• Title/Summary/Keyword: mutant protein

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Disturbance of $\alpha$-Amylase Secretion from Bacillus amyloliquefaciens Cells by the Treatment of Puromycin and Magnesium (Bacillus amyloliquefaciens에서 Puromycin 과 Magnesium에 의한 $\alpha$-Amylase 의 분비저해)

  • 안순자;김순옥;이동희;송방호
    • Microbiology and Biotechnology Letters
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    • v.17 no.5
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    • pp.412-420
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    • 1989
  • To know how the ribosomes involved in secretory protein synthesis were attached to the cytoplasmic membrane in Bacillus amyloliquefaciens, the cells were treated with puromycin combinated with magnesium at the logarithmic phase, and the variation of cell-bound and extracellular $\alpha$-amylase activity was assayed for determining the $\alpha$-amylase translocation blocking through the cytoplasmic membrane. In the abnormal $\alpha$-amylase producing mutant in which the C-terminal of the $\alpha$-amylase structure was deleted, B. umytotiquefaciens CH10-2, the $\alpha$-amylase was translocated normally through the cytoplasmic membranes, and the translocation blocking by puromycin was revealed to have a similar pattern as that in the wild type. This means that the C-terminal part of the enzyme structure may not have a signal for secretion. The cell death of the logarithmic phase cells in both strains was not affected much under 20$\mu\textrm{g}$/$m\ell$ of puromycin, however, the $\alpha$-amylase translocation was blocked markedly under less than 10$\mu\textrm{g}$/$m\ell$ of the puromycin concentration. The blocking of the enzyme secretion by puromycin may be due to the detachment of the ribosomes from cytoplasmic membranes by disturbing the nascent polypeptide synthesis. Further evidence for confirming this was that the detachment was increased in 50 mM of magnesium ion because the extracellular $\alpha$-amylase activity was decreased more under this condition. If the cells were treated with trypsin combinated with Iysozyme, the extracellular $\alpha$-amylase activity from the cultured medium was reduced markedly, however, the activity from the cells treated with trypsin only was not reduced. This means that the nascent polypeptides protruding from the cytoplasmic membrane were sensitive to the trypsin digestion, whereas the matured ones were not. Therefore, the protruding polypeptides from the cytoplasmic membranes may be truncated by trypsin before forming their final tertiary structures by folding in the cell wall layer.

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Polymorphisms in Epigenetic and Meat Quality Related Genes in Fourteen Cattle Breeds and Association with Beef Quality and Carcass Traits

  • Liu, Xuan;Usman, Tahir;Wang, Yachun;Wang, Zezhao;Xu, Xianzhou;Wu, Meng;Zhang, Yi;Zhang, Xu;Li, Qiang;Liu, Lin;Shi, Wanhai;Qin, Chunhua;Geng, Fanjun;Wang, Congyong;Tan, Rui;Huang, Xixia;Liu, Airong;Wu, Hongjun;Tan, Shixin;Yu, Ying
    • Asian-Australasian Journal of Animal Sciences
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    • v.28 no.4
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    • pp.467-475
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    • 2015
  • Improvement for carcass traits related to beef quality is the key concern in beef production. Recent reports found that epigenetics mediates the interaction of individuals with environment and nutrition. The present study was designed to analyze the genetic effect of single nucleotide polymorphisms (SNPs) in seven epigenetic-related genes (DNMT1, DNMT3a, DNMT3b, DNMT3L, Ago1, Ago2, and HDAC5) and two meat quality candidate genes (CAPN1 and PRKAG3) on fourteen carcass traits related to beef quality in a Snow Dragon beef population, and also to identify SNPs in a total of fourteen cattle populations. Sixteen SNPs were identified and genotyped in 383 individuals sampled from the 14 cattle breeds, which included 147 samples from the Snow Dragon beef population. Data analysis showed significant association of 8 SNPs within 4 genes related to carcass and/or meat quality traits in the beef populations. SNP1 (13154420A>G) in exon 17 of DNMT1 was significantly associated with rib-eye width and lean meat color score (p<0.05). A novel SNP (SNP4, 76198537A>G) of DNMT3a was significantly associated with six beef quality traits. Those individuals with the wild-type genotype AA of DNMT3a showed an increase in carcass weight, chilled carcass weight, flank thicknesses, chuck short rib thickness, chuck short rib score and in chuck flap weight in contrast to the GG genotype. Five out of six SNPs in DNMT3b gene were significantly associated with three beef quality traits. SNP15 (45219258C>T) in CAPN1 was significantly associated with chuck short rib thickness and lean meat color score (p<0.05). The significant effect of SNP15 on lean meat color score individually and in combination with each of other 14 SNPs qualify this SNP to be used as potential marker for improving the trait. In addition, the frequencies of most wild-type alleles were higher than those of the mutant alleles in the native and foreign cattle breeds. Seven SNPs were identified in the epigenetic-related genes. The SNP15 in CAPN1 could be used as a powerful genetic marker in selection programs for beef quality improvement in the Snow Dragon Beef population.

