1.Is the human dystrophin gene's intron structure related to its intron instability?
Wenli SHENG ; Jiangying CHEN ; Liangfu ZHU ; Zhuolin LIU
Chinese Medical Journal 2003;116(11):1733-1736
OBJECTIVETo study the human dystrophin gene molecular deletion mechanism, we analyzed breakpoint regions within junction fragments of deletion-type patients and investigated whether the dystrophin gene's intron structure might be related to intron instability.
METHODSJunction fragments corresponding to exon 46 and 51 deletions were cloned. The breakpoint regions were sequenced, and the features of introns with available Genebank sequences were analyzed.
RESULTSAn analysis of junction fragment sequences corresponding to exon 46 and 51 deletions showed that all 5' and 3' breakpoints are located within repeat sequences. No small insertions, small deletions, or point mutations are located near the breakpoint junctions. By analyzing the secondary structure of the junction fragments, we demonstrated that all junction fragment breakpoints are located in non-matching regions of single-stranded hairpin loops. A high concentration of repetitive elements is found to be a key feature of many dystrophin introns. In total, 34.8% of the overall dystrophin intron sequences is composed of repeat sequences.
CONCLUSIONRepeat elements in many dystrophin gene introns are the key to their structural bases and reflect intron instability. As a result of the primary DNA sequences, single-stranded hairpin loops form, increasing the instability of the gene, and forming the base for breaks in the DNA. The formation of the single-stranded hairpins can result in reattachment of two different breakpoints, producing a deletion.
Dystrophin ; genetics ; Humans ; Introns ; genetics ; Sequence Deletion
2.Analysis of gene inversion in Hemophilia A by Nanopore sequencing.
Peng DAI ; Yin FENG ; Chaofeng ZHU ; Xiangdong KONG
Chinese Journal of Medical Genetics 2021;38(6):521-525
OBJECTIVE:
To detect gene inversion in two pedigrees affected with Hemophilia A by using Nanopore sequencing technology.
METHODS:
Peripheral blood samples were taken from members of the two pedigrees. Following extraction of genome DNA, genetic variants of the carriers were detected by Nanopore sequencing and subjected to bioinformatic analysis.
RESULTS:
Nanopore sequencing has identified the niece of the proband of the pedigree 1 as carrier of Hemophilia A Inv22, and the mother of the proband of the pedigree 2 as carrier of Hemophilia A Inv1, which was consistent with clinical findings. Breakpoint sites in both pedigrees were accurately mapped. Statistical analysis of the sequencing results revealed a large number of variations in the carriers' genomes including deletions, duplications, insertions, inversions and translocations.
CONCLUSION
Nanopore sequencing can be used to analyze gene inversions associated with Hemophilia A, which also provided a powerful tool for the diagnosis of diseases caused by gene inversions.
Chromosome Inversion/genetics*
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Hemophilia A/genetics*
;
Humans
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Introns
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Nanopore Sequencing
;
Pedigree
3.Group Ⅱ introns and the application in biotechnology: a review.
Guzhen CUI ; Dengxiong HUA ; Junying GU ; Zhenghong CHEN
Chinese Journal of Biotechnology 2022;38(3):915-924
Group Ⅱ introns are self-splicing ribozymes, which insert directly into target sites in DNA with high frequency through "retrohoming". They specifically and efficiently recognize and splice DNA target sites, endowing themselves with great potential in genetic engineering. This paper reviewed the gene targeting principle of group Ⅱ introns and the application in microbial genetic modification, and then analyzed the limitations of them in multi-functional gene editing and eukaryotes based on the "retrohoming" characteristics and the dependence on high Mg2+ concentration. Finally, we dissected the potential of group Ⅱ introns in the development of novel gene editing tools based on our previous research outcome and the structural characteristics of the introns, hoping to provide a reference for the application of group Ⅱ introns in biotechnology.
DNA
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Eukaryota
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Gene Targeting
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Introns/genetics*
;
RNA, Catalytic/genetics*
4.Retrotransposon insertion polymorphism of the porcine esr gene and its association with production performances of Large White pigs.
