Short and sweet: foreleg abnormalities in Havanese and the role of the FGF4 retrogene

Abstract Background Cases of foreleg deformities, characterized by varying degrees of shortened and bowed forelegs, have been reported in the Havanese breed. Because the health and welfare implications are severe in some of the affected dogs, further efforts should be made to investigate the genetic background of the trait. A FGF4-retrogene on CFA18 is known to cause chondrodystrophy in dogs. In most breeds, either the wild type allele or the mutant allele is fixed. However, the large degree of genetic diversity reported in Havanese, could entail that both the wild type and the mutant allele segregate in this breed. We hypothesize that the shortened and bowed forelegs seen in some Havanese could be a consequence of FGF4RG-associated chondrodystrophy. Here we study the population prevalence of the wild type and mutant allele, as well as effect on phenotype. We also investigate how the prevalence of the allele associated with chondrodystrophy have changed over time. We hypothesize that recent selection, may have led to a gradual decline in the population frequency of the lower-risk, wild type allele. Results We studied the FGF4-retrogene on CFA18 in 355 Havanese and found variation in the presence/absence of the retrogene. The prevalence of the non-chondrodystrophic wild type is low, with allele frequencies of 0.025 and 0.975 for the wild type and mutant allele, respectively (linked marker). We found that carriers of the beneficial wild type allele were significantly taller at the shoulder than mutant allele homozygotes, with average heights of 31.3 cm and 26.4 cm, respectively. We further found that wild type carriers were born on average 4.7 years earlier than mutant allele homozygotes and that there has been a gradual decline in the population frequency of the wild type allele during the past two decades. Conclusions Our results indicate that FGF4RG-associated chondrodystrophy may contribute to the shortened forelegs found in some Havanese and that both the wild type and mutant allele segregate in the breed. The population frequency of the wild type allele is low and appear to be decreasing. Efforts should be made to preserve the healthier wild type in the population, increase the prevalence of a more moderate phenotype and possibly reduce the risk of foreleg pathology.

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Identification and Complementation of a Mutation Associated with Loss of Mycoplasma pneumoniae Virulence-Specific Proteins B and C

Journal of Bacteriology ◽

10.1128/jb.187.2.747-751.2005 ◽

2005 ◽

Vol 187 (2) ◽

pp. 747-751 ◽

Cited By ~ 16

Author(s):

Robert H. Waldo ◽

Jarrat L. Jordan ◽

Duncan C. Krause

Keyword(s):

Mycoplasma Pneumoniae ◽

Wild Type Allele ◽

Wild Type ◽

Type Allele ◽

Specific Proteins

ABSTRACT A mutation in gene MPN142 (orf6) was identified in the Mycoplasma pneumoniae cytadherence mutant III-4. MPN142 encodes virulence-specific proteins P90 and P40 (proteins B and C, respectively). Analysis of MPN142 in a cytadhering revertant and complementation using a recombinant wild-type allele confirmed the role of this mutation in the cytadherence defect.

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THE MUTATION SK(ad-3A) CANCELS THE DOMINANCE OF ad-3A + OVER ad-3A IN THE ASCUS OF NEUROSPORA

Genetics ◽

10.1093/genetics/97.2.237 ◽

1981 ◽

Vol 97 (2) ◽

pp. 237-246

Author(s):

A M Delange

Keyword(s):

Mutant Allele ◽

Wild Type Allele ◽

Wild Type ◽

Equal Frequency ◽

Type Allele ◽

Spore Killer ◽

Induced Mutant ◽

Ascospore Formation

ABSTRACT A newly induced mutant of Neurospora, when crossed with an ad-3A mutant, produces asci with four viable black and four inviable white ascospores. The survivors always contain the new mutant allele, never ad-3A. The new allele, which is called SK(ad-3A) (for spore killer of ad-3A), is located at or very near the ad-3A locus. —In crosses homozygous for ad-3A, each ascus contains only inviable white ascospores. This defect in ascospore maturation is complemented by the wild-type allele, ad-3A + (crosses heterozygous for ad-3A and ad-3A + produce mainly viable ascospores), but it is not complemented by the new SK(ad-3A) allele (all ad-3A ascospores from crosses heterozygous for SK(ad-3A) and ad-3A are white and inviable). In crosses homozygous for SK(ad-3A) or heterozygous for SK(ad-3A) and ad-3A +, each ascus contains only viable black ascospores. SK(ad-3A) does not require adenine for growth, and forced heterokaryons between SK(ad-3A) and ad-3A grow at wild-type rates and produce conidia of both genotypes with approximately equal frequency. Thus, the action of SK(ad-3A) is apparently restricted to ascospore formation. Possible mechanisms of the action of this new allele are discussed.

