LOCUS       sequence-249151-        7418 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expressing wildtype human Rad9b gene.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7418)
  AUTHORS   Cao X, Tian T, Steele JW, Cabrera RM, Aguiar-Pulido V, Wadhwa S, 
            Bhavani N, Bi P, Gargurevich NH, Hoffman EN, Cai CQ, Marini NJ, Yang
            W, Shaw GM, Ross ME, Finnell RH, Lei Y
  TITLE     Loss of RAD9B impairs early neural development and contributes to 
            the risk for human spina bifida.
  JOURNAL   Hum Mutat. 2020 Apr;41(4):786-799. doi: 10.1002/humu.23969. Epub 
            2020 Jan 16.
  PUBMED    31898828
REFERENCE   2  (bases 1 to 7418)
  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: 
            "10.1002/humu.23969"; journalName: "Hum Mutat"; date: "2020-04"; 
            volume: "41"; issue: "4"; pages: "786-799"
COMMENT     Sequence Label: R9-WT
FEATURES             Location/Qualifiers
     source          1..7418
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        69..448
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        449..652
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     603..623
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     697..716
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        697..715
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             757..1470
                     /codon_start=1
                     /product="green fluorescent protein"
                     /label=GFP
                     /translation="MSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKV
                     NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLLE
                     FVTAAGITHGMDELYK"
     CDS             join(757..759,760..1470)
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="MSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKV
                     NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLLE
                     FVTAAGITHGMDELYK"
     primer_bind     complement(799..820)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(1060..1079)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1407..1428
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     complement(2843..2860)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    2849..3073
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      3119..3547
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3206..3225)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3416..3437
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(3556..3576)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        3561..3890
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      3741..3876
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     3803..3822
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             3957..4751
                     /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
                     GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(4011..4030)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4621..4640
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    4925..5046
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4962..4981)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     5016..5035
