The characteristics and spore production of Gonji7ho, Bunhong, and Sunjung fruiting bodies were assessed at different growth stages. The shape of the Pleurotus species fruiting body starts out short and small, then takes on a typical mushroom shape as it grows. Gonji7ho has a long stalk, Bunhong has a short stalk and a wide cap, and Sunjung's cap and stalk dimensions are intermediate. Each variety displayed deep color at the beginning of growth but became steadily lighter with continued growth. The shape of the linkage between the mushroom stalk and cap changed from an initial central position to a lateral position after the growing stage. Gonji7ho cap diameter increased 7-fold from 15.5 mm (5 days of growth) to 37.9 mm (9 days of growth). Growth rates for each growth day measured using the growth percentage of the previous day were 285.5% (5 → 6th day), 182.2% (6 → 7th day), 129.4% (7 → 8th day), and 103.8% (8 → 9th day). This trend was also observed in Bunhong and Sunjung, but Bunhong’s growth rate was more rapid (4.9 fold on day 6, 2.7 fold on day 7) and continued to increase through day 9. Harvest yield, which is of greatest interest to farmers, displayed a similar trend spanning the growth period, as did cap diameter. Gonji7ho harvest yield increased rapidly until day 7 of growth (more than 177%), then growth slowed down beginning around day 8, and further decreased on day 9 (98%). Similar trends were observed in Bunhong and Sunjung. Bunhong showed characteristic rapid growth in harvest yield (4.9 fold compared to the previous day on day 6 and 2.7 fold on day 7), and the increase continued through day 9. A decrease in mushroom harvest yield commonly seen in the late growth stage is thought to be due to the death of some mushrooms and decomposition of cap tissue. Basidiospore content increased with number of growth days but decreased after day 8. Gonji7ho yielded the highest production on day 7 of growth, coinciding with harvest time, with 209,000,000 spores. This trend was also observed in Bunhong and Sunjung. These results will provide researchers with basal data and guide farmers in selecting the optimal harvest day.

Light plays an important role in fruit-body development and morphology during Pleurotus spp. cultivation. To understand the effects of light color on fruit-body properties, we evaluated the fruit-body characteristics of Pleurotus spp. varieties cultivated under blue, red, and purple LED light sources. The main results are as follows: The overall fruit-body shape showed differences depending on the color of the LED light. The fruit-bodies of mushroom cultivated under blue and purple light were generally similar to the mushroom shapes typically produced, while those of mushroom cultivated under green light were abnormally shaped, probably due to the absence of effective light source. The average cap lightness of mushrooms cultivated under blue, green, and purple LED lights was 57.0, 57.4, and 59.4, respectively. The average cap lightness of all varieties except Wonhyeong1ho and Hwang-geumsantari cultivated under the three LED light sources were statistically significantly different (P<0.05). The cap redness varied significantly depending on the LED lighting and variety. Only Gonji7hoM, the cap color mutant of Gonji7ho, showed negative cap redness values under all three LED light sources. Among the eight varieties excluding Gonji7ho, the highest cap redness was observed when cultivated under the blue LED. The average harvest weight of the varieties cultivated under purple, blue, and green LED light were 68.0, 58.3, and 50.1 g, respectively. The yield of Gonji7ho, the mushroom variety with the highest yield, cultivated under blue, green, and purple LED light were 92.8, 77.1, and 98.6 g, respectively. The earliness when grown under the purple, blue, and green LED lights were 5.3, 5.8, and 5.8 days, respectively. Among the varieties, six, three, and two cultivars showed the shortest earliness under the purple, green, and blue LED, respectively. The fruit-body lengths were 66.4, 51.8, and 46.8 mm when cultivated under green, purple, and blue lights, respectively. These results are expected to serve as a foundation for producing mushrooms with traits demanded in the market.

To elucidate how cultivation temperature affected various traits including pileus color, yield and morphology of Pleurotusspp. Main results were as follows. Pileus lightness of all cultivars of Pleurotustested became higher as cultivation temperature increased, while those of Santari, Hwang-geumsantari and Sunjung at 21oC were lower than at 18oC. Redness and yellowness of pileus decreased as cultivation temperature increased; those of chromatic pileus cultivars showed noticeable difference. Yellowness of cultivar with chromatic pileus was higher than that of cultivar with achromatic pileus. Yield was increased as cultivation temperature increased, Wonhyeung 1ho; low temperature favored cultivar showed high yield when it was cultivated at low temperature andno fruiting body at 21oC. Valid number of stipes were generally higher at 18oC, and its correlation coefficient with yield was low. Length and stipe thickness changed consistently (larger and thicker) upon cultivation temperature; the coefficient of determination(R2) 0.514 for lengthof Heuktari and 0.963for stipe thickness of Santari were high. Correlation coefficient of one trait was highly related with multiple traits. In the future, we will conduct research on the changes of expressed genes involved in the pigments for pileus color by RNA expression analysis.