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Positive control of sulphate reduction in Escherichia coli. The nature of the pleiotropic cysteineless mutants of E. coli K 12

Biochemical Journal ◽

10.1042/bj1100597 ◽

1968 ◽

Vol 110 (3) ◽

pp. 597-602 ◽

Cited By ~ 21

Author(s):

M. C. Jones-Mortimer

Keyword(s):

Escherichia Coli ◽

Mutant Allele ◽

Positive Control ◽

Wild Type Allele ◽

Wild Type ◽

E Coli ◽

Type Allele ◽

K 12 ◽

Sulphite Reductase

1. The function of the wild-type alleles of the pleiotropic mutants cysB and cysE of Escherichia coli was investigated. 2. The wild-type allele cysB+ is dominant to the mutant allele cysB in stable and transient heterozygotes. 3. The wild-type allele cysE+ is dominant to the mutant allele cysE, as predicted. 4. Sulphur-starved cultures of cysB or cysE strains contain less than 0·2nmole of free cysteine/mg. dry wt. 5. Complementation in vitro is not observed between extracts of cysB mutants and mutants lacking sulphite reductase only. 6. A scheme, involving positive control of the enzymes of sulphate activation and reduction, is suggested to account for the control of cysteine biosynthesis.

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THE REGULATION OF WHITE LOCUS EXPRESSION: A DOMINANT MUTANT ALLELE AT THE WHITE LOCUS OF DROSOPHILA MELANOGASTER

Genetics ◽

10.1093/genetics/95.2.341 ◽

1980 ◽

Vol 95 (2) ◽

pp. 341-353

Author(s):

Paul M Bingham

Keyword(s):

Drosophila Melanogaster ◽

Mutant Allele ◽

Chromosomal Rearrangements ◽

Wild Type Allele ◽

Wild Type ◽

Type Allele ◽

Dominant Mutant ◽

White Locus ◽

Somatic Pairing

ABSTRACT A new mutant allele (wDZL)at the white locus of Drosophila melanogaster is dominant to the wild-type allele, but apparently only when the two alleles are synapsed. When chromosomal rearrangements prevent somatic pairing between the two white alleles, wDZL is rendered recessive to wild type. This observation suggests that the dominance of wDZL is sensitive to a synapsis (transvection) effect. On the basis of this and other properties, it is proposed that wDZL causes the repression of transcription of a synapsed w+ allele, but not of a w+ allele elsewhere in the same nucleus. One model to account for this supposes that wDzL produces a repressor of white-locus transcription. This repressor is presumed to be so unstable that other white genes, removed from wDZL but in the same nucleus, are not detectably repressed. These properties may be simply understood if it is assumed that the repressor produced by the wDZL allele is an RNA molecule.

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MULTIPLEX PCR BASED SINGLE NUCLEOTIDE POLYMORPHISMS ANALYSIS OF HBS POINT MUTATION

10.36106/paripex/2702115 ◽

2020 ◽

pp. 1-4

Author(s):

Jignisha S Patel ◽

Jignaben P Naik ◽

Yazdi M Italia

Keyword(s):