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(5095..5111)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5095..5111)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(5108..5130)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5119..5135
                     /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(5143..5149,5150..5167,5168..5173))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5188..5209
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5326..5343)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5497..6085)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5577..5596)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6256..7047,7048..7116))
                     /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     6879..6898
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7117..7221)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(7296..7315)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
ORIGIN
        1 catgaagaat ctgcttaggg ttaggcgttt tgcgctgctt cgcgatgtac gggccagata
       61 tacgcgttga cattgattat tgactagtta ttaatagtaa tcaattacgg ggtcattagt
      121 tcatagccca tatatggagt tccgcgttac ataacttacg gtaaatggcc cgcctggctg
      181 accgcccaac gacccccgcc cattgacgtc aataatgacg tatgttccca tagtaacgcc
      241 aatagggact ttccattgac gtcaatgggt ggagtattta cggtaaactg cccacttggc
      301 agtacatcaa gtgtatcata tgccaagtac gccccctatt gacgtcaatg acggtaaatg
      361 gcccgcctgg cattatgccc agtacatgac cttatgggac tttcctactt ggcagtacat
      421 ctacgtatta gtcatcgcta ttaccatggt gatgcggttt tggcagtaca tcaatgggcg
      481 tggatagcgg tttgactcac ggggatttcc aagtctccac cccattgacg tcaatgggag
      541 tttgttttgg caccaaaatc aacgggactt tccaaaatgt cgtaacaact ccgccccatt
      601 gacgcaaatg ggcggtaggc gtgtacggtg ggaggtctat ataagcagag ctctctggct
      661 aactagagaa cccactgctt actggcttat cgaaattaat acgactcact atagggagac
      721 ccaagctggc tagcgtttaa acttaagctt gccaccatga gcaagggcga ggagctgttc
      781 accggggtgg tgcccatcct ggtcgagctg gacggcgacg taaacggcca caagttcagc
      841 gtgtccggcg agggcgaggg cgatgccacc tacggcaagc tgaccctgaa gttcatctgc
      901 accaccggca agctgcccgt gccctggccc accctcgtga ccaccctgac ctacggcgtg
      961 cagtgcttca gccgctaccc cgaccacatg aagcagcacg acttcttcaa gtccgccatg
     1021 cccgaaggct acgtccagga gcgcaccatc ttcttcaagg acgacggcaa ctacaagacc
     1081 cgcgccgagg tgaagttcga gggcgacacc ctggtgaacc gcatcgagct gaagggcatc
     1141 gacttcaagg aggacggcaa catcctgggg cacaagctgg agtacaacta caacagccac
     1201 aacgtctata tcatggccga caagcagaag aacggcatca aggtgaactt caagatccgc
     1261 cacaacatcg aggacggcag cgtgcagctc gccgaccact accagcagaa cacccccatc
     1321 ggcgacggcc ccgtgctgct gcccgacaac cactacctga gcacccagtc cgccctgagc
     1381 aaagacccca acgagaagcg cgatcacatg gtcctgctgg agttcgtgac cgccgccggg
     1441 atcactcacg gcatggacga gctgtacaag ggtaccatgg cagccatgct gaagtgcgtg
     1501 atgagcggca gtcaggtgaa agtatttggg aaagcagttc aagctctatc acgaattagt
     1561 gacgagttct ggctagaccc atctaaaaaa ggtcttgctc taagatgtgt gaattcttct
     1621 cggtcagcat atggatgtgt cctgttctct cctgtgtttt ttcagcatta tcaatggtca
     1681 gctttagtga aaatgagtga aaatgaactt gacacaacac tgcatttaaa atgcaaattg
     1741 ggaatgaagt caattttgcc catctttaga tgtctgaatt cccttgaaag aaatatagag
     1801 aagtgcagaa tattcaccag atctgataaa tgcaaagtag ttattcaatt cttctacaga
     1861 catggtatta aaagaactca taatatatgt tttcaagaaa gtcagccttt gcaagttatt
     1921 tttgacaaga atgtttgtac taatacgcta atgattcaac caagattgct tgctgatgcc
     1981 attgttcttt ttacatcaag tcaagaggaa gttactcttg ctgttactcc actgaatttt