국내의 주요 산느타리 품종인 호산47(일핵, 산타리 배우자), GB19(일핵, 산타리 배우자), 호산, 여름느타리1호, 삼복, 강산, 약산, 자산, 향산, 여름느타리2호의 유전체를 Hiseq을 이용하여 해독하였고 이 서열 정보에서 SSR을 분리하여 특성구명을 하였다. 일핵균사인 호산 47, GB19 의 유전체의 크기는 각각 37.3와 37.2 Mbp이고, 이핵균사인 나머지 산느타리 품종의 유전체 크기는 47.1~61.1 Mbp인 것으로 밝혀졌다. 품종별 총 SSR의 수는 HS47이 711개로 가장 적고, 강산이(GS)이 1.5배 많은 1,106개로 최다를 기록하였다. SSR의 repeat motif 중에서 hexanucleotide 와 octanucleotide가 가장 많은 빈도로 관찰되었고, 가장 많이 관찰되는 반복서열은 CGA/TCG, A/T, CTC/GAG이었다. SSR의 길이는 모든 품종에서 변이가 많아 유용성이 높은 20~30 nt가 가장 높은 비중인 70%를 차지하였다.

느타리 품종구분을 위한 마커의 개발을 위하여 곤지7호 의 어버이 일핵 균사중의 하나인 MT07156-97의 전체 유전자 염기서열을 바탕으로 제작한 251개의 SSR 프라이머를 제작하였다. 우선적으로 수한1호, 곤지7호, 흑타리 품종에 다형성 여부를 관찰하여 20개의 SSR을 선발하고, 이를 10개 품종에 적용하였다. 단일의 프라이머로는 일부 품종이 구분되지 않았으므로, 선발된 프라이머 간의 다양한 조합(multiplex 방식)을 적용한 결과 모든 품종을 판별 할 수 있는 분자마커 다형성을 보인 프라이머 "166+115" 조합을 선발하였다. 별도로 프라이머 115와 166가 만들어 낸 산술적인 유전자좌(loci) 31개보다 12개 많은 40개의 유전자좌가 증폭되어 다양한 품종에 특이적인 분자마커를 제공할 수 있었다. 개발된 분자마커는 종균의 품질관리, 품종의 판별, 신품종 보호에 활용될 수 있을 것이다.

큰느타리버섯 주요 수출시장인 유럽과 북미시장의 소비자에게 선호도가 높은 갓이 큰 형태의 큰느타리버섯 품종을 육종하기 위하여 육종모본 KNR2555를 자식교배하여 소규모 시험재배하여 갓형(Convex)와 갓직경(60.7 mm), 품질(4.9)을 기준으로 2×12계통을 선발하였다. 선발된 계통을 갓애린이라고 명명하고 대량재배로 큰느타리2호와 생육특성을 비교하였다. 병당수량은 갓애린이가 71.7 g으로 대조품종 71.4 g과 통계적 유의성 없었다. 품질의 경우 갓애린이는 6.8, 큰느타리2호는 6.5로 나타났다. 생육소요일, 길이, 갓직경은 독립 t test로 분석한 결과 통계적으로 유의성을 보였다(각각 P < 0.001, P < 0.05, P < 0.001, P < 0.05). 고유성에 있어서는 URP1와 URP10에서 대조 품종과 신품종이 다형성을 보였고, 대치배양에서도 뚜렷한 대선이 관찰되었다.

생육이 빠르고 중형인 큰느타리버섯 품종을 육종하기 위하여 육종모본 24×46과 KNR2539를 단교배하여 소규모 시험재배하여 생육소요일수(15.4일)와 수량(81.5 g/850 cc), 품질(7.5)을 기준으로 17×15계통을 선발하였다. 선발된 계통을 ‘애린이6’라고 명명하고 대량재배로 큰느타리2호와 생육특성을 비교하였다. 병당수량은 ‘애린이6’가 76.0 g으로 대조품종 61.4 g의 113% 수준이었다. 품질의 경우 ‘애린이6’는 6.8, ‘큰느타리2호’는 5.7로 나타났다. 생육소요일, 수량, 품질은, 독립 t test로 분석한 결과 통계적으로 유의성을 보였다. 갓의 명도(L)에 있어서 ‘애린이6’는 61.7로 대조품종의 58.3보다 높아서 밝은 색을 띄었다. 고유성에 있어서는 URP2와 URP11에서 대조품종과 신품종이 다형성을 보였고, 대치배양에서도 뚜렷한 대선이 관찰되었다.