Biological Functions of N- and O-linked Oligosaccharides of Equine Chorionic Gonadotropin and Lutropin/Chorionic Gonadotropin Receptor

  • Min, K.S.
    • Korean Journal of Animal Reproduction
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    • v.24 no.4
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    • pp.357-364
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    • 2000
  • Members of the glycoprotein family, which includes CG, LH, FSH and TSH, comprise two noncovalently linked $\alpha$- and $\beta$-subunits. Equine chorionic gonadotropin (eCG), known as PMSG, has a number of interesting and unique characteristics since it appears to be a single molecule that possesses both LH- and FSH-like activities in other species than the horse. This dual activity of eCG in heterologous species is of fundamental interest to the study of the structure-function relationships of gonadotropins and their receptors. CG and LH $\beta$ genes are different in primates. In horse, however, a single gene encodes both eCG and eLH $\beta$ -subunits. The subunit mRNA levels seem to be independently regulated and their imbalance may account for differences in the quantities of $\alpha$ - and $\beta$-subunits in the placenta and pituitary. The dual activities of eCG could be separated by removal of the N-linked oligosaccharide on the $\alpha$-subunit Asn 56 or CTP-associated O-linked oligosaccharides. The tethered-eCG was efficiently secreted and showed similar LH-like activity to the dimeric eCG. Interestingly, the FSH-like activity of the tethered-eCG was increased markedly in comparison with the native and wild type eCG. These results also suggest that this molecular can implay particular models of FSH-like activity not LH-like activity in the eCG/indicate that the constructs of tethered molecule will be useful in the study of mutants that affect subunit association and/or secretion. A single-chain analog can also be constructed to include additional hormone-specific bioactive generating potentially efficacious compounds that have only FSH-like activity. The LH/CG receptor (LH/CGR), a membrane glycoprotein that is present on testicular Leydig cells and ovarian theca, granulosa, luteal, and interstitial cells, plays a pivotal role in the regulation of gonadal development and function in males as well as in nonpregnant and pregnant females. The LH/CGR is a member of the family of G protein-coupled receptors and its structure is predicted to of a large extracellular domain connected to a bundle of seven membrane-spanning a-helices. The LH/CGR phosphorylation can be induced with a phorbol ester, but not with a calcium ionophore. The truncated form of LHR also was down-regulated normally in response to hCG stimulation. In contrast, the cell lines expressing LHR-t631 or LHR-628, the two phosphorylation-negative receptor mutant, showed a delay in the early phase of hCG-induced desensitization, a complete loss of PMA-induced desensitization, and an increase in the rate of hCG-induced receptor down-regulation. These results clearly show that residues 632~653 in the C-terminal tail of the LHR are involved in PMA-induced desensitization, hCG-induced desensitization, and hCG-induced down-regulation. Recently, constitutively activating mutations of the receptor have been identified that are associated with familial male-precocious puberty. Cells expressing LHR-D556Y bind hCG with normal affinity, exhibit a 25-fold increase in basal cAMP and respond to hCG with a normal increase in cAMP accumulation. This mutation enhances the internalization of the free and agoinst-occupied receptors ~2- and ~17- fold, respectively. We conclude that the state of activation of the LHR can modulate its basal and/or agonist-stimulated internalization. Since the internalization of hCG is involved in the termination of hCG actions, we suggest that the lack of responsiveness detected in cells expressing LHR-L435R is due to the fast rate of internalization of the bound hCG. This statement is supported by the finding that hCG responsiveness is restored when the cells are lysed and signal transduction is measured in a subcellular fraction (membranes) that cannot internalize the bound hormone.

