• Journal of Mushroom
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• v.19 no.1
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• pp.33-40
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• 2021

The genome sequence of various Flammulina species has recently been reported, thereby revealing a diverse genetic repertoire. In this study, we identified laccase genes and analyzed their structural characteristics in Flammulina elastica (F. elastica) genome. Through genome analysis and bioinformatics approaches, three laccase genes (Fe-lac1, -lac2, and -lac3) were identified, ranging from 1,548 to 1,602 bp in length. These genes contained a copper ion-binding region with ten histidine residues and one cysteine residue and a disulfide bond-forming region with four cysteine residues. Full-length cDNA sequencing analysis revealed that laccase genes contain 12 to 16 introns and signal peptides between 17 and 22 bp in the N-terminus. Structural characterization of the laccase genes identified in this study should help in better understanding the biomass decomposition of F. elastica.

• Journal of Life Science
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• v.32 no.8
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• pp.659-665
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• 2022

Chlorella vulgaris is an important freshwater alga that is widely used as a food source by humans and animals. Recently, Chlorella has received considerable attention with regard to its potential application in aquaculture and the production of biofuels, nutrients, and therapeutic proteins. Recently, our laboratory acquired a new strain of C. vulgaris, PKVL7422, characterized by fast growth, ease of culture, and cultivability under dark conditions. However, the genes involved in its nitrogen assimilation are unknown. In this work, we identified the nitrate reductase (NR) gene of C. vulgaris PKVL7422 using rapid amplification of cDNA ends and genome walking. The NR gene of C. vulgaris PKVL7422 is approximately 8 kb long and composed of 18 introns and 19 exons, which encode 877 amino acids. An alignment analysis of the NR gene showed that it possesses the five domains and several invariant residues found in plant NRs. These results provide new insight into the molecular organization of the NR gene in algae.

• Horticultural Science & Technology
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• v.30 no.2
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• pp.162-170
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• 2012

The objectives of this study were to isolate and the sequence of novel $F3'H$ gene related to an anthocyanin pathway, and to confirm the expression patterns of the gene involved in the flower color variations of chrysanthemum mutants. In this study, we isolated the full-length cDNAs and the genomic DNAs of an $F3'H$ gene from a wild type (WT) chrysanthemum (cv. Argus) and its three color mutants. The sequence analysis revealed a putative open reading frame of 1,527 bp that encodes a polypeptide of 509 amino acids. Sequence homology ranged from 97% to 99% between 'Argus' and its three color mutants. The sequence analysis from the genomic DNA revealed that the chrysanthemum $DgF3'H$ gene consisted of three exons and two introns spanning a 3,830 bp length. The sizes of the gene for three mutants ranged from a shorter size of 3,828 bp to a longer size of 3,838 bp when compared to the size of WT. The total size of the two introns was 2,157 bp for WT, but those for three color mutants ranged from 2,154 bp to 2,159 bp. A result of an RT-PCR analysis indicated that the color variations of the mutants AM1 and AM2 can be partly explained by the structural modification derived from the sequencial changes in the gene caused by gamma ray. A Southern blot analysis revealed that the $DgF3'H$ gene existing as multiple copies in the chrysanthemum genome. A systemic study will be further needed to provide a genetic mechanism responsible for the color mutation and to uncover any involvement of genetic elements for the expression of the $DgF3'H$ gene for the color variation in chrysanthemum.

• Asian-Australasian Journal of Animal Sciences
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• v.21 no.7
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• pp.928-935
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• 2008

The myostatin gene of seven important meat (Beltex (Australia), Beltex$\times$Huyang (F1), Meat and Multi-Prolific Chinese Merino Fine Wool, Meat Chinese Merino Fine Wool and Dorper (South Africa)) and non-meat (Huyang and Kazak) sheep breeds was analyzed to study the genetic basis of muscular hypertrophy (double muscling) phenotype in sheep. SNPs, four in regulatory regions and several in the introns in the myostatin gene, were identified, and the former four SNPs were used for further studies. Twelve haplotypes were predicted by PHASE program, of which four main haplotypes (1, 3, 7, 9) were present in 90% of the 364 sheep in the study. Haplotypes 1-4 were mainly present in meat breeds while haplotypes 7 and 9 dominated the non-meat breeds. The association between haplotypes and average daily gain (ADG) was analyzed among 116 sheep with production data, Haplo2 (CGAA) and Haplo8 (TGAA) were identified to have significant (p<0.05) effect on ADG by the model (JMP5.1 software) taking into account the effects of breed, family background, haplotype, birth weight and sex. ADG of these haplotype groups also correlated well (r = 0.82) with hypertrophic phenotype scores. In conclusion, the mutations -956 (T$\rightarrow$C), -41 (C$\rightarrow$A) and 6223 (G$\rightarrow$A) involved in Haplo2 and 8 may be associated with the double-muscling trait by influencing myostatin function and be suitable markers in selecting meat sheep.

