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Biomedical subjects

Craig A Jones

Publications and source records attributed to Craig A Jones.

18 recordsLinked to original sources
PubMed2006

Improvement of asthma control with omalizumab in 2 obese pediatric asthma patients.

BACKGROUND: Dosing of the anti-IgE antibody (omalizumab) in the treatment of allergic asthma depends on serum IgE concentration and body weight. It is unclear whether omalizumab is therapeutically effective in obese patients with difficult-to-control asthma whose body weights fall outside dosing guidelines. OBJECTIVE: To report the efficacy of omalizumab in 2 obese pediatric patients with severe persistent asthma whose high body weights placed them outside current dosing guidelines. METHODS: Omalizumab was given at maximum doses to both patients. Standard asthma therapy was continued throughout the treatment period. Multiple parameters of asthma disease activity were followed at every health encounter for approximately 1 year during and before omalizumab treatment. These parameters included asthma disease severity and activity, pulmonary functions, daily inhaled corticosteroid requirements, systemic steroid rescue, and urgent care and emergency department visits and hospitalizations. RESULTS: While receiving omalizumab, both patients had improvements in asthma disease activity and reduction in systemic steroid requirements. One patient required fewer daily inhaled corticosteroids and achieved total asthma control. CONCLUSION: Patients with difficult-to-treat asthma may benefit from receiving omalizumab even if they fall outside current dosing guidelines because of high body weight.

Administration, Inhalation↗
PubMed2006

Validation of a single survey that can be used for case identification and assessment of asthma control: the Breathmobile Program.

BACKGROUND: Underdiagnosis of asthma and underrecognition of disease severity in lower socioeconomic populations continue to be significant health care concerns despite national efforts to better educate health care providers. OBJECTIVE: To validate a 1-page survey as a point-in-time tool identifying uncontrolled vs controlled asthma and moderate-to-severe disease activity in an urban, lower-socioeconomic pediatric population. METHODS: A previously validated survey (the Breathmobile Case Identification Survey) was evaluated as a point-in-time tool for identifying children with poorly controlled disease. Clinical validation was achieved in children (n = 1,826) presenting to a school-based asthma program for either an initial (n = 666) or a follow-up (n = 1,170) visit. Responses were compared with a comprehensive evaluation by a physician specialist as the gold standard. Response patterns were used to construct multimodel tiered scoring algorithms for baseline and follow-up visits that identify children with uncontrolled asthma, and children are likely to have moderate-to-severe disease activity at that time. RESULTS: Surveys scored using the developed algorithms identified children with uncontrolled asthma (sensitivity: baseline, 77.0%; follow-up, 71.6%; specificity: baseline, 72.7%; follow-up, 71.5%) and detected moderate-to-severe disease activity (sensitivity: baseline, 69.2%; follow-up, 77.4%; specificity: baseline, 70.2%; follow-up, 70.3%). CONCLUSIONS: The Breathmobile Case Identification Survey can be used in lower-socioeconomic, urban populations as a point-in-time tool for identifying children with uncontrolled vs controlled asthma and moderate-to-severe disease activity.

Adolescent↗
PubMed2006

Symptom profile and asthma control in school-aged children.

BACKGROUND: Children with poorly controlled asthma are at high risk of airway remodeling, sleep disruption, school absenteeism, and limited participation in activities. OBJECTIVE: To determine asthma prevalence and characterize disease severity and burden in school-aged children. METHOD: A case-finding study was conducted via a multiple-choice questionnaire and asthma algorithm. Items used for analysis include physician diagnosis of asthma, symptom severity, and health care utilization. The chi2 test was used to determine the significance of differences among cases. Logistic regression was used to evaluate the association of patient factors and asthma indicators. RESULTS: Of the 5,417 children surveyed, 1,341 (25%) were classified as being at risk of asthma. Of these asthma cases, 55% were positive by diagnosis and algorithm (active), 10% were positive per algorithm alone (suspected), and 35% were positive per diagnosis alone (nonactive). Only 14% of all asthma cases reported experiencing no respiratory symptoms (< 1% active, 2% suspected, and 40% nonactive) compared with 75% of noncases. Also, 75% of noncases reported never missing school compared with 19%, 33%, and 54% of active, suspected, and nonactive asthma cases. African American race, Medicaid enrollment, and male sex were independent predictors of asthma risk. Similarly, African American race, Medicaid enrollment, age, and persistent asthma were independent predictors of emergency department use among asthma cases. DISCUSSION: Prevalence of active symptoms suggestive of poor asthma control was extremely high among urban, minority children enrolled in Arkansas' largest public school district. Poor asthma control greatly affects quality of life, including school attendance and performance. Interventions should raise expectations and emphasize the importance of achieving asthma control.