Multiplex Pcr ◽

Point Mutation ◽

Sickle Cell ◽

Mutant Allele ◽

Snp Analysis ◽

Wild Type Allele ◽

Wild Type ◽

Single Nucleotide ◽

Type Allele ◽

Cell Carrier

Introduction: Sickle hemoglobin (HbS), an autosomal recessive hemoglobinopathy cause of Sickle cell disease (SCD), is widely sprayed around the globe affecting millions of people . SCD results from single nucleotide polymorphism (SNP) or point mutation causing amino acid substitution from Glutamic acid to Valine leads to sickled shape red blood cells. SNPs can be well studied by using allele-specific amplification (ASA) technique. Aims & Objective: To develop a simple, rapid, easy and accurate genotyping method for SNP analysis of SCD. Materials and methods: By performing different tests, a well characterized sample panel of 150 different types of samples was prepared. From this sample panel DNA was extracted and used for SNP-genotyping of SCD. Specific primers were used for performing monoplex PCR amplification of wild type allele (HbAA) and mutant allele (HbSS) were performed individually. By using the same primers multiplex PCR assay was experimented. Results and conclusion: This is a simple and low cost molecular method for the detection of point mutation and useful tool for the diagnosis of SCD. The entire analysis can be performed in one reaction mixture, which results in higher speed, higher accuracy, and the need for smaller samples. This technique might be of great value for genotyping of homozygous sickle cell patients (SS) and heterozygous sickle cell trait (AS). But we found one discrepancy with double heterozygous (sickle β-thalassaemia) samples. We were not able to differentiate sickle cell carrier state (AS) from the double heterozygous like sickle β-thalassaemia state. So we conclude that for simultaneous detection of thalassaemia along with sickle cell requires addition of more primers specific for thalassaemia mutation. In addition to this when two bands, one for wild type allele and second for mutant allele appears, care must be taken to conclude whether the person is a sickle cell carrier (AS) or having double heterozygous (sickle β-thalassaemia) like condition.

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SELECTIVE ALLELE LOSS IN MIXED INFECTIONS WITH T4 BACTERIOPHAGE

Genetics ◽

10.1093/genetics/73.1.1 ◽

1973 ◽

Vol 73 (1) ◽

pp. 1-11

Author(s):

Wendy C Benz ◽

Hillard Berger

Keyword(s):

Mutant Allele ◽

Deletion Mutants ◽

Mixed Infections ◽

Repair System ◽

Wild Type Allele ◽

Wild Type ◽

Heteroduplex Dna ◽

E Coli ◽

Type Allele ◽

T4 Bacteriophage

ABSTRACT Evidence is presented that when E. coli B is mixedly infected with T4D wild type and rII deletion mutants, the excess DNA of the wild type allele is lost. No loss is seen in mixed infections with rII point mutants and wild type. In similar experiments with lysozyme addition mutants, the mutant allele is lost. We believe these results demonstrate a repair system which removes "loops" in heteroduplex DNA molecules. A number of phage and host functions have been tested for involvement in the repair of the excess DNA, and T4 genes x and v have been implicated in this process.

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The Role of MLL1 and MLL2 in MLL Fusion Oncoprotein-Initiated Leukemia

Blood ◽

10.1182/blood.v128.22.573.573 ◽

2016 ◽

Vol 128 (22) ◽

pp. 573-573

Author(s):

Patricia Ernst ◽

Konstantinos Anastassiadis ◽

Andrea Kranz ◽

A. Francis Stewart ◽

Kathrin Arndt ◽

...

Keyword(s):