• Asian-Australasian Journal of Animal Sciences
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• v.21 no.7
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• pp.917-922
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• 2008

A disintegrin-like and metalloprotease (reprolysin type) with thrombospondin type 1 motif (ADAMTS1) plays a critical role in follicular rupture and represents a major advance in the proteolytic events that control ovulation. In this study, a 9,026-bp DNA sequence containing the full coding region, all 8 introns and part of the 5'and 3' untranslated region of the porcine ADAMTS1 gene was obtained. Analysis of the ADAMTS1 gene using the porcine radiation hybrid panel indicated that pig ADAMTS1 is closely linkage with microsatellite marker S0215, located on SSC13q49. The open reading frame of its cDNA covered 2,844 bp and encoded 947 amino acids. The coding region of porcine ADAMTS1 as determined by sequence alignments shared 85% and 81% identity with human and mouse cDNAs, respectively. The deduced protein contained 947 amino acids showing 85% sequence similarity both to the human and mouse proteins, respectively. Comparative sequencing of three pig breeds revealed one single nucleotide polymorphism (SNP) within exon 7 of which a G-C substitution at position 6006 changes a codon for arginine into a codon for proline. The substitution was situated within a PvuII recognition site and developed as a PCR-RFLP marker for further use in population variation investigations and association analysis with litter size. Allele frequencies of this SNP were investigated in seven pig breeds/lines. An association analysis in a new Qingping female line suggested that different ADAMTS1 genotypes have significant differences in litter size (p<0.01).

• Korean Journal of Animal Reproduction
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• v.20 no.4
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• pp.371-378
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• 1997

Human lactoferrin (hLF) was expressed in the mammary gland of transgenic mice. Expresion of hLF was achieved by palcing its cDNA under the control of bovine $\beta$-casein gene. To improve the hLF expression level, two artificial introns were introduced into the expression vector. One intron is a hybrid-splice consisting of bovine $\beta$ casein intron 1 and rabbit $\beta$-casem intron II. The other intron is a DNA fragment spanning intron 8 of bovine $\beta$ casein gene. Trans sgenic mice were developed which expressed hLF in their milk. Twenty lines of transgenic mice were produced. hLF was present in the milk at concentrations of 1 ~ 200 ${\mu}\textrm{g}$ / ml. hLF RNA was only detected in the mammary gland of transgenic mice. The expressed RNA was cor r rectly spliced at the exon /intron junctions. To generate transgenic cows secreting active hLF in their milk, we transferred the DNA-injected bovine embryos to recipient heifers by surgical a and non-surgical methods out of 68 embryos transferred to 51 recipients by surgical or non-surgical method, 7 calves were normally born. Effect of embryo quality of DNA-injected blastocysts on pregnancy rate after transfer was investig a ated. Higher pregnancy rate of (38.9%) DNA-injected embryos was shown in excellent embryos. Pregnancy rates in the groups of good a and fair embryos were 15.4 and 14.3%, respectively. Effect of culture period of DNA-injected b bovine embryos on pregnancy rate after transfer was investigated. When Day-6 blastocysts of cuI ture were transferred, there was no pregnancy. Pregnancy rates of Day-7 and -8 blastocysts were 28.6 and 33.3%, respectively. There was no difference on pregnancy rate between Day-7 a and -8 bovine blastocysts after DNA injection. Thus, we established the techniques for transfer a and culture of DNA-injected bovine embryos. In a addition, factors affecting the pregnancy rate of DNA-injected embryos after transfer were investigated .

• Asian-Australasian Journal of Animal Sciences
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• v.23 no.1
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• pp.116-121
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• 2010

Long-chain polyunsaturated fatty acids (LC-PUFA) promote the development of brain and vision of the fetus, relieve inflammation, inhibit oral dysplasia of rumor cell, decrease the incidence of cardiovascular disease and regulate arrhythmia. ${\Delta}-6$ desaturase is the rate-limited enzyme in the desaturation process. This study reports the cloning, characterization and tissue expression of a ${\Delta}-6$ desaturase gene in the chicken. PCR primers were designed based on the predicted sequence of chicken ${\Delta}-6$ desaturase (accession number: XM421053) and used to isolate a cDNA fragment of 1,323 bp from chicken liver. Based on the 1,323 bp fragment an EST (BI390105) was obtained by BLAST. The EST and 5'nd of the 1,323 bp fragment were partially overlapped. Gene specific primers derived from the EST were used for amplification of the 5'nd. Another gene-specific primer derived from the 1,323 bp fragment was used for amplification of the 3'nd by 3'ACE. Then the three overlapping cDNA sequences obtained were assembled with DNAMAN software and a full-length ${\Delta}-6$ desaturase of 2,153 bp was obtained. The full-length cDNA contained an ORF of 1,335 bp with a 5'ntranslated region of 147 nucleotides followed by an ATG initiation codon. Stop codon TGA was at position 1,481-1,483 bp. The deduced amino acids shared an homology above 77% with bovine, mice, orangutan, rat and human. The protein sequence had three histidine-rich regions HDFGH (HisI region), HFQHH (HisII region) and HH (HisIII region), a cytochrome $b_{5}$-like domain containing a heme-binding motif and two transmembrane domains. Sequence analysis of the chicken genomic DNA revealed that the coding sequence of chicken ${\Delta}-6$ desaturase included 12 exons and 11 introns. Semi-quantitative RT-PCR showed that the ${\Delta}-6$ desaturase expression levels were in turn liver, spleen, pancreas, lung, breast muscle, heart, and abdominal fat. The expression of ${\Delta}-6$ desaturase in liver was significantly higher than that in breast muscle (p<0.01). The expression of ${\Delta}-6$ desaturase in lung was significantly higher than that in abdominal fat (p<0.01). This is the first clone of chicken ${\Delta}-6$ desaturase.