Adolescent↗
PubMed2006

School-based asthma case finding: the Arkansas experience.

This population-based case-finding study sought to determine asthma prevalence and characterize disease severity and burden among school-aged children in the Little Rock School District. Asthma cases were identified by validated algorithm and parental report of asthma diagnosis. The overall response rate was low. Among schools with greater than 50% response rate, prevalence of physician-diagnosed asthma was comparable to other studies in public school settings. Prevalence of symptoms in cases identified as current is suggestive of poor asthma control. Improved systems for case identification of children with asthma are needed in schools. Efforts should focus on strategies to improve asthma control.

Adolescent↗
PubMed2005

Activation of the rat renin promoter by HOXD10.PBX1b.PREP1, Ets-1, and the intracellular domain of notch.

Renin gene expression is subject to complex developmental and tissue-specific regulation. A comparison of the promoter sequences of the human, rat, and mouse renin genes has revealed a highly conserved sequence homologous to the DNA recognition sequence for CBF1 (CSL/RBP-Jkappa/Su(H)/LAG1/RBPSUH). Electrophoretic mobility shift assays document that As4.1 cell nuclear protein complex binding to the putative rat renin CBF1-binding site (-175 to -168 bp) contains CBF1. Transient transfection analyses in COS-7 cells further document that a CBF1-VP16 fusion protein and the intracellular domain of Notch1 robustly activate a promoter containing multiple copies of the rat renin CBF1-binding site. An Ets-binding site (-143 to -138 bp) has also been identified in the rat renin promoter by sequence comparisons and electrophoretic mobility shift assays. Transcription factor Ets-1 is capable of activating the rat renin promoter through the Ets-binding site. Mutation of the CBF-binding site significantly increases transcriptional activity of the rat renin promoter in Calu-6 and COS-7 cells but not in As4.1 cells, whereas mutation of the Ets-binding site reduces promoter activity of the rat renin gene in all three cell lines. Finally, we show that the intracellular domain of Notch1, Ets-1, and HOXD10.PBX1b.PREP1 activate the rat renin promoter cooperatively in COS-7 cells. These results strongly suggest that the renin gene is a downstream target of the Notch signaling pathway.

Animals↗
PubMed2005

School absenteeism in children with asthma in a Los Angeles inner city school.

OBJECTIVE: To investigate the pattern of school absenteeism in asthmatic children within a Los Angeles inner city school. STUDY DESIGN: Five hundred twenty-eight students of predominant Hispanic ethnicity, from a Los Angeles inner city school were divided into 3 groups: known asthma, high probability of asthma, and low probability of asthma using a previously validated instrument. Attendance records of these students were analyzed to determine total and respiratory absences over a year. School records were compared to the corresponding answers on 513 surveys to determine the accuracy of parental responses in regard to their children's absenteeism. RESULTS: Children with known asthma missed on average 2 more days of school than children with low probability of asthma and high probability of asthma. This was only significant in the younger age groups. Survey responses were found to have a 45.6% agreement with school attendance records. Underestimation occurred more often when school-recorded absentee rates were highest. Overestimation occurred more by parents of children with known asthma or a high probability of asthma. CONCLUSION: In a Los Angeles inner city population, younger children with known asthma miss more days of school than those with no asthma. Survey-reported absenteeism is less accurate than school attendance records.

Absenteeism↗
PubMed2005

The Breathmobile Program: structure, implementation, and evolution of a large-scale, urban, pediatric asthma disease management program.