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Identification of multiple key genes involved in pathogen defense and multi-stress tolerance using microarray and network analysis (Microarray와 Network 분석을 통한 병원균 및 스트레스 저항성 관련 주요 유전자의 대량 발굴)

  • Kim, Hyeongmin;Moon, Suyun;Lee, Jinsu;Bae, Wonsil;Won, Kyungho;Kim, Yoon-Kyeong;Kang, Kwon Kyoo;Ryu, Hojin
    • Journal of Plant Biotechnology
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    • v.43 no.3
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    • pp.347-358
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    • 2016
  • Brassinosteroid (BR), a plant steroid hormone, plays key roles in numerous growth and developmental processes as well as tolerance to both abiotic and biotic stress. To understand the biological networks involved in BR-mediated signaling pathways and stress tolerance, we performed comparative genome-wide transcriptome analysis of a constitutively activated BR bes1-D mutant with an Agilent Arabidopsis $4{\times}44K$ oligo chip. As a result, we newly identified 1,091 (562 up-regulated and 529 down-regulated) significant differentially expressed genes (DEGs). The combination of GO enrichment and protein network analysis revealed that stress-related processes, such as metabolism, development, abiotic/biotic stress, immunity, and defense, were critically linked to BR signaling pathways. Among the identified gene sets, we confirmed more than a 6-fold up-regulation of NB-ARC and FLS2 in bes1-D plants. However, some genes, including TIR1, TSA1 and OCP3, were down-regulated. Consistently, BR-activated plants showed higher tolerance to drought stress and pathogen infection compared to wild-type controls. In this study, we newly developed a useful, comprehensive method for large-scale identification of critical network and gene sets with global transcriptome analysis using a microarray. This study also showed that gain of function in the bes1-D gene can regulate the adaptive response of plants to various stressful conditions.

Effect of Lactic Acid Bacteria and Formic Acid on the Silage Quality of Whole Crop Rice at Different Maturity (유산균 및 개미산 첨가가 수확시기별 벼 사일리지의 발효 품질 및 사료성분에 미치는 영향)

  • 김병완;김곤식;성경일
    • Journal of The Korean Society of Grassland and Forage Science
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    • v.24 no.1
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    • pp.61-70
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    • 2004
  • Silage additives are needed to increase the quality of whole crop rice silage which seldom produce without the additives due to both high pH and butyric acid concentrations. Little information, however, is available about the silage fermentation of whole crop rice added with silage additives in Korea. This study was conducted to determine the optimum levels of silage additives by evaluating the effects of latic acid bacteria (LAB) and formic acid concentrations on the silage quality of whole crop rice harvested at different mature stages. Field study was established early in May until October 7th on a rice field at Yupori, Sinbuk-yeup, Chunchon, Kangwon-Do. "Ilpum" mutant rice was harvested at six different mature stages; booting stage (17 Aug.), milk-ripe stage (27 Aug.), dough stage (7 Sep.), yellow ripe stage (17 Sep.), dead ripe stage (27 Sep.) and full ripe stage (7 Oct.). Each sample was ensiled in three different ways; with 1) LAB (0.05, 0.1 and 0.2% of sample wt), 2) formic acids (0.2, 0.3 and 0.4% of sample wt.) and 3) no additive. The additive levels did not affect dry matter content, crude protein, fiber and total digestable neutriant concentrations at all stages. Addition of additives significantly decreased the silage pH and butyric acid concentrations which tended to be more decreased with higher levels of additives. Latic acid concentrations were higher with the use of additives, especially with LAB. The lower concentrations of ammonia-N were observed in additive treatments at all stages, but the concentrations of ammonia-N did not differ according to the additve levels after yellow ripe stage (0.69, 0.60 and 0.71% of DM in 0.05, 01 and 0.2% of LAB, respectively; 0.64 0.59 and 0.75% of DM in 0.2, 0.3 and 0.4% of formic acid, respectively). These results indicate that the optimum addition levels of LAB and formic acid are 0.5∼0.1% and 0.2∼0.3%, respectively, on which the high quality of rice whole crop silage was produced. produced.

Expression and Purification of Recombinant Human Interferon-gamma Produced by Escherichia coli (대장균이 생산한 재조합 인체 감마인터페론의 발현과 정제)

  • Park, Jung-Ryeol;Kim, Sung-Woo;Kim, Jae-Bum;Jung, Woo-Hyuk;Han, Myung-Wan;Jo, Young-Bae;Jung, Joon-Ki
    • KSBB Journal
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    • v.21 no.3
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    • pp.204-211
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    • 2006
  • For the production of the recombinant human interferon-gamma(rhIFN-${\gamma}$) in Escherichia coli, human glucagon and ferritin heavy chain were used as fusion partners. Even though rhIFN-${\gamma}$ is expressed as an inclusion body form in E. coli because of strong hydrophobicity of itself, over 50% of fused rhIFN-${\gamma}$ was expressed as soluble form in E. coli $Origami^{TM}$(DE3) harboring pT7FH(HE)-IFN-${\gamma}$ which encodes ferritin heavy chain-fused rhIFN-${\gamma}$. In the case of using glucagon-ferritin heavy chain hybrid mutant as a fusion partner, 6X His-tag was additionally introduced to N-terminus of GFHM(HE)-IFN-${\gamma}$ for enhancing purification yields of rhIFN-${\gamma}$. Fusion protein HGFHM(HE)-IFN-${\gamma}$ with two 6X His-tag was more effectively bound to Ni-NTA agarose bead than GFHM(HE)-IFN-${\gamma}$ with a 6X His-tag. rhIFN-${\gamma}$ was completely purified from enterokinase-treated HGFHM(HE)-IFN-${\gamma}$ by Ni-NTA affinity column. For high-level production of rhIFN-${\gamma}$, glucose was used as the sole carbon source with simple exponential feeding rate($2.4{\sim}7.2g/h$) in fed-batch process. The effective lactose concentration for the expression of the rhIFN-${\gamma}$ was $10{\sim}20mM$. Under the fed-batch culture conditions, rhIFN-${\gamma}$ production yield reached 11 g DCW/L for 6 hours after lactose induction.