• BMB Reports
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• v.40 no.2
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• pp.247-260
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• 2007

Cinnamate 4-hydroxylase (C4H) is a key enzyme of phenylpropanoid pathway, which synthesizes numerous secondary metabolites to participate in development and adaption. Two C4H isoforms, the 2192-bp BnC4H-1 and 2108-bp BnC4H-2, were cloned from oilseed rape (Brassica napus). They both have two introns and a 1518-bp open reading frame encoding a 505-amino-acid polypeptide. BnC4H-1 is 57.73 kDa with an isoelectric point of 9.11, while 57.75 kDa and 9.13 for BnC4H-2. They share only 80.6% identities on nucleotide level but 96.6% identities and 98.4% positives on protein level. Showing highest homologies to Arabidopsis thaliana C4H, they possess a conserved p450 domain and all P450-featured motifs, and are identical to typical C4Hs at substrate-recognition sites and active site residues. They are most probably associated with endoplasmic reticulum by one or both of the N- and C-terminal transmembrane helices. Phosphorylation may be a necessary post-translational modification. Their secondary structures are dominated by alpha helices and random coils. Most helices locate in the central region, while extended strands mainly distribute before and after this region. Southern blot indicated about 9 or more C4H paralogs in B. napus. In hypocotyl, cotyledon, stem, flower, bud, young- and middle-stage seed, they are co-dominantly expressed. In root and old seed, BnC4H-2 is dominant over BnC4H-1, with a reverse trend in leaf and pericarp. Paralogous C4H numbers in Brassicaceae genomes and possible roles of conserved motifs in 5' UTR and the 2nd intron are discussed.

• Korean Journal of Microbiology
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• v.49 no.1
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• pp.8-16
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• 2013

A GAL4 type transcription factor gene (formally annotated as AN3912) locating downstream of sndA (AN3911) was characterized. The putative transcription factor carries both Zn(II)2Cys6 binuclear cluster DNA-binding domain and transcription activator domain. The gene named gtfA (gal4 type transcription factor) had an open reading frame which consisted of 762 amino acids and was disrupted by three introns. The deletion mutant produced reduced amount of conidia but increased amount of fruiting bodies, suggesting that the GtfA make function in decision of asexual preferential to sexual development. The forced over expression of gtfA caused the retardation of fruiting body formation on high glucose concentration. The transcript level of gtfA was kept constant through the life cycle except late vegetative stage and early sexual development stage during which slight increase was found. The expression of gtfA was not significantly affected by sexual or asexual development regulators, such as VeA, NsdD or FluG, FadA, and SfaD. The GtfA repressed the nsdC transcription, which suggested that GftA control sexual development negatively via negative regulation of nsdC expression.

• BMB Reports
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• v.38 no.6
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• pp.739-747
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• 2005

Full-length cDNA sequences of four novel SPATA4 genes in chimpanzee, cow, chicken and ascidian were identified by bioinformatic analysis using mouse or human SPATA4 cDNA fragment as electronic probe. All these genes have 6 exons and have similar protein molecular weight and do not localize in sex chromosome. The mouse SPATA4 sequence is identified as significantly changed in cryptorchidism, which shares no significant homology with any known protein in swissprot databases except for the homologous genes in various vertebrates. Our searching results showed that all SPATA4 proteins have a putative conserved domain DUF1042. The percentages of putative SPATA4 protein sequence identity ranging from 30% to 99%. The high similarity was also found in 1 kb promoter regions of human, mouse and rat SPATA4 gene. The similarities of the sequences upstream of SPATA4 promoter also have a high proportion. The results of searching SymAtlas (http://symatlas.gnf.org/SymAtlas/) showed that human SPATA4 has a high expression in testis, especially in testis interstitial, leydig cell, seminiferous tubule and germ cell. Mouse SPATA4 was observed exclusively in adult mouse testis and almost no signal was detected in other tissues. The pI values of the protein are negative, ranging from 9.44 to 10.15. The subcellular location of the protein is usually in the nucleus. And the signal peptide possibilities for SPATA4 are always zero. Using the SNPs data in NCBI, we found 33 SNPs in human SPATA4 gene genomic DNA region, with the distribution of 29 SNPs in the introns. CpG island searching gives the data about CpG island, which shows that the regions of the CpG island have a high similarity with each other, though the length of the CpG island is different from each other.This research is a fundamental work in the fields of the bioinformational analysis, and also put forward a new way for the bioinformatic analysis of other genes.