Despite more than a decade of education and research-oriented intervention programs, inner city children with asthma continue to engage in episodic "rescue" patterns of healthcare and experience a disproportionate level of morbidity. The aim of this study was to establish and evaluate a sustainable community-wide pediatric asthma disease management program designed to shift inner city children in Los Angeles from acute episodic care to regular preventive care in accordance with national standards. In 1995 the Southern California Chapter of the Asthma and Allergy Foundation of America (AAFA), the Los Angeles County Department of Health Services (LAC DHS), and the Los Angeles Unified School District (LAUSD) established an agreement to initiate and sustain the Breathmobile Program. This program includes automated case identification, mobile school-based clinics, and highly structured clinical encounters supported by an advanced information technology solution. Interdisciplinary teams of asthma care specialists provide regular and ongoing care to children at school and county clinic sites over a wide geographic area of urban Los Angeles. Each team operates in a specially equipped mobile clinic (Breathmobile), efficiently moving a structured healthcare process to school and county clinic sites with large numbers of children. Demographic, clinical, and participation data is tracked carefully in an electronic medical record system. Program operations, clinical oversight, and patient tracking are centralized at a care coordination center. Clinical operations and methods have been replicated in fixed specialty clinic sites at the Los Angeles County + University of Southern California Medical Center. Clinical and process measures are regularly evaluated to assure quality, plan iterative improvement, and support evidence-based care. Four Breathmobiles deliver ongoing care at more than 90 school sites. The program has engaged over five thousand patients and their families in a continuity care model that has demonstrated efficacy over usual episodic care. More than 90% of patients in all asthma severity categories achieved clinical control of asthma with significant reductions in inpatient (IP) and emergency department (ED) use. On February 14, 2002, the program became the first program in the United States to receive the award of disease-specific care certification by the Joint Commission on Accreditation of Healthcare Organizations (JCAHO). Proper design and resource allocation can sustain a school-based community-wide pediatric asthma disease management program and shift a population of inner city children from acute episodic care to routine preventive care in accordance with national standards. An evidence-based approach to evaluating and maintaining quality, coupled with stratified care delivery, can assure the efficient use of safety net healthcare resources.

Adolescent↗
PubMed2005

Discussing the COSTS of asthma: controlling outcomes, symptoms, and treatment strategies.

Asthma is a very common chronic disease characterized by clinically recurrent episodes of wheezing, dyspnea, chest tightness, and coughing. Both genetic and environmental factors appear to play major roles in the etiology of asthma. The economic and social costs of asthma are varied and far reaching. Patients suffer a significant decline in quality of life, and managed care organizations have become increasingly aware of its impact on health care expenditures. This article explores current treatment therapies for asthma, the challenges of managing asthma effectively, and strategies for improving outcomes in patients with asthma.

Asthma↗
PubMed2004

Enhancer-dependent inhibition of mouse renin transcription by inflammatory cytokines.

Inflammatory cytokines have been shown to inhibit renin gene expression in the kidney in vivo and the kidney tumor-derived As4.1 cell line. In this report, we show that cytokines oncostatin M (OSM), IL-6, and IL-1beta inhibit transcriptional activity associated with 4.1 kb of the mouse renin 5'-flanking sequence in As4.1 cells. The 242-bp enhancer (-2866 to -2625 bp) is sufficient to mediate the observed inhibitory effects. Sequences within the enhancer required for inhibition by each of these cytokines have been determined by deletional and mutational analysis. Results indicate that a 39-bp region (CEC) containing a cAMP-responsive element, an E-box, and a steroid receptor-binding site, previously identified as the most critical elements for enhancer activity, is sufficient for the inhibition induced by IL-1beta. However, mutation of each of the three component sites does not abolish the inhibition by IL-1beta, suggesting that the target(s) of cytokine action may not be the transcription factors binding directly to these sites. This CEC region is also critical, but not sufficient, for the inhibition mediated by OSM and IL-6. These data suggest that the direct target of the associated cytokines may be coactivators interacting with transcription factors binding at the enhancer. Finally, we show that OSM treatment caused a 17-fold increase in promoter activity when only 2,625 bp of the Ren-1(c) flanking sequence were tested, in which the enhancer is not present. Three regions including -2625 to -1217 bp, the HOX.PBX binding site at -60 bp, and -59 to +6 bp have been found to contribute to this induction.

Animals↗
PubMed2004

Genomic characterization and expression analysis of the first nonmammalian renin genes from zebrafish and pufferfish.

