Mosquitoes are primary medical insect pests due to their diseases transmission as vectors. In Korea, the insecticide-resistant populations of disease vector mosquito species, such as Anopheles sinensis, Culex pipiens and Culex tritaeniorhynchus, have constantly increased. Thus, management of insecticide resistance to major insecticides including pyrethroids and organophosphates is required for more efficient control of resistant populations. In this study, the quantitative sequencing (QS) protocols were established to detect the frequencies of three mutations (the L1014F on voltage sensitive sodium channel and the G119S and F331W on acetylcholinesterase 1) that are associated with either pyrethroids or organophosphates. Based on the QS protocol using newly designed non-polymorphic primers, resistance allele frequencies (RAFs) were estimated in field populations of An. sinensis, Cx. pipiens and Cx tritaeniorhynchus collected from an identical site in Korea. The dynamics of each resistance allele frequency over time in the same populations were also evaluated.

Amphiregulin (AREG), a glycoprotein that is a member of the epidermal growth factor (EGF) family, is expressed by the porcine conceptus and endometrium. AREG genotypes were determined based on an SNP in the intron 3 of the gene. Contradictory effects of AREG genotypes on reproductive traits in different pig breeds were reported previously. G allele had undesirable effect on reproductive trait in Meishan breed, while it had favorable effects in Polish Landrace and Large White. We determined AREG genotypes of 179 pigs including the Duroc, Landrace, Yorkshire, Korean native pig (KNP), and Meishan breeds. Two new SNPs were identified near the previously reported SNP in the intron 3 of AREG. Frequencies of AREG alleles among the Duroc, Landrace, Yorkshire, and KNP sows were significantly different (p<0.001), indicating association between AREG genotypes and pig breeds. The first parity litter size was significantly affected by the breeds (p=0.014), but not by AREG genotypes (p=0.148). However, there were breed and AREG genotype associated trends in the first parity litter size. The first parity litter size appeared to be higher in Duroc and KNP sows with G allele, while it appeared to be lower in Landrace sows with G allele. Significant variability of AREG alleles among pig breeds, for the first time in Duroc and KNP sows, was identified. AREG genotypes may influence reproductive traits differentially for each breed and thus, AREG genotypes may need to be considered when sows are bred to increase litter size.

A quantitative sequencing (QS) protocol was established for predicting the frequencies of the A298S and G324A mutations in the diamondback moth (Plutella xylostella) type-1 acetylcholinesterase locus, putatively involved with organophosphate and carbamate insecticideresistance. The nucleotide signal ratio at each mutation site was generated from sequencing chromatograms and plotted against the corresponding resistance allele frequency. Frequency prediction equations were generatedfrom the plots by linear regression, and the signal ratios were shown to highly correlated with resistance allele frequencies (r2>0.987). QS analysis of 15 representative regional field populations of DBM in Korea revealed that the allele frequencies of both A298S and G324A were over 70% in most field populations. As judged by inhibition assay, all populations showed resistance to paraoxon, DDVP, carbaryl, and carbofuran. In addition, different DBM strains exhibited differential sensitivities to both OPs and CBs depending on the structure of inhibitor, implying that the resistance of DBM against OPs and CBs is saturated and widespread in Korea.

A quantitative sequencing (QS) protocol that detects the frequencies of sodium channel mutations (M815I, T917I and L920F) responsible for knockdown resistance in permethrin-resistant head lice was tested as a population genotyping method. Genomic DNA fragments of the sodium channel α-subunit gene that encompass the three mutation sites were PCR-amplified from individual head lice with either resistant or susceptible genotypes, and combined together in various ratios to generate standard DNA template mixtures for QS. Following sequencing, the signal ratios between resistant and susceptible nucleotides were calculated and plotted against the corresponding resistance allele frequencies. Quadratic regression coefficients of the plots were close to 1, demonstrating that QS is highly reliable for the prediction of resistance allele frequencies. Prediction of resistance allele frequencies by QS in several globally collected lice samples including 12 Korean lice populations suggested that permethrin resistance varied substantially amongst different geographical regions. Three local populations of Korean lice were determined to have 9.8-36.7% resistance allele frequencies, indicating that an urgent resistance management is needed. QS should serve as a preliminary resistance monitoring tool for proper management strategies by allowing early resistance detection.