     2041 tgcctcaaga gttctaatga ggaatcaatg gatttgagca atgctgtaca cagtgagatg
     2101 tttgttggct cagatgagtt tgacttcttt caaattggaa tggacactga gataacattt
     2161 tgtttcaaag aattgaaggg aatactgaca ttttcagaag ctacacatgc tcctatatcc
     2221 atttattttg atttccctgg gaaacctctg gctttgagta ttgatgatat gttagtggaa
     2281 gctaacttta ttttggccac attagctgat gaacaaagta gagcatcttc accacagtca
     2341 ctgtgtcttt cacagaaacg aaaaaggtca gatctgattg aaaaaaaggc tggcaaaaat
     2401 gtaactggcc aggccctgga atgtatttca aaaaaagcag caccaagaag gctttatcct
     2461 aaggagactc tcacaaacat atctgcattg gaaaactgtg gcagccctgc aatgaaaaga
     2521 gtggatggag atgtcagtga agtatcagaa agcagtgtca gcaacacaga ggaagtgcca
     2581 gggtctctgt gtctcagaaa gttttcttgc atgttctttg gagcagtttc ttctgaccag
     2641 caagaacact tcaaccaccc tttcgacagt ctggcaagag caagtgacag tgaagaggac
     2701 atgaataatg tgtgctgcag gaaagaattt aatggaagtg atgccaaata tttctgtatt
     2761 atctgaggat ccgaattctg cagatatcca gcacagtggc ggccgctcga gtctagaggg
     2821 cccgtttaaa cccgctgatc agcctcgact gtgccttcta gttgccagcc atctgttgtt
     2881 tgcccctccc ccgtgccttc cttgaccctg gaaggtgcca ctcccactgt cctttcctaa
     2941 taaaatgagg aaattgcatc gcattgtctg agtaggtgtc attctattct ggggggtggg
     3001 gtggggcagg acagcaaggg ggaggattgg gaagacaata gcaggcatgc tggggatgcg
     3061 gtgggctcta tggcttctga ggcggaaaga accagctggg gctctagggg gtatccccac
     3121 gcgccctgta gcggcgcatt aagcgcggcg ggtgtggtgg ttacgcgcag cgtgaccgct
     3181 acacttgcca gcgccctagc gcccgctcct ttcgctttct tcccttcctt tctcgccacg
     3241 ttcgccggct ttccccgtca agctctaaat cgggggctcc ctttagggtt ccgatttagt
     3301 gctttacggc acctcgaccc caaaaaactt gattagggtg atggttcacg tagtgggcca
     3361 tcgccctgat agacggtttt tcgccctttg acgttggagt ccacgttctt taatagtgga
     3421 ctcttgttcc aaactggaac aacactcaac cctatctcgg tctattcttt tgatttataa
     3481 gggattttgc cgatttcggc ctattggtta aaaaatgagc tgatttaaca aaaatttaac
     3541 gcgaattaat tctgtggaat gtgtgtcagt tagggtgtgg aaagtcccca ggctccccag
     3601 caggcagaag tatgcaaagc atgcatctca attagtcagc aaccaggtgt ggaaagtccc
     3661 caggctcccc agcaggcaga agtatgcaaa gcatgcatct caattagtca gcaaccatag
     3721 tcccgcccct aactccgccc atcccgcccc taactccgcc cagttccgcc cattctccgc
     3781 cccatggctg actaattttt tttatttatg cagaggccga ggccgcctct gcctctgagc
     3841 tattccagaa gtagtgagga ggcttttttg gaggcctagg cttttgcaaa aagctcccgg
     3901 gagcttgtat atccattttc ggatctgatc aagagacagg atgaggatcg tttcgcatga
     3961 ttgaacaaga tggattgcac gcaggttctc cggccgcttg ggtggagagg ctattcggct
     4021 atgactgggc acaacagaca atcggctgct ctgatgccgc cgtgttccgg ctgtcagcgc
     4081 aggggcgccc ggttcttttt gtcaagaccg acctgtccgg tgccctgaat gaactgcagg
     4141 acgaggcagc gcggctatcg tggctggcca cgacgggcgt tccttgcgca gctgtgctcg
     4201 acgttgtcac tgaagcggga agggactggc tgctattggg cgaagtgccg gggcaggatc
     4261 tcctgtcatc tcaccttgct cctgccgaga aagtatccat catggctgat gcaatgcggc
     4321 ggctgcatac gcttgatccg gctacctgcc cattcgacca ccaagcgaaa catcgcatcg
     4381 agcgagcacg tactcggatg gaagccggtc ttgtcgatca ggatgatctg gacgaagagc
     4441 atcaggggct cgcgccagcc gaactgttcg ccaggctcaa ggcgcgcatg cccgacggcg
     4501 aggatctcgt cgtgacccat ggcgatgcct gcttgccgaa tatcatggtg gaaaatggcc
     4561 gcttttctgg attcatcgac tgtggccggc tgggtgtggc ggaccgctat caggacatag
     4621 cgttggctac ccgtgatatt gctgaagagc ttggcggcga atgggctgac cgcttcctcg
     4681 tgctttacgg tatcgccgct cccgattcgc agcgcatcgc cttctatcgc cttcttgacg