Chenglin CHI ; Yalong AN ; Kaiyuan LI ; Hao GU ; Saisai WANG ; Cai CHEN ; Bo GAO ; Chengyi SONG ; Xiaoyan WANG
Chinese Journal of Biotechnology 2021;37(8):2794-2802
Estrogen receptor (esr) mediates the effects of estrogen on the expression of related genes, thereby regulating the growth and reproduction of mammals. To investigate the effect of retrotransposon insertion polymorphism (RIP) of the porcine esr gene on porcine growth performance, retrotransposon insertion polymorphism of the esr gene were predicted by comparative genomics and bioinformatics, and PCR was used to verify the insertion polymorphisms in different porcine breeds. Finally, the correlation analysis between the genotypes and performance of Large White pigs was conducted. The results showed that four retrotransposon polymorphic sites were identified in the esr1 and esr2 genes, which are esr1-SINE- RIP1 located in intron 2 of the esr1 gene, esr1-LINE-RIP2 and RIP3-esr1- SINE located in intron 5 of the gene, and esr2-LINE-RIP located in intron 1 of the esr2 gene, respectively. Among them, insertion of a 287 bp of SINE into intron 2 of the esr1 gene significantly affected (P<0.05) the live back fat thickness and 100 kg body weight back fat thickness of Large White pigs. Moreover, the live back fat thickness and back fat thickness at 100 kg body weight of homozygous with insertion (SINE+/+) was significantly greater than that of heterozygous with insertion (SINE+/-) and homozygous without insertion (SINE-/-). Therefore, esr1-SINE-RIP1 could be used as a molecular marker to assist the selection of deposition traits in Large White pigs.
Animals
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Genotype
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Introns/genetics*
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Phenotype
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Polymorphism, Genetic/genetics*
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Retroelements/genetics*
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Swine/genetics*
5.Intron 1 and 22 inversions in factor VIII gene in patients with haemophilia A.
Tan LI ; Jing DAI ; Jing-Sheng WU ; Qiu-Lan DING ; Kai-Yang DING ; Chang-Cheng ZHENG ; Ping SUN ; Xue-Feng WANG
Chinese Journal of Hematology 2009;30(3):150-153
OBJECTIVETo analyze intron 1 and 22 inversions in factor VIII (FVIII) gene in hemophilia A (HA) patients and and their families and to investigate the correlation between intron inversion and FVIII antibody.
METHODSAll patients were detected FVIII: C and FVIII antibody. In addition, 81 unrelated HA patients were directly detected by multiplex PCR and long-distance PCR for intron 1 and 22 inversions in FVIII gene. Pedigree investigation for some patients were conducted.
RESULTSIn 81 unrelated HA patients, 3 severe cases were found intron 1 inversion which accounted for 4.6% of total 65 severe cases. Of the 3 cases, one was FVIII antibody positive. Two female family members of a intron 1 inversion patient were identified as one carrier and one non-carrier. Twenty five of 65 (38.5%) severe cases were found intron 22 inversion. Of the 25 cases 1 was FVIII antibody positive. Nine female members in 5 HA families which had patients with intron 22 inversion were identified as 7 carries and 2 non-carriers.
CONCLUSIONBesides intron 22 inversion, intron 1 inversion was another important molecular defect in resulting in severe HA. Intron inversion analysis can also be used for deviation rectification of experiment grouping in HA patients. Intron 1 and 22 inversions may be one of the higher risk factors for resulting in FVIII antibodies.
Chromosome Inversion ; Chromosomes, Human, X ; Factor VIII ; genetics ; Female ; Hemophilia A ; genetics ; Humans ; Introns ; Male
6.Detection and genetic counseling of F8 gene inversions for patients with severe hemophilia A.
Nan BAI ; Qinghua WU ; Ning LIU ; Duo CHEN ; Zhenhua ZHAO ; Xiangdong KONG
Chinese Journal of Medical Genetics 2016;33(4):508-510
OBJECTIVEInversions of intron 1 (Inv1) or intron 22 (Inv22) of the coagulation factor VIII gene (F8) may be found in 40%-50% of patients with severe hemophilia A. Such inversions cannot be detected by conventional sequencing. Due to homologous recombination, family-based linkage analysis may yield false positive or false negative results. In this study, Inverse-shifting PCR (IS-PCR) was used to detect potential inversions in two families affected with hemophilia A.
METHODSPeripheral venous blood, fetal amniotic fluid and fetal chorionic cells were harvested for genome DNA extraction. IS-PCR was used to detect Inv1 or Inv22 detection or its subtypes.
RESULTSIS-PCR has accurately detected Inv22 and Inv1 in both families and verified the subtypes of Inv22.
CONCLUSIONCarriers of Inv22 or Inv1 may be precisely detected with IS-PCR. The results have provided valuable information for genetic counseling and prenatal diagnosis for the affected families.
Child ; Chromosome Inversion ; Factor VIII ; genetics ; Genetic Counseling ; Hemophilia A ; diagnosis ; genetics ; Humans ; Introns ; Male ; Prenatal Diagnosis
7.Relationship of von Willebrand factor gene single-nucleotide polymorphism with thrombosis diseases.