Insulin-like growth factor 2 (Igf-2) and H19 genes are closely linked imprinted genes which have a pivotal role in embryogenesis and fetal development. Igf-2 and H19 are coexpressed in tissues of mesodermal, endodermal and neuroectodermal origin. Rat Igf-2 has a complex structure with three promoters and a complicate imprinting mechanism having an exception of biallelic expression in the choroid plexus, leptomeninges, and fetal tissues of neuroectodermal origin. To detect the expression of maternal and paternal alleles of Igf-2 and H19 during orofacial development, fetal and neonatal hybrid rats, obtained from Wistar and Fisher interstrain rat crosses were used. We also detected the promoter-specificity of Igf-2 transcripts by primers selected from P1, P2, and P3 of Igf-2 gene. RT-PCR analysis of Igf-2 and H19 showed the monoallelic expression of Igf-2 from the paternal allele and H19 from the maternal allele in E15.5 to E19.5 orofaciall structures including the maxilla, tongue, and salivary gland. P3 promoters were active in all tested samples, whereas transcripts derived from P2 promoter arised with approximately half of the tested cases and showed variable alternation. P1 promoter was not transcribed in all tested samples. These results suggest that Igf-2 and H19 may be involved in orofacial development and exhibit parent-of-origin monoallelic expression. On the other hand, in orofaciall development, P2 and P3 promoters except for P1 promoter are transcribed with variable alternative transcripts

Aroma in rice (Oryza sativa L.) results from the loss of function of the betaine aldehyde dehydrogenase (Badh2) gene on chromosome 8. An 8-bp deletion in exon 7 of Badh2 was reported to be the main allele functionally associated with fragrance. The discovery of new functional alleles will provide additional genetic resources to improve fragrant rice. In this study, we sequenced the Badh2 gene in 30 rice accessions and filtered the Badh2 polymorphisms from whole genome re-sequence data of 295 rice accessions. Seven alleles were detected from the sequence data. Six of the seven were known alleles and one was a novel allele (badh2-E12). The novel allele was a 3-bp deletion in exon 12. Five functional markers, targeting six of the seven alleles, were identified. Fourteen accessions were selected to test the utility of these markers. The five molecular markers reliably distinguishing this fragrant rice from other fragrant or non-fragrant rice accessions. Analysis of two F2 rice population validated the genetic markers FME12-3 and FME14I as functional markers. These two markers co-segregated with the fragrance phenotype. These markers will be used in a Badh2 diversity study and to breed improved fragrant rice accessions via marker-assisted selection.

사과는 배우체형 자가불화합성을 나타내는데 이는 S-locus의 복대립유전자에 의해 조절된다. 본 연구는 S-allele specific PCR분석을 통해 신품종을 포함한 24종의 사과 주요 재배품종과 7종의 꽃사과 품종의 자가불화합성 유전자형(S-genotype)을 결정하고자 수행하였다. 31종의 재배품종과 꽃사과 품종 을 23종의 S-allele specific primer을 이용하여 분석한 결과 12개의 S-allele (S1, S2, S3, S5, S7, S9, S10, S16, S21, S23, S26, S29)이 동정되었다. 그 중에서 24종의 재배품종에는 S1(41.7%), S3(58.3%), S7(29.2%), S9(54.2%)의 S-allele이 흔히 존재하는 것으로 확인되었다. 국내육성 신품종인 ‘아리수’와 ‘황옥’의 S-genotype은 각각 S3S7과 S3S9으로 동정되었다. 본 실험에서 얻은 S-genotype 정보는 안정적인 사과 과실생산에 적합한 수분수 선발과 육종프로그램에서 교배조합 작성에 유용 하게 활용될 것이다.

