtgtgca cgaacccccc gttcagcccg accgctgcgc cttatccggt aactatcgtc
     6121 ttgagtccaa cccggtaaga cacgacttat cgccactggc agcagccact ggtaacagga
     6181 ttagcagagc gaggtatgta ggcggtgcta cagagttctt gaagtggtgg cctaactacg
     6241 gctacactag aagaacagta tttggtatct gcgctctgct gaagccagtt accttcggaa
     6301 aaagagttgg tagctcttga tccggcaaac aaaccaccgc tggtagcggt ggtttttttg
     6361 tttgcaagca gcagattacg cgcagaaaaa aaggatctca agaagatcct ttgatctttt
     6421 ctacggggtc tgacgctcag tggaacgaaa actcacgtta agggattttg gtcatgagat
     6481 tatcaaaaag gatcttcacc tagatccttt taaattaaaa atgaagtttt aaatcaatct
     6541 aaagtatata tgagtaaact tggtctgaca gttaccaatg cttaatcagt gaggcaccta
     6601 tctcagcgat ctgtctattt cgttcatcca tagttgcctg actccccgtc gtgtagataa
     6661 ctacgatacg ggagggctta ccatctggcc ccagtgctgc aatgataccg cgagacccac
     6721 gctcaccggc tccagattta tcagcaataa accagccagc cggaagggcc gagcgcagaa
     6781 gtggtcctgc aactttatcc gcctccatcc agtctattaa ttgttgccgg gaagctagag
     6841 taagtagttc gccagttaat agtttgcgca acgttgttgc cattgctaca ggcatcgtgg
     6901 tgtcacgctc gtcgtttggt atggcttcat tcagctccgg ttcccaacga tcaaggcgag
     6961 ttacatgatc ccccatgttg tgcaaaaaag cggttagctc cttcggtcct ccgatcgttg
     7021 tcagaagtaa gttggccgca gtgttatcac tcatggttat ggcagcactg cataattctc
     7081 ttactgtcat gccatccgta agatgctttt ctgtgactgg tgagtactca accaagtcat
     7141 tctgagaata gtgtatgcgg cgaccgagtt gctcttgccc ggcgtcaata cgggataata
     7201 ccgcgccaca tagcagaact ttaaaagtgc tcatcattgg aaaacgttct tcggggcgaa
     7261 aactctcaag gatcttaccg ctgttgagat ccagttcgat gtaacccact cgtgcaccca
     7321 actgatcttc agcatctttt actttcacca gcgtttctgg gtgagcaaaa acaggaaggc
     7381 aaaatgccgc aaaaaaggga ataagggcga cacggaaatg ttgaatactc atactcttcc
     7441 tttttcaata ttattgaagc atttatcagg gttattgtct catgagcgga tacatatttg
     7501 aatgtattta gaaaaataaa caaatagggg ttccgcgcac atttccccga aaagtgccac
     7561 ctgacgtc
//