Renin is a key enzyme in the renin-angiotensin system (RAS), a pathway which plays an important physiological role in blood pressure and electrolyte homeostasis. The origin of the RAS is believed to have accompanied early evolution of vertebrates. However, renin genes have so far only been unequivocally identified in mammals. Whether or not a bona fide renin gene exists in nonmammalian vertebrates has been an intriguing question of physiological and evolutionary interest. Using a genomic analytical approach, we identified renin genes in two nonmammalian vertebrates, zebrafish (Danio rerio) and pufferfish (Takifugu rubripes). Phylogenetic analysis demonstrates that the predicted fish renins cluster together with mammalian renins to form a distinct subclass of vertebrate aspartyl proteases. RT-PCR results confirm generation of the predicted zebrafish mRNA and its expression in association with the opisthonephric kidney of adult zebrafish. Comparative in situ hybridization analysis of wild-type and developmental mutants indicates that renin expression is first detected bilaterally in cells of the interrenal primordia at 24 h postfertilization, which subsequently migrate to lie adjacent to, but distinct from, the glomerulus of the developing pronephric kidney. Our report provides the first molecular evidence for the existence of renin genes in lower vertebrates. The observation that the earliest renin-expressing cells, arising during ontogeny of this teleost vertebrate, are of adrenocortical lineage raises an interesting hypothesis as regards the origin of renin-expressing cells in the metanephric kidney of higher vertebrates.

Adrenal Glands↗
PubMed2004

Identification of a novel region in the proximal promoter of the mouse renin gene critical for expression.

An enhancer at -2.6 kb and a HOX.PBX-binding site at -60 bp have been demonstrated to be critical to expression of the mouse renin gene (Ren-1(c)) in As4.1 cells. In this report, we show that a region (-197 to -70) immediately 5' to the HOX.PBX-binding site is also critical for Ren-1(c) expression. Deletion of this region in a construct containing 4.1 kb of the Ren-1(c) 5'-flanking sequence resulted in a 99% reduction in Ren-1(c) promoter activity in As4.1 cells, suggesting the pivotal role for the region in the regulation of the mouse renin gene. Electrophoretic mobility shift and supershift assays have identified two nuclear factor I-binding sites and a Sp1/Sp3-binding site within the distal portion of the region (-197 to -103). Mutation of these three sites caused a 90% decrease in Ren-1(c) promoter activity. Mutational analysis and electrophoretic mobility shift assays have also identified three additional transcription factor-binding sites within the region from -103 to -69, each of which contributes to high-level expression of Ren-1(c) in As4.1 cells. Finally, we have shown that the Ren-1(c) enhancer is the target for endothelin-1 (ET-1)-induced inhibition of Ren-1(c) expression and the transcription factor-binding sites in the proximal promoter are required for the maximal ET-1 inhibitory effect.

Animals↗
PubMed2004

Identification of preschool children with asthma from low-income families in Los Angeles, CA.

BACKGROUND: Few studies have addressed asthma screening in the preschool age group. Early asthma recognition and intervention in preschool children may reduce costs related to unscheduled medical care and missed school and work. OBJECTIVE: To facilitate an early recognition and referral process for asthma in a preschool education program in Los Angeles, CA. METHODS: We administered a 7-question survey to parents and guardians of children aged 12 months to 6 years in the prekindergarten program of a large school district in Southern California. English and Spanish survey questions addressed health care use, school absenteeism, and asthma symptoms. Postsurvey reports to parents recommended clinical evaluation of children who had probable asthma. RESULTS: Of the 609 surveys returned from 8 centers (> or = 80% survey return rate), 12% were positive for probable asthma and only 5.4% of these cases had been previously diagnosed. Of the 12% found to have a high probability of asthma, 3 independent factors were associated with a lower likelihood of prior asthma diagnosis: Hispanic descent; Spanish speaking; and medicine use 2 or more times per week for symptoms such as cough, chest tightness, trouble breathing, or wheezing. Symptoms at play, during the day, and at night were noted in 35% to 44% of the preschoolers. Cough was the most frequently reported symptom (71.9%, n = 424). CONCLUSIONS: A school-based screening process in an early education program can help identify preschool children with a high probability of asthma and offer a basis for early recognition and intervention.

Asthma↗
PubMed2004

A school-based case identification process for identifying inner city children with asthma: the Breathmobile program.