A New Auricleless Barley Cultivar "Dami" for Whole Crop Forage (밀식적응 양질 다수성 무엽이 청보리 품종 "다미")

  • Park, Tae-Il;Seo, Jae-Hwan;Han, Ouk-Kyu;Park, Ki-Hun;Choi, Jae-Seong;Kim, Jung-Gon;Park, Jong-Chul;Kim, Hong-Sik;Heo, Hwa-Young;Baek, Seong-Bum;Kwon, Young-Up;Park, Hyoung-Ho;Kang, Moon-Seok;Park, Kwang-Geun;Suh, Sae-Jung
    • Korean Journal of Breeding Science
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    • v.41 no.3
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    • pp.349-353
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    • 2009
  • We developed a new barley cultivar "Dami" (Hordeum vulgare L.) with the auricleless gene lig (al, li, aur-a). The characteristic of auricleless is a spontaneous mutant type which has known as a monogenic recessive gene. The plant with the gene has erect leaf blades because of no auricle. The cultivar was derived from a cross between 'BGS60' and 'Kangbori'. 'BGS60' has the auricleless gene (li), while 'Kangbori' showed a high biomass with winter hardiness and resistance to BaYMV (Barley Yellow Mosaic Virus). Subsequent generations were handled by the bulk method in a pedigree selection program. A promising line showed both high yield and lodging resistance in the yield trials at Iksan in 2003 to 2004, and designated as Iksan414. The line was subsequently evaluated for winter hardiness, earliness, and yield in the seven locations around Korea for three years from 2005 to 2007 and was designated as "Dami" and released. It has the growth habit of III, erect plant type, green leaf and stem similar to the check cultivar 'Sunwoo' Its heading date was April 30, and maturing date May 31 in paddy field conditions, which were similar to those of 'Sunwoo' respectively. The cultivar Dami was 97 cm in culm length, had 643 spikes per $m^2$ and higher leaf dry weight, and better adaptability to dense planting, winter hardiness, and resistance to BaYMV than the check cultivar did. The average forage yield of "Dami" was about 12 ton $ha^{-1}$ in dry matter (33 ton $ha^{-1}$ in fresh matter) in paddy field. "Dami" also showed 7.5% of crude protein content, 28.5% of ADF (Acid Detergent Fiber), 50.1% of NDF (Neutral Detergent Fiber), and 66.4% of TDN (Total Digestible Nutrients), including higher grade of silage quality for whole crop barley. This cultivar would be suitable for the area where the daily minimum temperature of January is above $-8^{\circ}C$ in Korean peninsula.

A New Mid-late Maturing Rice Variety, 'Honong' with High-Quality and Multiple Disease Resistance (중만생 고품질 복합내병성 벼 신품종 '호농')

  • Kim, Ki-Young;Shin, Mun-Sik;Ko, Jae-Kwon;Nam, Jeong-Kwon;Ha, Ki-Yong;Baek, Man-Gee;Ko, Jong-Cheol;Kim, Bo-Kyeong;Park, Hyun-Su;Shin, Woon-Chul;Kang, Hyun-Jung;Choung, Jin-Il;Kim, Woo-Jae;Mo, Young-Jun;Kim, Jung-Gon
    • Korean Journal of Breeding Science
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    • v.42 no.3
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    • pp.252-256
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    • 2010
  • 'Honong' is a new japonica rice variety developed and registered by the rice breeding team of Department of Rice and Winter Cereal Crop, NICS, RDA in 2009. This variety was derived from mutagen MNU (N-methyl-N-nitrosourea) treatment on fertilized egg cells of Unbong31. This variety has about 126 days growth duration from transplanting to harvesting in west-southern coast, Honam and Youngnam plain of Korea. It has 79 cm in culm length and is tolerant to lodging. In reaction to biotic and abiotic stresses, it show resistance to blast, bacterial blight pathogen races from $K_1$ to $K_3$ and stripe virus, but susceptible to other major diseases and insect pests. The milled rice of 'Honong' exhibits translucent, relatively clear non-glutinous endosperm and medium short grain. It has higher amylose content (20.3%) and lower protein content (6.3%), and good palatability of cooked rice compared with Nampyeong. The milled rice yield performance of this variety is about 5.44MT/ha in local adaptability test for three years. 'Honong' would be adaptable to west-southern coast, Honam and Youngnam plain of Korea.