Fusion Protein ◽

Histone Methyltransferase ◽

Myelogenous Leukemia ◽

Leukemia Cells ◽

Wild Type Allele ◽

Wild Type ◽

Double Knockout ◽

Hematopoietic Stem ◽

Type Allele

Abstract The MLL1 histone methyltransferase gene at 11q23 undergoes many distinct chromosomal translocations to yield poor prognosis leukemia. In these MLL-rearranged (MLL-r) leukemias, the remaining wild-type allele is typically expressed, and the fusion oncoprotein lacks the C terminal histone methyltransferase domain. The role of the wild-type, endogenous MLL1 allele in MLL-fusion protein mediated leukemogenesis is not fully understood, with some evidence suggesting an important functional role. On the other hand, the wild-type allele is lost in some patients and in the MLL-r ML2 cell line, suggesting that the wild-type allele is not absolutely required. Here we rigorously address the role of endogenous MLL1 in MLL-r leukemia. Using precise and independent animal models, we show that MLL1 is dispensable for MLL-AF9- or MLL-AF6-mediated acute myelogenous leukemia (AML). In parallel experiments, deletion of Menin, encoding a chromatin targeting cofactor for MLL-fusion proteins, has severe effects on the proliferation and survival of MLL-r leukemia, as previously published. Potential redundancy between MLL family methyltransferases was addressed by co-deleting the closest paralog, Mll2. Surprisingly, deletion of Mll2 by itself had a significant impact on survival of MLL-AF9 leukemia cells, and co-deletion of Mll1 and Mll2 more effectively killed these cells, suggesting a redundant or synthetic lethal relationship between the two genes. A comparison of gene expression and histone modification changes in Mll1, Mll2 and Mll1;Mll2 double knockout leukemia cells showed that 1) global defects in histone H3, lysine 4 methyation can be exclusively attributed to MLL2, 2) MLL1- versus MLL2-dependent genes are largely distinct, and 3) Mll1;Mll2 double knockout leukemia cells exhibit deficiencies in NFκB and integrin ß3 pathways, but exhibit little evidence of deregulation of either MLL1- or MLL-fusion protein target genes. These findings reshape our thinking about the role of endogenous MLL family proteins in MLL-fusion protein leukemogenesis. The data presented here, in conjunction with the minimal effect of Mll2 deletion on hematopoietic stem and progenitor cells make MLL2 an attractive candidate for targeted therapy in MLL-r leukemia. Disclosures Ernst: Amgen: Other: stocks. Yokoyama:Sumitomo Dainippon Pharma: Research Funding. Neff:Bristol Myers Squibb: Other: Travel; Janssen: Other: Travel; Epizyme: Patents & Royalties: patent filed.

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O-090 Correcting a PLCζ mutation in the human germ line to overcome hereditary infertility

Human Reproduction ◽

10.1093/humrep/deab125.020 ◽

2021 ◽

Vol 36 (Supplement_1) ◽

Author(s):

B Bekaert ◽

A Boel ◽

M Popovic ◽

P Stamatiadis ◽

S M Chuva de Sousa Lopes ◽

...

Keyword(s):

Loss Of Heterozygosity ◽

Base Pair ◽

Mutant Allele ◽

Germ Line ◽

Wild Type Allele ◽

Fertilization Failure ◽

Wild Type ◽

Type Allele ◽

Synonymous Variant ◽

Repair Template

Abstract Study question Can clustered regularly interspaced short palindromic repeats (CRISPR)/Cas9 gene editing result in the correction of a single base pair substitution that causes male infertility? Summary answer CRISPR/Cas9 administration during intracytoplasmic sperm injection (ICSI) leads to correction attempts of mutant phospholipase C zeta (PLCζ), howeverc loss-of-heterozygosity (LOH). What is known already Failed fertilization after ICSI can be caused by mutations in the sperm-related oocyte factor PLCζ which can be overcome by assisted oocyte activation (AOA). In this way, children may inherit the infertility-causing mutation. Mutation transmission can be overcome through CRISPR/Cas9 delivery during ICSI. In previous studies using CRISPR/Cas9 in the human germline for mutation correction, loss-of-heterozygosity (LOH, loss of the allele of one of the parents) was observed. Two different explanations were given, namely partial or complete paternal chromosomal loss or the correction of the mutation by using the maternal wild-type allele instead of the exogeneous supplied repair template. Study design, size, duration We injected a gRNA-Cas9 protein complex to target the PLCζ mutant allele, a repair template harboring the desired nucleotide substitution and an additional synonymous variant to track template usage, together with patient’s sperm. To overcome fertilization failure, AOA was applied during ICSI. After a culture period of maximal 6 days the embryos were collected. At day 3, some embryos were dissociated in individual blastomeres. The extracted DNA was analyzed through different genetic sequencing techniques. Participants/materials, setting, methods Donated sperm of a patient experiencing complete fertilization failure after routine ICSI, harboring a heterozygous base pair substitution in PLCZ1 (c.136-1G>C), was utilized. Sperm was injected in donated in vitro matured oocytes or in vivo matured oocytes containing clusters of smooth endoplasmic reticulum. Next-generation sequencing was used to assess correction potential. Short tandem repeat (STR) and single nucleotide polymorphism (SNP) assays were used to determine whether the sperm contained the mutation and to evaluate LOH. Main results and the role of chance CRISPR/Cas9 injections had no significant impact (p > 0.05) on embryonic development. Due to the heterozygous nature of the mutation, 47% (27/58) of the embryos originated from mutated sperm injection. The CRISPR components showed a high specificity with absence of insertions/deletions in 97% of the embryos originating from wild-type sperm (n = 31). Embryos originating from mutant sperm (n = 27) fall into three categories:(1) 22% showed the untargeted mutant allele, (2) 52% showed additional mutagenesis and (3) 26% showed the wild-type allele, which could be explained by correction. Mosaicism, defined as various editing events, was present in 17% (1), 21% (2) and 71% (3) of the embryos. The low occurrence of the synonymous variant, incorporated in the repair template, suggests that the template is not used during correction attempts. In only 29% (2/7) and 14% (1/7) of the ‘corrected embryos’, respectively long (>18Mb) or medium width LOH (4Mb) was observed through STR analysis. SNP analysis in closer proximity showed in 71% (5/7) of the embryos LOH, even in the absence of LOH through STR, suggesting also the occurrence of short width LOH. These results will be studied in more detail before definitive conclusions can be made. Chromosomal LOH will be studied by ddRADseq. Limitations, reasons for caution The occurrence of mosaicism and LOH might complicate the use of traditional CRISPR/Cas9 in human embryos and should be studied in detail to draw definite conclusions on its potential future use. To this end, genomic data have been produced from both individual blastomeres and whole-embryos which will be further analyzed. Wider implications of the findings Our findings demonstrate caution to use CRISPR/Cas9 to correct mutations in the germ line. They seem to contradict other reports that show predominant lack of mosaicism and presence of long width LOH. A deeper evaluation will be undertaken to define the length and type of LOH in this study. Trial registration number Not Applicable