BACKGROUND: Striking increases in the prevalence and morbidity of asthma among inner city children have been documented. OBJECTIVE: To establish and evaluate a large-scale, school-based case-detection process designed to efficiently and reliably identify inner city children with asthma. METHODS: A bilingual, seven-question, self-administered, parental asthma screening survey was developed. Clinical validation was achieved in a sample of 675 consecutive parents bringing a child to the school-based Breathmobile Program for initial evaluation, using a comprehensive evaluation by a physician specialist (ie, allergist) as the standard. Survey response patterns were used to construct a novel seven-model, tiered scoring algorithm and an abbreviated algorithm that predict the probability of a child being clinically classified as "yes asthma" or "no asthma." A systematic survey distribution process administered by a single coordinator was developed, and the impact of a classroom-oriented incentive offering a 25 dollars school supply gift certificate for survey return rates of >or= 80% was evaluated. RESULTS: A total of 636 parents provided one or more survey responses and information sufficient for clinical classification. The scoring algorithm correctly identified children with asthma (>or= 80% probability) with a sensitivity of 86.5%, a specificity of 83.6%, and a misclassification rate of 14.3% (91 of 636 children). The sensitivity for identifying persistent asthma was 91.3%. Asthma prevalence estimates derived using survey results from a larger sampling of the general population were similar to rates previously reported for comparable populations. The inclusion of an inexpensive incentive increased the median survey return rates from 35.3 to 65% (z= -11.9; p <.001). The screening process has been used to conduct 27,526 surveys at inner city schools. CONCLUSIONS: The Breathmobile case-detection process offers a validated, comprehensive, large-scale method with which to identify children with asthma at their school sites.

Adolescent↗
PubMed2004

Predictive value of a cross-cultural asthma case-detection tool in an elementary school population.

OBJECTIVE: Bronchial asthma, which affects approximately 5 million US children, is vastly underdiagnosed and treated, particularly among minorities and those of low socioeconomic status. Because current methods of detecting those at greatest risk of asthma in a multicultural setting appear inadequate, we assessed the validity and reliability of a new asthma questionnaire across 3 dominant cultures in Orange County, California (white, Hispanic, and Vietnamese). METHODS: Children in grades 1, 3, and 5 and their families, in 3 different schools representative of these major ethnic groups, were randomly selected to participate in the validation process. Two schools with low socioeconomic status and dominant Hispanic or Vietnamese minorities were designated inner-city schools, whereas the third school was a suburban school with predominately white students. Participants completed a 7-question, 11-element questionnaire in their primary language, followed by an asthma evaluation (history, physical examination, and spirometry) by an asthma specialist (who was blinded with respect to the results of the questionnaire), at their respective schools. The physician then made a determination regarding the presence and severity (according to National Institutes of Health guidelines) of asthma. Several weeks later, the entire student body was asked to complete the questionnaire at home and return it to school for analysis. Validation of each item was evaluated for sensitivity, specificity, and positive and negative predictive values, and application of univariate analyses provided an estimated probability of an asthma diagnosis by the asthma specialist. A "best-fit" algorithm was determined with all 11 elements, if possible, and an abbreviated algorithm that selected the fewest-question combination that yielded the best asthma predictability was established. Reliability was established with the percent agreement between the 2 questionnaires and the kappa statistic. RESULTS: Of the 401 children/families who participated in the validation analysis, 45% were Hispanic, 22% white, 19% Vietnamese, and 15% other. The overall prevalence of asthma specialist-diagnosed asthma was 28%, with 65% of cases being graded as intermittent and 35% as persistent. Sixty-two percent of the children had not been previously diagnosed with asthma. There were no significant differences among cultures in sensitivity or specificity for any of the individual questions or the complete or abbreviated algorithms. The abbreviated algorithm with 3 questions, ie, question 1 (asthma in the past 2 years), question 4 (cough, chest tightness, trouble breathing, or wheezing with exercise), and question 6 (same symptoms in the morning or day in the past 4 weeks) yielded comparable sensitivity and specificity for the complete algorithm in all groups. The abbreviated algorithm had >86% predictability in detecting children with persistent asthma and 56% predictability in detecting children with intermittent asthma. Reliability was also excellent, with percent agreement usually > 80% and kappa values of >.70. CONCLUSIONS: This asthma detection tool has been shown to be suitable for detecting persistent asthma in a multicultural inner-city population, as well as in a suburban setting. An abbreviated algorithm with 3 questions and >80% predictability in detecting persistent asthma seems ideal for evaluating large numbers of school-aged children. The school setting is an excellent site for identifying children with asthma. Although there is concern that subjects detected in the school setting might not have access to ongoing medical care, case detection is an important first step that could lead to earlier diagnosis and treatment. Reducing the barriers to good care in inner-city environments is the next step.

Algorithms↗
PubMed2003

Evaluation of shedding of bovine herpesvirus 1, bovine viral diarrhea virus 1, and bovine viral diarrhea virus 2 after vaccination of calves with a multivalent modified-live virus vaccine.