Genetic resources play a great role in crop breeding because of containing a broad array of useful genes. Currently, the harder are rice breeders trying to develop new rice cultivars with the improved traits, they are more often handicapped by the limited availability of germplasm resources. Thus, a desirable core or heuristic (HS) set of germplasm with maximum genetic diversity can be usefully exploited to breakthrough the present and future challenges of the rice breeding. As such we previously developed the rice HS sets of 166 diverse accessions out of a total 24,368 rice germplasms. Here, we report a large-scale analysis of the patterns of genome-wide genetic variations accumulated in the HS as well as Korean rice over the time. We characterized a total of about 11.8 millions of single nucleotide polymorphisms (SNPs) across the rice genome from resequencing a total of 295 rice genomes including 137 HS and 158 KB rice sets, with an average of approximately 10x depth and > 90% coverage. Using about 460,000 high-quality SNPs (HQSNPs), we specified the population structure, confirming our HS set covers all the rice sub-populations. We further traced the relative nucleotide variabilities of HQSNPs and found the level of the diversity was dynamically changing across the KB genome, which reveals the selection history of KB lines in the past and present. In addition, the results of our genome wide association study (GWAS) suggests that our HS can be also a good reservoir of valuable alleles, pinpointing those alleles underlying the important rice agronomical traits. Overall, the resequencing of our HS set re-illuminates the past, present of the germplasm utilization, which will support the Korean rice breeding in the future.

벼줄무늬잎마름병은 애멸구에 의해 매개되며 우리나라, 일본, 중국 등 동아시아 지역에서 벼에 가장 많은 피해를 끼치는 바이러스병이다. 벼 줄무늬잎마름병 저항성 유전자는 6번 염색체에 존재하는 Stv-a 유전자와 11번 염색체에 존재하는 Stv-b 유전자 및 Stv-b 유전자의 복대립 유전자형인 Stv-bi 등이 알려져 있다. 단일유전자에 의한 저항성의 경우 바이러스 집단의 변화, 저항성 유전자의 기능소실 등에 의해 저항성의 붕괴가능성이 제기되면서 기존의 유전자 이외의 저항성 유전자를 추가 탐색하는 노력이 진행되고 있다. 본 연구는 국내 통일형 품종 및 국외 도입 유전자원을 중심으로 총 155 품종에 대한 저항성 생물검정과 기존에 개발된 Stv-bi 유전자 영역을 분자마커를 이용한 유전자형 검정을 통해 신규 유전자를 포함하는 저항성 자원을 선발함을 목적으로 수행하였다. 분자마커를 이용한 유전자원의 군집분석 결과 크게 2개의 군으로 분류되었으며 이는 또한 InDel 7에 의한 유전자형 구분결과와 일치하였다. 그 중 Daw dam 등 6품종은 InDel 7 marker에 의한 유전형 분석결과에서 감수성 allele을 나타내었으나 RSV 생물검정에서는 유전형과 반대인 저항성을 나타내어 InDel 7 marker로서 구분이 불가능 한 것으로 나타났으며 ST10에 대한 유전형에서도 감수성 allele을 나타내었다. 따라서 이들 6품종은 11번 염색체에 존재하는 Stv-bi 유전자가 아닌 다른 유전자에 의해 저항성을 나타낼 가능성이 높은 것으로 추정되었다. 이들 품종은 향후 유전분석과 유전자지도작성을 통하여 신규 유전자 여부 확인을 위한 좀 더 상세한 검토가 필요한 것으로 생각되며 향후 저항성 증진 또는 저항성 붕괴에 따른 대책으로 gene pyramiding등에도 이용 가능할 것으로 판단된다.