     4741 agttcttctg agcgggactc tggggttcga aatgaccgac caagcgacgc ccaacctgcc
     4801 atcacgagat ttcgattcca ccgccgcctt ctatgaaagg ttgggcttcg gaatcgtttt
     4861 ccgggacgcc ggctggatga tcctccagcg cggggatctc atgctggagt tcttcgccca
     4921 ccccaacttg tttattgcag cttataatgg ttacaaataa agcaatagca tcacaaattt
     4981 cacaaataaa gcattttttt cactgcattc tagttgtggt ttgtccaaac tcatcaatgt
     5041 atcttatcat gtctgtatac cgtcgacctc tagctagagc ttggcgtaat catggtcata
     5101 gctgtttcct gtgtgaaatt gttatccgct cacaattcca cacaacatac gagccggaag
     5161 cataaagtgt aaagcctggg gtgcctaatg agtgagctaa ctcacattaa ttgcgttgcg
     5221 ctcactgccc gctttccagt cgggaaacct gtcgtgccag ctgcattaat gaatcggcca
     5281 acgcgcgggg agaggcggtt tgcgtattgg gcgctcttcc gcttcctcgc tcactgactc
     5341 gctgcgctcg gtcgttcggc tgcggcgagc ggtatcagct cactcaaagg cggtaatacg
     5401 gttatccaca gaatcagggg ataacgcagg aaagaacatg tgagcaaaag gccagcaaaa
     5461 ggccaggaac cgtaaaaagg ccgcgttgct ggcgtttttc cataggctcc gcccccctga
     5521 cgagcatcac aaaaatcgac gctcaagtca gaggtggcga aacccgacag gactataaag
     5581 ataccaggcg tttccccctg gaagctccct cgtgcgctct cctgttccga ccctgccgct
     5641 taccggatac ctgtccgcct ttctcccttc gggaagcgtg gcgctttctc atagctcacg
     5701 ctgtaggtat ctcagttcgg tgtaggtcgt tcgctccaag ctgggctgtg tgcacgaacc
     5761 ccccgttcag cccgaccgct gcgccttatc cggtaactat cgtcttgagt ccaacccggt
     5821 aagacacgac ttatcgccac tggcagcagc cactggtaac aggattagca gagcgaggta
     5881 tgtaggcggt gctacagagt tcttgaagtg gtggcctaac tacggctaca ctagaagaac
     5941 agtatttggt atctgcgctc tgctgaagcc agttaccttc ggaaaaagag ttggtagctc
     6001 ttgatccggc aaacaaacca ccgctggtag cggtggtttt tttgtttgca agcagcagat
     6061 tacgcgcaga aaaaaaggat ctcaagaaga tcctttgatc ttttctacgg ggtctgacgc
     6121 tcagtggaac gaaaactcac gttaagggat tttggtcatg agattatcaa aaaggatctt
     6181 cacctagatc cttttaaatt aaaaatgaag ttttaaatca atctaaagta tatatgagta
     6241 aacttggtct gacagttacc aatgcttaat cagtgaggca cctatctcag cgatctgtct
     6301 atttcgttca tccatagttg cctgactccc cgtcgtgtag ataactacga tacgggaggg
     6361 cttaccatct ggccccagtg ctgcaatgat accgcgagac ccacgctcac cggctccaga
     6421 tttatcagca ataaaccagc cagccggaag ggccgagcgc agaagtggtc ctgcaacttt
     6481 atccgcctcc atccagtcta ttaattgttg ccgggaagct agagtaagta gttcgccagt
     6541 taatagtttg cgcaacgttg ttgccattgc tacaggcatc gtggtgtcac gctcgtcgtt
     6601 tggtatggct tcattcagct ccggttccca acgatcaagg cgagttacat gatcccccat
     6661 gttgtgcaaa aaagcggtta gctccttcgg tcctccgatc gttgtcagaa gtaagttggc
     6721 cgcagtgtta tcactcatgg ttatggcagc actgcataat tctcttactg tcatgccatc
     6781 cgtaagatgc ttttctgtga ctggtgagta ctcaaccaag tcattctgag aatagtgtat
     6841 gcggcgaccg agttgctctt gcccggcgtc aatacgggat aataccgcgc cacatagcag
     6901 aactttaaaa gtgctcatca ttggaaaacg ttcttcgggg cgaaaactct caaggatctt
     6961 accgctgttg agatccagtt cgatgtaacc cactcgtgca cccaactgat cttcagcatc
     7021 ttttactttc accagcgttt ctgggtgagc aaaaacagga aggcaaaatg ccgcaaaaaa
     7081 gggaataagg gcgacacgga aatgttgaat actcatactc ttcctttttc aatattattg
     7141 aagcatttat cagggttatt gtctcatgag cggatacata tttgaatgta tttagaaaaa
     7201 taaacaaata ggggttccgc gcacatttcc ccgaaaagtg ccacctgacg tcgacggatc
     7261 gggagatctc ccgatcccct atggtgcact ctcagtacaa tctgctctga tgccgcatag
     7321 ttaagccagt atctgctccc tgcttgtgtg ttggaggtcg ctgagtagtg cgcgagcaaa
     7381 atttaagcta caacaaggca aggcttgacc gacaattg
//