OBJECTIVE: To determine whether bovine herpesvirus 1 (BHV1), bovine viral diarrhea (BVDV) virus 1 (BVDV1), or BVDV 2 (BVDV2) were shed after parenteral administration of a multivalent modified-live virus vaccine. DESIGN: Prospective study. ANIMALS: 28 healthy beef calves and 4 healthy pregnant beef cows. PROCEDURE: A commercially available modified-live virus multivalent vaccine was administered to steers and heifers (n = 18) that were seronegative to BHV1, BVDV1, and BVDV2. Four seronegative pregnant control cows were held in contact with the vaccinated calves for 103 days. Unvaccinated calves (n = 10) were held as controls in a separate double-fenced pen. Seroconversion was monitored by determining serum neutralization titers after vaccination. Viral shedding and viremia were assessed via analysis of nasal swab specimens and blood by use of polymerase chain reaction (PCR) and reverse transcriptase-PCR assays and virus isolation. RESULTS: A transient BVDV1 viremia was detected in most vaccinated calves 3 to 10 days after vaccination. All vaccinated calves seroconverted to BVDV1 and BVDV2. Seventeen of 18 vaccinated calves seroconverted to BHV1. Viral shedding was not detected in the vaccinated calves. All control cattle remained seronegative to BHV1, BVDV1, and BVDV2 throughout the study. CONCLUSIONS AND CLINICAL RELEVANCE: Shedding of BHV1, BVDV1, and BVDV2 after vaccination was either nonexistent or undetected and did not result in transmission of BHV1, BVDV1, or BVDV2 vaccine viruses to pregnant contact control cows.

Animals↗
PubMed2003

Regulation of renin enhancer activity by nuclear factor I and Sp1/Sp3.

Transcription of the mouse Ren-1(c) gene in kidney tumor-derived As4.1 cells, which express high levels of renin mRNA, is dependent on a proximal promoter element and a 242-bp enhancer region located 2.6 kb upstream of the transcription start site. We showed previously that the enhancer contains a cAMP responsive element (CRE) and an E-box. Mutation of either element resulted in almost complete loss of the Ren-1(c) expression. In this report we show that there are additional transcription factor-binding sites within the Ren-1(c) enhancer contributing to the enhancer activity. Electrophoretic mobility shift and supershift assays have identified four nuclear factor I (NFI)-binding sites, an Sp1/Sp3 site and an unidentified transcription factor-binding site (Ei) located upstream of the CRE and E-box. Mutation of the Sp1/Sp3 site or Ei reduced Ren-1(c) expression by 40% or 30%, respectively, while mutations of four NFI-binding sites resulted in an 89% decrease in expression. Thus, these protein-DNA interaction sites are essential for transcription of mouse renin genes. There are four homologous NFI genes (NFI-A, -B, -C and -X) in vertebrates and multiple alternatively spliced isoforms from each gene. Real-time reverse transcriptase-polymerase chain reaction (RT-PCR) assays have demonstrated that NFI-X is the predominant NFI mRNA expressed in As4.1 cells. Direct study of involvement of NFI-X in regulation of renin genes is underway.

Base Sequence↗
PubMed2002

Rapid localization of transgenes in mouse chromosomes with a combined Spectral Karyotyping/FISH technique.

We explored the feasibility of combined Spectral Karyotyping (SKY) and Fluorescence In Situ Hybridization (FISH) as means to rapidly map a chromosomally integrated renin/green fluorescent protein (GFP) fusion gene construct (Ren-GFP) in the transgenic mouse, Tg(Ren-GFP)1Kwg. A sequential hybridization with SKY probes followed by FISH gave consistently satisfactory results, demonstrating that multiple copies of the Ren-GFP transgene in this transgenic mouse line are integrated into a single chromosomal site of Chromosome (Chr) 4, most probably in the juxta-centromeric euchromatic region consisting of the A2-A3 domain. Chr 4 as a sole carrier of the transgene also was confirmed by co-hybridization to p1 BAC clone DNA containing telomeric sequences specific for mouse Chr 4 and the Ren-GFP construct in pGEM4Z vector. The hemizygosity of the Ren-GFP transgene is maintained not only in bone marrow cells, but also in lung cells proliferating in vitro, indicating that stable integration of the Ren-GFP transgene into chromosomal DNA was established at a very early embryonic stage. We conclude that the SKY/FISH technique is a reliable and facile method for establishing the integration site of a transgene. As such, this protocol has obvious advantages over traditional backcross methods in terms of time, cost and labor for determining the chromosomal location of transgenes.

Animals↗