Journal of Experimental Hematology 2010;18(2):549-552
Recently it has been discovered that not only von Willebrand factor (vWF) decrease results in von Willebrand disease, but also vWF increase can lead to several thrombosis diseases. Plasma vWF level is affected by genetic factors. Individuals with different single nucleotide polymorphism (SNP) genotype in vWF have different susceptibility to disease; individuals with different blood group have different plasma vWF level. Environment factors also affect plasma vWF level. Understanding relationship of polymorphisms in promoter, exon and intron with thrombosis diseases contribute to prevent and cure these diseases. In this review, the relationship of SNP in promoter, exon and intron of vWF gene with thrombosis diseases is summarized.
Exons
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Genotype
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Humans
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Introns
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Polymorphism, Single Nucleotide
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Promoter Regions, Genetic
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Thrombosis
;
genetics
;
von Willebrand Factor
;
genetics
8.FVIII intron 22 homologous region mediated recombinations and genotyping methods of hemophilia A.
Journal of Experimental Hematology 2014;22(3):865-868
The intron 22 inversion is caused by intrachromosomal homologous recombination of the Int22h (intron 22 homologous region) repeats in opposite orientation, accounting for almost 45%-50% of all cases of severe hemophilia A. By contrast, recombinations between similarly oriented int22h repeats in the same chromosome, homologous chromosomes or chromatids may cause deletion or duplication of the intermediate region between the two int22h copies, resulting in false-positive or false-negative possibility that obscure the characterization of intron 22 inversion. The modified long distance PCR(LD-PCR) and inverse shifting PCR(IS-PCR) would distinguish all the possible int22h-mediated rearrangements.
Factor VIII
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genetics
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Genotype
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Hemophilia A
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genetics
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Humans
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Introns
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Polymerase Chain Reaction
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methods
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Recombination, Genetic
9.Analysis of pathogenic variants of USH2A gene in a child with Usher syndrome type II.
Kefeng TANG ; Liyan JIANG ; Juan YAO ; Sheng YANG ; Guosong SHEN
Chinese Journal of Medical Genetics 2021;38(10):966-968
OBJECTIVE:
To detect pathogenic variant in a child featuring Usher syndrome type II.
METHODS:
Peripheral blood samples of the child and his parents were collected for the analysis of variants of hearing impairment-related genes. The findings were verified in 100 individuals with normal hearing.
RESULTS:
The child was found to harbor compound heterozygous variants of the USH2A gene, namely c.8224-1G>C in intron 41 and c.5678C>G(p.Ser1893X) in exon 28, which were inherited respectively from his mother and father. Based on the American College of Medical Genetics and Genomics standards and guidelines, both c.8224-1G>C and c.5678C>G(p.Ser1893X) variants of USH2A gene were predicted to be pathogenic(PVS1+PM2+PM3).
CONCLUSION
The compound heterozygous variants c.8224-1G>C and c.5678C>G of the USH2A gene probably underlay the disease in this child. Above finding has enriched the spectrum of USH2A gene variants.
Child
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Exons
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Extracellular Matrix Proteins/genetics*
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Family
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Humans
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Introns
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United States
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Usher Syndromes/genetics*
10.A pedigree analysis of a rare RhD 336-1G>A intron variant.
Yuefeng WANG ; Haijiang CHEN ; Yan ZENG ; Xiaoliang SHI ; Jianjun ZHANG
Chinese Journal of Medical Genetics 2021;38(8):803-806
OBJECTIVE:
To explore the molecular mechanism of a case where RhD genotyping did not match serological results.
METHODS:
The serological results of 8 members from two generations of this family were analyzed. And according to Mendelian law of inheritance, RhD genotyping, zygotic type determination and gene sequencing were performed for the family members.
RESULTS:
The proband and one of her cousins have the same RhD alleles, both of them have a 336-1G>A intron variant RhD allele and a complete RhD deletion allele. The variant alleles are inherited from two of their parents with blood relationship, while the complete-deleted alleles come from the other. 336-1G>A means that the last base G of the second intron of the RhD gene is mutated to A, which leads to a negative RhD serology and a positive genotype in the proband.
CONCLUSION
There was a rare 336-1G> A intron variant gene (RhD * 01N.25) in this family, which was a recessive gene relative to the RhD gene and resulted in RhD phenotype negative.
Alleles
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Female
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Genotype
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Humans
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Introns/genetics*
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Pedigree
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Phenotype
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Rh-Hr Blood-Group System/genetics*