The use of functional markers, it is expected to make direct identification about genetic diversity at DNA level and overcome the problem of recombination /linkage. These markers can be used to identify interesting alleles in a breeding program and indirectly select for the trait, saving money, time and labor. Bacterial blight of rice caused by Xanthomonas oryzaepv. Oryzae is a destructive disease in rice production worldwide. No bactericide is effective to control the bacterial blight disease yet. Xa3, which is a gene conferring resistance to BB of the rice plant has been previously characterized by map-based cloning. We have cloned and sequenced the Xa3/xa3 gene in Korean cultivar, Hwayoung, Ilmi and Goun with gene specific primers. Our work detected polymorphisms and PCR-based allele specific SNP markers were developed. Susceptible or resistant individuals from an F2 population developed from across between Milyang244 and Ilmi, Korean germplasms and near isogenic lines carrying BB resistance genes were screened with allele specific markers. We found that the genotype completely matched their phenotype to BB using ASP-primers. These markers could be effective to marker-assisted selection for the Xa3 gene in rice breeding programs.

Brown planthopper (BPH) is a destructive insect pest of rice in Korea. Identification and the incorporation of newBPH resistance genes into modern rice cultivars are important breeding strategies to control the damage caused by BPH. To expand genetic resource against BPH in Korea, we screened more than 2,500 mutant lines, which were derived from EMS treatment on Namil, a high yielding Korean japonica cultivar. One mutant line, Namil(EMS)M2-1463-1-1-1-1, designated as ‘Namil(EMS)-bl10,bph1’ performed high level of resistance against BPGH and rice blast, while the wild type, Namil, performed highly susceptible to rice blast as well as BPH. A mapping population was constructed by using F2 progeny lines derived from cross between Namil(EMS)-bl10,bph1 and Milyang23, a BPH susceptible Tongil type cultivar. DNAs prepared from F2 individuals were used for SSR marker based linkage map skeleton, and F2:3 seeds were subjected to BPH infestation to infer resistance level of corresponding F2 plant. Association analysis between marker genotype and evaluated survival ratio of each progeny line were used to localize the putative chromosomal location(s) involved to BPH resistance. The location was initially located on the subterminal region of the long arm of chromosome 12 flanked by the SSR markers RM1337 and RM277, where at least three BPH resistance genes, Bph1, Bph18, and Bph21, were localized previously.

본 연구의 목적은 알츠하이머 질병과 관련된 유전자 (아포지단백 e4 유전형질)를 보유함으로써 유발되는 뇌기능의 결함 (뇌 활성화 감소, 대뇌피질 두께 감소)을 유산소능력이 상쇄시켜줄 수 있는지를 심리생리학적 측정법을 통해 조사하는 것이다. 연구대상자는 평균연령 20세의 건강한 성인 남성들로 29명의 아포지단백 e4 유전형질 보유자와 30명의 무보유자로 구성되었으며, 이들은 YMCA 싸이클 프로토콜을 통해 예측된 최대산소섭취량에 의해 고 유산소능력 집단과 저 유산소능력 집단으로 구분되었다. 인지기능은 오드볼 과제 수행 중 Fz, Cz, Pz 영역에서 측정된 신경전기적 반응인 사건유발전위의 P300 진폭과 잠재기를 통해 조사되었다. 연구 결과 e4 유전형질을 보유한 집단에서 고 유산소능력 집단의 Fz와 Cz에서 측정된 P300 진폭이 저 유산소능력 집단의 진폭보다 유의하게 컸다. 그러나 유산소능력에 따른 P300 진폭의 차이는 유전형질을 보유하지 않은 집단에서는 나타나지 않았다. P300 잠재기 분석에서는 유전형질 보유여부에 상관없이 고 유산소능력 집단의 Pz에서 측정된 P300 잠재기가 저 유산소능력 집단보다 짧았다. 이러한 결과들은 유산소능력이 높을수록 정보처리 속도가 빠르고, 과제 수행시 특정 뇌영역에 주의자원의 양이 많다는 것을 의미한다. 유산소능력이 유산소 운동훈련과 정적으로 상관되어 있고 유산소능력의 효과가 유전형질을 보유한 집단에게 더 크다는 사실은 알츠하이머 질병의 유전적인 위험에 노출된 사람들에게 유산소운동에 꾸준한 참여가 더욱 중요할 수 있음을 의미하는 것이다.