Simple sequence repeats (SSR), also referred to “microsatellites” consist of tandemly repeated short DNA sequence motifs and have been applied in various marker-based studies. SSRs were isolated and characterized from ‘Heuktari’ and ‘Miso’, which are major oyster mushroom cultivars in Korea, by genome sequencing and bioinformatic analysis. The genome sizes of ‘Heuktari’ and ‘Miso’ were estimated to be 40.8 and 40.3 Mb, respectively, which are larger than those of other P. ostreatus species (PC9 and PC10) and smaller than those of P. eryngii (KNR2312P5). In total, 949 and 968 SSRs were found in the ‘Heuktari’ and ‘Miso’ genomes, respectively. Comparative analysis of five mushrooms including P. ostreatus var. florida (PC9 and PC15) and P. eryngii revealed that the number of SSRs in ‘Heuktari’ and ‘Miso’ were the highest among them. All mushrooms studied showed similar SSR distribution patterns. Tri-, hexa-, and octanucleotide motifs accounted for the top three fractions of all SSRs.

To breed a new Korean Pleurotus eryngii cultivar with high quality and yield, single crosses between 24×46 and KNR2539 were performed, and a new cultivar, 6×13, was selected based on the days to harvest (14.6 days), quality (7.2), and yield (85.9 g/850 cc bottle). The strain was named Aeryni 5 and cultivated on a large scale at the mushroom farms to compare with Keuneutari 2ho. The yield of Aeryni 5 (82.2 g) was 122.7% of Keuneutari 2ho, and the quality of the new cultivar was 7.7 while reference cultivar was 6.3. The yield and quality of the two cultivars were statistically different. The lightness of the pileus of Aeryni 5 (61.7) was greater than that of Keuneutari 2ho by 3.4 points; thus, the pileus of Aeryni 5 looked brighter. PCR with URP2 was used to discriminate between Aeryni 5 and Kenneutari 2ho.

Differentiation of Pleurotus eryngii is laborious and time-consuming tasks especially in mycelial status. For development of a method for differentiation of P. eryngii cultivars, simple sequence repeats (SSR) from whole genomic DNA sequence analysis was used for genotyping and two multiplex-SSR primer sets were developed. These SSR primer sets were employed to distinguish 12 cultivars and strains. Five polymorphic markers were selected based on the genotypes. PCR with the each primer produced one to four distinct bands ranging in size from 200 to 300 bp. Polymorphism information content (PIC) values of the five markers were in range of 0.6627 to 0.6848 with an average of 0.6775. Unweighted pair-group method with arithmetic mean clustering analysis based on genetic distances using five SSR markers classified 12 cultivars into 2 clusters. Cluster I and II comprised of 4 and 8 cultivars, respectively. Two multiplex sets, Multi-1 (SSR312 and SSR366) and Multi-2 (SSR178 and SSR277) completely discriminated 12 cultivar and strains with 21 allele with a PIC value of 0.9090. These results might be useful to provide an efficient method for the identification of P. eryngii cultivars with separate PCR reactions. (This work was supported by a grant from the Golded Seed Project (213003-04-3-SBY20), MIFAFF, Republic of Korea.]

Pleurotus eryngii has recently become a major cultivated mushroom; it uses tetrapolar heterothallism as a part of its reproductive process. Sexual development progresses only when the A and B mating types are compatible. Such mating incompatibility occasionally limits the efficiency of breeding programs in which crossing within loci-shared strains or backcrossing strategies are employed. Therefore, understanding the mating system in edible mushroom fungi will help provide a short cut in the development of new strains. We isolated and identified pheromone and receptor genes in the B3 locus of P. eryngii and performed a functional analysis of the genes in the mating process by transformation. A genomic DNA library was constructed to map the entire mating-type locus. The B3 locus was found to contain four pheromone precursor genes and four receptor genes. Remarkably, receptor PESTE3.3.1 has just 34 amino acid residues in its C-terminal cytoplasmic region; therefore, it seems likely to be a receptor-like gene. Real-time quantitative RT-PCR (real-time qRT-PCR) revealed that most pheromone and receptor genes showed significantly higher expression in monokaryotic cells than dikaryotic cells. The pheromone genes PEphb3.1 and PEphb3.3 and the receptor gene PESTE3.3.1 were transformed into P5 (A3B4). The transformants were mated with a tester strain (A4B4), and the progeny showed clamp connections and a normal fruiting body.