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DOMINANCE MODIFIERS IN NEUROSPORA CRASSA: PHENOCOPY SELECTION AND INFLUENCE ON CERTAIN ASCUS MUTANTS

Genetics ◽

10.1093/genetics/71.2.233 ◽

1972 ◽

Vol 71 (2) ◽

pp. 233-245

Author(s):

Peter J Russell ◽

Adrian M Srb

Keyword(s):

Neurospora Crassa ◽

Mutant Allele ◽

Detectable Effect ◽

Wild Type Allele ◽

Wild Type ◽

Group V ◽

Type Allele ◽

Dominance Relationships ◽

Mutant Strains ◽

Dominant Peak

ABSTRACT When homozygous in zygotes, mutant alleles at the peak locus in linkage group V of Neurospora crassa initiate aberrant asci that are nonlinear, in contrast to the linear asci characteristic of wild type. Most mutant alleles are recessive, inasmuch as crosses of the mutant strains with wild type give linear asci. However, five different mutant alleles, when heterozygous with the wild-type allele, act in varying degrees as zygote dominants, initiating both linear and nonlinear asci, the relative proportions depending on the allele. Five modifiers that act on the dominance relationships of at least one of the five possible heterozygotes of a dominant peak and its wild-type allele have been characterized, four of them having been obtained by selection directed against a phenocopy of these mutants induced by treatment of wild type with l-sorbose. The pattern of modifier specificity observed among the various dominant peak heterozygotes indicates that the phenotypic effects are produced by a complex relationship between the modifiers and the dominant peak alleles in relation to their wild-type allele. In all but two cases the direction of modification, where present, is towards decreasing the dominance of the mutant allele in the heterozygote, evidenced by an increase in the percentage of linear asci when compared with control data. The modifiers exert their maximum modification when they themselves are heterozygous with their wild-type alleles and when the dominant peak allele is heterozygous with its wild-type allele. No modification occurs when heterozygous modifiers are included in zygotes homozygous for a dominant peak allele, reinforcing the notion that the modifiers act on the dominance relationship existent between a dominant peak allele and its wild-type allele, rather than influencing some activity of the mutant allele itself. The modifiers have no detectable effect of their own on ascus morphology, since homozygous modifier zygotes initiate entirely linear asci when only wild-type alleles of peak are present in the zygotes. Their only detectable effect, other than dominance modification, appears to be in conferring sorbose resistance to the mycelium. The modifiers are unlinked to the peak locus, and, except for two of them, they are nonallelic.

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