An Early-Maturing and High-Biomass Tetraploid Rye (Secale cereale L.) Variety 'Daegokgreen' for Forage Use (조생 다수성 조사료용 4배체 호밀 '대곡그린')

  • Ku, Ja-Hwan;Han, Ouk-kyu;Oh, Young-Jin;Park, Tae-Il;Kim, Dae-Wook;Kim, Byung-Joo;Park, Myoung Ryoul;Ra, Kyung-Yoon
    • Journal of The Korean Society of Grassland and Forage Science
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    • v.40 no.4
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    • pp.209-215
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    • 2020
  • A winter forage tetraploid rye (Secale cereale L.) cultivar, 'Daegokgreen', was developed at the Department of Central Area Crop Science, NICS, RDA in 2016. The mutant line 'CG11003-8-B', which was induced from rye cultivar 'Gogu' (diploid) by colchicine treatment, was selected for its excellent agronomic performance and was placed in preliminary yield trials for one year, 2013. The line was designated "Homil59" and was tested for regional yield trials at the four locations in Korea from 2014 to 2016. Finally, the new cultivar was named as the 'Daegokgreen' (grant number 8274). The leaf of cultivar 'Daegokgreen' is wide, long and dark-green color. The cultivar also has a big-size grain with light-brown color. The heading date of cultivar 'Daegokgreen' was April 17 which was 2 days later than that of check cultivar 'Gogu'. The tolerance to cold and wet injury, and resistance to powdery mildew and leaf rust of the new cultivar were similar to those of the check cultivar but the resistance to the lodging of the new cultivar was stronger than that of the check. The average roughage fresh and dry matter yield of the new cultivar after 10 days from heading were 37.0 and 7.7 MT ha-1, respectively, which were similar to those (38.4 and 8.0 MT ha-1) of the check cultivar. The roughage quality of 'Daegokgreen' was higher in crude protein content (8.9%) than that of the check cultivar (7.9%), while was similar to the check in total digestible nutrients (56.9%). This cultivar is recommended for fall sowing forage crops at all of crop cultivation areas in Korea.

Agricultural Characteristics of Inbred Korean Waxy Corn Lines and Relationships (국내 찰옥수수 계통의 농업형질 특성 및 연관 연구)

  • Jun Young Ha;Young Sam Go;Jae Han Son;Beom Young Son;Tae Wook Jung;Hwan Hee Bae
    • KOREAN JOURNAL OF CROP SCIENCE
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    • v.67 no.4
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    • pp.265-273
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    • 2022
  • Waxy corn (Zea mays L.), which contains homozygous mutant alleles for the waxy1 (wx1) gene, is widely consumed as a snack food in Asia. This study evaluated sixteen agronomic characteristics of inbred Korean waxy corn lines to aid development of high-quality waxy corn cultivars. The plant materials studied were 177 inbred waxy corn lines developed by the National Institute of Crop Science, Rural Development Administration, Republic of Korea. For the tested lines, days to tasseling and silking averaged 77.69±2.22 days (with a range of 56-97 days), and 81.12±7.56 days (66-99 days), respectively. Plant length ranged from 88 to 237 cm (averaged 164.88±22.67 cm), ear length averaged 11.75±2.52 cm (5.0-18.5 cm), and ear width averaged 2.94±0.68 cm (1.4-4.5 cm). The number of rows on each ear of corn averaged 12.22±2.22 (7-32 rows) and the kernel number averaged 24.30±4.22 (9-37 kernels) per row. The crude protein content was 12.05±1.53% (8.90-21.80%) and total starch content was 69.27±5.74% (49.5-83.9%). Principal component analysis revealed that ear width, grain length, ear length, days to tasseling, days to silking, percentage of ear setting height, and total starch are features that allow distinction between the 177 waxy inbred corn lines. Hierarchical cluster analysis identified twelve waxy inbred lines that produce tall plants and have a short silking period. These lines may improve yield among quickly growing corn varieties.