큰느타리버섯 품종 육성을 위해 품질이 우수한 특성을 지니는 큰느타리버섯 KNR2598과 경도가 우수한 특성을 지니는 큰느타리버섯 KNR2610 모본으로부터 단핵균주를 분리 한 뒤 단포자간 교잡을 통해 고품질의 저온 저장성이 우수한 신품종 “단비5호”를 육성하였다. 신품종의 균사 생 육 적정온도는 25 o C이며 자실체 발생 적정 온도는 15~ 16 o C 이었다. 솎음 재배에서 발이소요일수를 포함한 수확 소요일수는 대조품종인 큰느타리버섯 2호에 비해 0.5~1.7 일 정도 빠른 특성을 보였다. 갓 색깔은 중간수준의 갈색이 며 대 색깔은 흰색을 나타내었다. 갓모양은 우산형으로 850cc 플라스틱 병재배에서 한 개체의 평균무게는 90.4 ±16.9 g이었다. “단비5호”와 대조품종간의 RAPD 분석결과 서로 다른 DNA 밴드양상을 보여 주었으며 대치배양을 통 해 두품종간에 대치선을 확인 하였다. “단비5호”의 경우 4 o C에서의 저장성이 대조품종 대비 24.6~30.6일 이상 길게 유지되는 특성이 있어 수출적합형 품종으로 가치가 있을 것으로 기대된다.

본 연구는 큰느타리버섯의 베타글루칸 고함유 형질에 관련된 SCAR marker를 개발하기 위해 수행되었다. operon사의 OPA(20개), OPB(20개), OPL(20개), OPP(20개), OPR(20개), OPS(20개) 등 총 120개 primer를 random primer(10 mer)로 사용하여 대립 계통 9종과 베타글루칸 고함유 계통 9 종을 대상으로 RAPD를 이용한 bulked segregant analysis를 실시하여 OP-R03 primer로부터 대립 계통에는 나타나지 않고 베타글루칸 고함유 계통에만 나타나는 특이적인 RAPD 밴드를 얻었다. OP-R03 primer를 이용한 RAPD 결과, 약 91 bp 부근에서 베타글루칸 고함유 계통에 특이적인 DNA 밴드가 관찰되었으며 이 DNA 밴드의 염기서열 말단을 근거로 SCAR 마커로 사용할 specific primer인 OP-R03-1-F와 OP-R03-1-R를 디자인하였다. SCAR 마커 OP-R03-1-F/-1-R primer를 이용하여 PCR을 수행한 결과에서도 91 bp 부근에서 대립 계통과 구별되는 DNA 밴드가 베타글루칸 고함유 계통에서 확인되었으며 random primer인 OP-R03 primer를 이용하여 PCR을 수행했을 때보다 재현성이 높고 진한 DNA 밴드임을 확인할 수 있었다.

본 연구는 큰느타리버섯의 고온적응성 형질에 관련된 SCAR marker를 개발하기 위해 수행되었다. operon 사의 OPA(20개), OPB(20개), OPL(20개), OPP(20개), OPR(20개), OPS(20개) 등 총 120개 primer를 random primer(10 mer)로 사용하여 대립 계통 7종과 고온성 계통 7 종을 대상으로 RAPD를 이용한 bulked segregant analysis를 실시하여 OP-A06 primer로부터 대립 계통에는 나타나지 않고 고온성 계통에만 나타나는 특이적인 RAPD 밴드를 얻었다. OP-A06 primer를 이용한 RAPD 결과, 약 385 bp 부근에서 고온성 계통에 특이적인 DNA 밴드가 관찰되었으며 이 DNA 밴드의 염기서열 말단을 근거로 SCAR 마커로 사용할 specific primer인 OP-A06-1-F와 OP-A06-1-R를 디자인하였다. SCAR 마커 OP-A06-1-F/-1-R primer를 이용하여 PCR을 수행한 결과에서도 385 bp 부근에서 대립 계통과 구별되는 DNA 밴드가 고온성 계통에서 확인되었으며 random primer인 OP-A06 primer를 이용하여 PCR을 수행했을 때보다 재현성이 높고 진한 DNA 밴드임을 확인할 수 있었다.