Aims/hypothesis The escalating rate of childhood obesity is a public health concern worldwide, with children in certain ethnic groups being disproportionately affected. Our objective was to examine the joint effects of pre-pregnancy adiposity, pregnancy weight gain and gestational diabetes (GDM) in relation to excess fetal growth and to identify susceptible races or ethnic populations. Methods The risk for delivery of a large-for-gestational-age (LGA) infant, specific to race and fetal sex, was evaluated in 105,985 pregnancies in the Consortium on Safe Labor from [2002][2003][2004][2005][2006][2007][2008]. Generalised estimating equations were used to estimate the risk for delivery of LGA infants. Joint effects were employed to evaluate the interplay of three risk factors.Models were stratified by racial group considering one, two or three factors (i.e. pre-pregnancy adiposity, pregnancy weight gain and GDM, with 0 factors as the reference group). Results Greater pre-pregnancy adiposity, pregnancy weight gain and GDM were independently associated with increased risk of giving birth to an LGA infant across all races (except GDM among non-Hispanic whites), in both underweight and normal-weight women. Among non-Hispanic white, non-Hispanic black and Hispanic women, the threefactor joint effect was associated with substantially increased odds of LGA (OR [95% CI] 11. 27 [8.40, 15.11], 7.09 [4.81, 10.45] and 10.19 [6.84, 15.19], respectively). However, for Asian women the joint effect of all three factors (OR [95% CI] 5.14 [2.11, 12.50]) was approximately the same as any of the two factors. Conclusions/interpretation GDM, pre-pregnancy obesity and excessive pregnancy weight gain were jointly associated with elevated risk of giving birth to an LGA infant and the effects varied by race. This suggests that those involved in public health efforts aimed at preventing LGA deliveries should consider variations in racial groups when devising effective strategies.
The purpose of this study was to investigate the effects of maternal gestational diabetes mellitus (GDM) and breast feeding on childhood overweight and obesity in a mainland Chinese population. The incidence of and factors associated with overweight and obesity were compared between children of mothers with (n=1068) and without (n=1756) GDM. The independent roles of the associated factors were examined by multiple logistic regression analysis. The incidence of overweight was higher (16.6 v. 12.6%, P=0.002) in the GDM group, but that of obesity was not different (10.7 v. 12.0%, P=0.315). At age 1-2 and 2-5 years, no difference in overweight (11.0 v. 12.0%, P=0.917, and 15.7 v. 14.6%, P=0.693, respectively) was found, while obesity (8.0 v. 13.6%, P=0.019, and 8.4 v. 13.4%, P=0.014, respectively) was less frequent in the GDM offspring. At age 5-10 years, increased overweight (22.2 v. 12.1%, P<0.001) and obesity (15.9 v. 9.0%, P=0.001) were found in the GDM group, which was associated with maternal obesity, being born large-for-gestational age, male gender and formula feeding. After adjusting for confounding factors, GDM remained an independent determinant of offspring overweight and obesity (aOR 2.28, 95% CI 1.61-3.22), suggesting that the effects of GDM were independent of breast feeding, as well as of maternal obesity and birth size.
The polymerase chain reaction is a powerful technique used to amplify nucleic acids in vitro. The reaction produces linear products, and as of yet, closed circular products have not been possible. Since the replicatively competent form of many DNA molecules is the closed circular form, it would be adventitious to amplify closed circular DNA as closed circular molecules. Until now, these molecules could only be amplified in vivo in appropriate host cells. Here, we describe an in vitro procedure, ligation-during-amplification (LDA), for selective amplification of closed circular DNA using sequence-specific primers. LDA is useful for sitedirected mutagenesis, mutation detection, DNA modification, DNA library screening and circular DNA production.The polymerase chain reaction (PCR) (1-3) is a powerful technique for in vitro amplification of nucleic acids. Although circular and linear nucleic acids can serve as templates for PCR, the resulting products have always been linear molecules. Until now, closed circular DNA, the replicatively competent form of many DNA molecules, could only be amplified in appropriate host cells. Here, we describe and demonstrate an in vitro procedure, ligation-during-amplification (LDA), for selective amplification of closed circular DNA using sequence-specific primers. The essence of LDA is the inclusion of a thermostable DNA ligase in a PCR reaction that uses a closed circular DNA as template. After a primer is fully extended on the circular template, the ligase closes the gap to form a double-stranded DNA. Following thermal denaturation, the two circular DNA strands serve as templates for the next round of extension and ligation. Through thermal cycling, closed circular DNA is amplified exponentially.To demonstrate the feasibility of using LDA to generate and amplify closed circular DNA, two 5′ phosphorylated primers (16 and 17 nt long) were used to mutate and amplify a circular plasmid of 1990 bp. The primers are complementary to different strands of the plasmid in an inward orientation (Fig. 1A). One possesses a single G to A mismatch on an HphI site in the plasmid. The reaction mixture (50 µl) contained 10 ng of native plasmid, 10 pmol of each primer, 10 nmol of dNTPs, 5 nmol of ATP, 2.5 U Pfu DNA polymerase (Stratagene, La Jolla, CA), 4 U Pfu DNA ligase (Stratagene), in 1 cloned pfu DNA polymerase reaction buffer consisting of 20 mM Tris-HCl (pH 8.8), 10 mM KCl, 10 mM (NH 4 ) 2 SO 4 , 2 mM MgSO 4 , 1% Triton X-100 and 100 µg/ml BSA. The mixture was pre-incubated at 70_C for 10 min allowing the ligase to repair any nicks in the template. It was then subjected to thermal cycling at 95_C for 1 s (denaturation), 50_C for 1 s (annealing), 72_C for 4 min (extension), 95_C for 1 s (denaturation) and 72_C for 4 min (annealing, extension and ligation) for 20 cycles. As a control, a PCR was performed under similar conditions, except that the DNA ligase was not included. The reaction mixtures were analyzed directly by electrophoresis into 1% agarose gels followed by staining with ethid...
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