With the development of plant genetic engineering, many transformation methods can be used to transfer heterogeneous genes into plants to develop genetic crops. However, a lot of research results have shown that transgene expression remains largely unpredictable and there is great variation of expression level in different transgenic plant lines. Plant genetic engineering research is reviewed in the present paper. We analysed the reasons why low efficiency of heterogeneous gene expression has happened in transgenic plants in terms of DNA modification, localization of proteins and methods of transformation used. Some strategies for improving heterogeneous gene expression in transgenic plants are also discussed.

The microsatellite, or short sequence repeat (SSR), is a powerful genetic marker, useful in many areas of fish genetics and breeding. Polymorphic microsatellite loci have been frequently applied to the analysis of genetic diversity, population genetic structure, and genomic mapping. These co-dominant markers have also been applied to the classification and systematics, parentage identification, germplasm conservation, and breeding programme of food fish.

Pancreases obtained from 1–9-day-old rabbits were digested with 1 mg/ml ?? collagenase and cultivated with low-glucose Dulbecco's modified Eagle's medium (DMEM). The cells were subcultured almost until confluence. They were identified as pancreatic stem/progenitor cells using two methods: an immunohistochemical staining technique and the response of glucose-stimulated insulin secretion after β-cell induction. Results showed that the cells of rabbits acquired strong proliferation ability, and expressed the markers of pancreatic stem/progenitor cells: PDX-1, CK19 and nestin. After β-cell induction for 2 days, 20% of cells were dithizone (DTZ) positive, rapidly increasing to 80% after 6 days. Insulin secretion was stimulated by a high glucose concentration (26 mmol/l). These powerful tests demonstrated that isolated and cultivated cells in rabbits were pancreatic stem/progenitor cells. This result has not yet been reported elsewhere.

The effects of hormone additions at various stages and different basic media, with or without serum, on porcine oocyte maturation in vitro were studied. The results showed that the rate of maturation was not significantly different with three different stages of hormone supplement; the rate of maturation on modified TCM199 medium (54.01%) was higher than that on TCM199 (46.16%) and (47.14%), but these differences were not significant; and the rate of maturation on serum-free medium (67.10%) was significantly higher than that on medium plus serum (52.22%). Therefore, modifed tissue culture medium 199 (mTCM199)+10I U/ml pregnant mare serum gonadotrophin (PMSG)+10I U/ml human chorionic gonadotrophin (hCG)+2.5 IU/ml follicle stimulating hormone (FSH) was a suitable medium for culture of porcine oocytes in vitro, and the rate of maturation was 67.10%.

The genetic variations of the 5′-upstream region and the second intron of the porcine H-FABP gene were investigated by polymerase chain reaction–restriction fragment length polymorphism (PCR-RFLP) in 256 pigs, including Duroc, Large White, Landrace, Neijiang, Rongchang, Bamei pig, Hanjiang Black, Hanzhong White, and wild pigs. The effect of the H-FABP gene on intramuscular fat (IMF) content was analysed by the least square method. Results showed a HinfI-RFLP in these eight pig breeds and wild pigs, among which Large White, Bamei pig, Hanjiang Black, Hanzhong White and wild pigs presented low polymorphism, while the other breeds had intermediate polymorphism. There were no HaeIII or MspI-RFLPs in the four Chinese local pig breeds tested, but Duroc, Landrace, Large White, Hanzhong White and wild pig displayed polymorphism. Landrace, Large White and wild pigs had low levels of HaeIII- and MspI-RFLP, while the others had intermediate polymorphism. H-FABP genotypes significantly affected IMF content (P<0.05). IMF content according to H-FABP genotypes were HH>Hh>hh, DD<Dd<dd, AA<Aa<aa. The genetic effect values were 3.95 (HH), 3.48 (Hh), 3.23 (hh), 2.33 (DD), 2.55 (Dd), 2.97 (dd), 2.34 (AA), 2.76 (Aa) and 3.01 (aa). Results suggest that porcine meat quality may be improved by increasing the frequency of genotype aa-dd-HH in pig breeds.

Sox9 is one of the important genes related to sex differentiation and determination. Two partial cDNAs encoding sox9 were derived from brain, testis and ovary of yellow catfish Pelteobagrus fulvidraco using reverse transcriptase-polymerase chain reaction (RT-PCR). Sequence analysis revealed that the ovarian sox9a2 was different from sox9a1 derived from brain and testis. The identity rate of cDNA was 77%, that of amino acids was 84%, and the different nucleotides were distributed in different sites all over the cDNA. This finding confirmed that there were transcripts from two genes. Based on the homology of the amino-acid sequences for sox9 of other animals, the similarity for sox9a1 from yellow catfish and sox9 of other animals averaged 86%. The similarity for sox9a2 reached a mean of 83%. The phylogenetic analysis revealed yellow catfish sox9a1 and sox9a2 were clustered into the fish branch. And yellow catfish sox9a2 was different from the other fish sox9. There was no significant branch including all sox9 derived from testis and brain or including all ovarian sox9. The genetic distance was the same with the results of the traditional taxonomy. RT-PCR analysis indicated that sox9a1 was expressed in male and female yellow catfish forebrain as well as in gonads, whereas sox9a2 was only expressed in ovaries. The function of sox9a2 in female sex formation and ovary development needs to be studied in depth.

Bovine mastitis caused by Streptococcus agalactiae is mainly subclinical and therefore can be diagnosed only in the laboratory. A nested polymerase chain reaction (PCR)-based method for specific, sensitive and rapid detection of S. agalactiae in raw milk was developed. The general streptococci primers, which anneal to conserved areas within the 16S rRNA subunit gene, were used as positive controls. The specificity of S. agalactiae primers is based on various areas within conserved areas of the 16S rRNA genes of S. agalactiae. Results have indicated that the method enables the detection of 1 CFU/ml of S. agalactiae in raw milk after enrichment, followed by DNA extraction using a rapid and simple procedure developed for this purpose, and specific PCR reaction. The method developed can be used efficiently in the early infectious status investigation of S. agalactiae in the dairy herd and in prevention and control of S. agalactiae spread in a herd.

Genetic diversity in 122 accessions of pummelo (Citrus grandis Osbeck) and its related varieties was assessed using simple sequence repeat (SSR) markers. Thirty-one pairs of SSR informative primers generated a total of 335 alleles. The average number of alleles per locus was 9.85. The value of allelic polymorphism information content (PIC) ranged from 0.1939 to 0.9073, with an average of 0.7085 per primer. The 122 accessions of pummelo and its related varieties could be clustered into seven groups by the unweighted pair-group method arithmetic average (UPGMA), in which the 110 pummelo accessions could be divided into 18 subgroups at similarity coefficient of 0.712. These subgroups were mainly composed of the Shatian pummelo variety group, the Wendan variety group and many of the hybrid pummelo groups. The classification method can be used in targeting many varieties in order to widen the genetic background of pummelo.

The genetic variation of isozymes [esterase (EST), superoxide dismutase (SOD), malate dehydrogenase (MDH), malic enzyme (ME) and glucose-6-phosphate dehydrogenase (G6PD)] extracted from two different adult tissues (adductor muscle and gill) of Pteria penguin R. and offspring from two stocks of Pinctada martensii D. were analysed by vertical polyacrylamide gel electrophoresis. The result showed an obvious and stable variation in the isozyme phenotypes in two different pearl oyster species. The SOD and EST isozymes from gill and MDH, ME and G6PDH from adductor muscle were species-specific. The electrophoretograms of these isozymes could be used as markers to differentiate the two pearl oysters. Between inbreeding and hybridized stocks of P. martensii, the electrophoresis patterns of isozymes were alike, but the similarities of electrophoretograms in the inbreeding progenies were higher than in the hybridized ones. The characteristic electrophoretograms of isozymes could be applied as molecular markers in the breeding of pearl oysters.

Primary adipocytes from subcutaneous adipose tissue samples obtained from 7-day-old Yorkshire×Landrace crossbreed piglets were exposed to 0–400 nmol/l of insulin for 48 h. The accumulated triglyceride was measured through Oil Red O staining and the cumulative glycerol released was determined to assess lipolytic activity in adipocytes. Transcription levels of sterol regulatory element binding protein (SREBP)-1c, carbohydrate response element binding protein (ChREBP), acetyl-CoA carboxylase 1 (ACC1), fatty acid synthase (FAS), and hormone-sensitive lipase (HSL) were assessed using reverse transcriptase-polymerase chain reaction (RT-PCR). The results showed that ChREBP and ACC1 mRNA levels were not influenced by insulin alone under low glucose (5 nmol/l). FAS mRNA level was markedly stimulated by all doses of insulin except 200 nmol/l, and SREBP-1c mRNA level increased with 100–300 nmol/l insulin. High insulin doses (300 and 400 nmol/l) increased the HSL mRNA level as well as lipolytic activity.

The Arabidopsis homeotic gene AP2 is essential for floral meristem development and organ specification. In this study, we report the isolation and sequence comparison of the AP2 homologue of Brassica rapa (BAP2). The results showed that the BAP2 DNA sequence was 2138 bp in length and contained nine introns. It shared 90% identity with AP2 in the coding region. The putative BAP2 polypeptide contained a nuclear localization signal (NLS) sequence and two copies of highly conserved AP2 domain, suggesting that BAP2 may function similarly to AP2. Two nucleotide variations were detected in DNA sequences from wild-type and apetalous B. rapa plants at the BAP2 locus, while the putative polypeptides were identical. We propose that BAP2 is not likely to be responsible for the apetalous mutation in B. rapa.

This study used three winter wheat (Triticum aestivum L.) genotypes (H6756, H311 and SP8581) to compare the effects of sampling time, callus induction media, differentiation media and rooting media on in vitro culture of young spikes in wheat. In all these three genotypes, the frequencies of green plantlet differentiation were high when their young spikes were cultured between the stages of protective glume primordium formation and pistil and stamen primordium formation, but low at other stages. The optimum medium for callus induction was Murashige and Skoog (MS) medium+2 mg/l 2,4-dichlorophenoxyacetic acid (2,4-D). The optimum green plantlet differentiation medium was MS medium. Some abnormal plantlets regenerated from calli. When these plantlets were transferred to another differentiation medium [MS+1.0 mg/l 1-naphthaleneacetic acid (NAA)+0.2 mg/l 6-benzylaminopurine (6-BA)], shoot formation and elongation were induced. This allowed 90.91% of them to develop into normal green plantlets. The optimum rooting medium was 1/2MS+0.2 mg/l 3-Indolylacetonitrile (IAA)+80 g/l sucrose. An efficient regeneration system for young spike culture of wheat was set up based on such methods. Using this wheat-regeneration system, young spikes and immature embryos of 17 genotypes of wheat were in vitro cultured to study and compare the callus induction frequencies and green plantlet differentiation frequencies. The results of two successive years showed that in 15 out of the 17 genotypes (88.24%) the green plantlet differentiation frequencies were higher than those of immature embryos by 6.2–65.1%. These results showed that the regeneration system established in this trial for young spike culture of wheat was effective.